Is butyrate a natural alternative to dexamethasone in the management of CoVID-19?

Coronavirus disease 2019 (CoVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 has affected more than 100 million lives. Severe CoVID-19 infection may lead to acute respiratory distress syndrome and death of the patient, and is associated with hyperinflammation and cytokine storm. The broad spectrum immunosuppressant corticosteroid, dexamethasone, is being used to manage the cytokine storm and hyperinflammation in CoVID-19 patients. However, the extensive use of corticosteroids leads to serious adverse events and disruption of the gut-lung axis. Various micronutrients and probiotic supplementations are known to aid in the reduction of hyperinflammation and restoration of gut microbiota. The attenuation of the deleterious immune response and hyperinflammation could be mediated by short chain fatty acids produced by the gut microbiota. Butyric acid, the most extensively studied short chain fatty acid, is known for its anti-inflammatory properties. Additionally, butyric acid has been shown to ameliorate hyperinflammation and reduce oxidative stress in various pathologies, including respiratory viral infections. In this review, the potential anti-inflammatory effects of butyric acid that aid in cytokine storm depletion, and its usefulness in effective management of critical illness related to CoVID-19 have been discussed.


Introduction
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative factor for the Coronavirus disease 2019 (CoVID-19) 1 . SARS-CoV-2 has affected nearly 100 million people around the globe 2 . SARS-CoV-2 enters the host by binding to its receptor, angiotensin converting enzyme 2 (ACE2), which is expressed mainly in the lungs and intestine [3][4][5] . Upon infection, SARS-CoV-2 causes mild to severe inflammation, which disrupts homeostasis and the integrity of infected organs 6,7 . Furthermore, severe infection of CoVID-19 results in systemic inflammation, thrombosis, acute respiratory distress syndrome (ARDS) and multiple organ failure, which may lead to death 8-10 . The corticosteroid immunosuppressant, dexamethasone, which attenuates hyperinflammation and cytokine storm, is being used to treat seriously ill CoVID-19 patients and has been found to improve survival in hospitalised patients 11,12 . However, the prolonged usage of dexamethasone causes serious adverse effects and gut dysbiosis 13,14 . Besides, the hyperinflammation and thrombotic complications associated with CoVID-19 can also be alleviated by various nutrients including vitamins, polyunsaturated fatty acids, minerals, and even amino acids 15-20 . The growing number of studies indicate the potential role of nutritional supplement, probiotics, and gut microbiome in mitigating the inflammation and in preventing viral infections including respiratory viral infections 21 . Alterations of the gut microbiome has been observed during SARS-CoV-2 infection, which significantly reduces the abundance of beneficial microbiome and its metabolites, such as short chain fatty acids (SCFAs) including butyric acid 22-24 .
Butyric acid or butyrate can act primarily as an anti-inflammatory molecule and various studies have reported its role in mitigating hyperinflammation via several mechanisms [25][26][27] . For the past several years, our group has worked on the role of proinflammatory regulators in the pathogenesis of various inflammatory disorders and identified that the role of histone deacetylase (HDAC) inhibitor in activating anti-inflammatory molecules. Further, this leads to the simultaneous down regulation of proinflammatory membrane receptors, downstream signalling molecules and respective cytokines, resulting in inflammatory homeostasis. Our in vitro preliminary experiments using various cell lines have revealed that the molecular mechanism of butyrate in neutralising inflammatory devastation, induction of anti-inflammatory molecular expression and its translocation to the site of action, is almost similar to dexamethasone 28 . Consequently, we hypothesise if the SCFA, butyric acid, a HDAC inhibitor, which is synthesized by the gut microbiota, could have strong anti-inflammatory functions with anti-fibrotic properties. Therefore, this article reviews the anti-inflammatory properties of butyric acid or butyrate and its associated molecular pathways involved in controlling the cytokine storm and hyperinflammation associated with SARS-CoV-2 infection. Based on the various positive reports, we presume that butyric acid possesses potent anti-inflammatory activity, which suggests it as an alternative to dexamethasone for the preventive management of primary and secondary complications related to CoVID-19.

Coronavirus disease 2019
The CoVID-19 pandemic is caused by SARS-Cov-2, which belongs to genera β-coronaviruses and is the seventh known coronavirus to infect humans 4 . Spike protein of SARS-CoV-2 binds to ACE2, a type I membrane protein 29 expressed in the lung, heart, kidney, and intestine 3,30,31 . The majority of SARS-CoV-2 infected cases present with mild symptoms like dry cough, sore throat, fatigue and fever 9,10,32 . Less common symptoms such as myalgia, expectoration, pharyngalgia, dizziness, nausea, headache, haemoptysis, diarrhoea, abdominal pain, and vomiting have also been reported. Lymphocytopenia along with elevated expression of C-reactive proteins (CRP) and inflammatory cytokines are also common 9,10,32,33 . Infection can progress into severe disease with dyspnoea, grinding glass-like abnormalities and patchy consolidation areas in lungs observed upon imaging; viral pneumonia usually appears after 2-3 weeks of infection 9,10,32,33 . However, some patients have developed organ failure, septic shock, myocarditis, acute cardiac injury, arrhythmia, pulmonary oedema, severe pneumonia, acute kidney injury, and ARDS. Inflammation, oxidative stress, and fibrosis associated with CoVID-19 is perhaps partially mediated by angiotensin II (AngII), a substrate for ACE2, which degrades it to anti-inflammatory angiotensin (1-7). The accumulation of AngII results in hyperinflammation induced by nucleotide-binding oligomerization domain (NOD) like receptors family pyrin domain-containing 3 (NLRP3) inflammasome and nuclear factor kappa B (NF-κB) activation 34 . Disrupted immune response in CoVID-19 is further characterized by decreased expression of human leukocyte antigen D related (HLA-DR) on CD(cluster of differentiation)14 monocytes, accompanied by decrease in number of CD4 and CD19 lymphocytes, and natural killer cells along with the continuous production of proinflammatory tumour necrosis factor (TNF)-α and interleukin (IL)-6 secreted by circulating monocytes, subsequently leading to cytokine storm and hyperinflammation 7 . Hypercytokinemia and hyperinflammation associated with CoVID-19 results in acute lung injury, ARDS and death of the patients 8,35,36 . Furthermore, the SARS-CoV-2 infection may lead to liver injury and its dysfunction, and dysbiosis in the gut, where high expression of ACE2 is observed. Myocardial damage by interaction of SARS-CoV-2 with ACE2 expressed in cardiac pericytes has also been observed 37 . In addition, the high complication of disseminated intravascular coagulation is known to be associated with the severe form of CoVID-19 38 .

Nutrients in mitigating the Covid-19 pathogenesis
Nutrition and nutrients play a vital role in enhancing immune response along with reduction of inflammation and oxidative stress 44-46 . Better nutritional status of CoVID-19 patients is associated with less adverse outcomes 18,47-52 . Vitamin D is involved in reducing respiratory infections, such as influenza, and a reduced plasma 25-hydroxyvitamin D (25(OH)D) concentration in SARS-CoV-2 patients has been observed 53 . Moreover, people with vitamin D deficiency are at higher risk of getting infected with SARS-CoV-2 54,55 . Co-supplementation of vitamin D along with glutathione precursor L-cysteine significantly increases serum 25(OH)D levels and augments vitamin D regulatory gene expression, which in turn reduces the oxidative stress and inflammatory responses in CoVID-19 patients 56 . Vitamin D supplementation in SARS-CoV-2 infected patients attenuates the production of proinflammatory cytokines like Interferon (IFN)-γ, IL-6, IL-2 and TNF-α by inhibiting NF-κB and other pathways 57-59 . CoVID-19 associated inflammatory signalling pathways including NF-κB, Mitogen-Activated Protein Kinase (MAPK) and phosphatidylinositol 3-kinase/ protein kinase B (PI3K/ AKT) and innate immune response pathways, such as Toll-like signalling and NOD-like signalling modulation and regulation can be mediated by the combination of curcumin, vitamin C, and glycyrrhizic acid 60 . Vitamin C has been known to improve the immune condition by enhancing differentiation and proliferation of B-and T-cells, but severe vitamin C deficiency is associated with pneumonia and respiratory tract infections 61 . Intravenous administration of vitamin C can significantly decrease IL-6 levels 62,63 . Glycyrrhizic acid and curcumin exhibits anti-viral, anti-inflammation, anti-cancer, and immune system benefits 60 . The combination of vitamin D/magnesium/vitamin B12 significantly reduced the subsequent need for oxygen therapy and/or intensive care support in older CoVID-19 patients 57 . Vitamin B12 is crucial in maintaining the healthy gut microbiome which plays a vital role in immune responses 57 . Fat soluble vitamin E acts as an antioxidant that scavenges Reactive Oxygen species (ROS) and inhibits devastating effects of hyperinflammation 64 . Moreover, the supplementation of vitamin E stimulates T cell function and confers protection against upper respiratory infections 65 .
Selenium is one of the key micronutrients known to positively impact CoVID-19 patient recovery 66,67 . Selenium status regulates the expression of glutathione peroxidase 1 (GPX1), a cytosolic selenoenzyme known for its antioxidative properties. The antioxidant enzyme GPX1 mitigates the production of ROS and further leading to mutations in the viral genome 68 . In addition, attenuating ROS also helps in the inhibition of proinflammatory NF-κB activation and further nuclear translocation 69 . Severe endothelial injury and widespread pulmonary micro thromboses are accompanied with platelet activation and aggregation in patients with severe CoVID-19 manifestations. The synthetic Rupatadine (histamine1 receptor antagonist) and natural flavonoids with anti-inflammatory properties are known to inhibit the platelet activating factor 70 . Elderly individuals with deficiency of nutrients, such as vitamin C, vitamin D, calcium, folate, and zinc are prone to increase severity of SARS-CoV-2 infection 71 . Folic acid may inhibit furin protease and inactivates chymotrypsin-like protease (3CL pro ) 72 . Zinc (Zn 2+ ) deficiency contributes to impaired cell mediated immune response and increased susceptibility to various infections. However, increased intracellular levels of Zn 2+ disrupt viral RNA replication including SARS-CoV-2, where Zn 2+ inhibits RNA (Ribonucleic acid) dependent RNA polymerase (RdRp) elongation and template binding 73 . Among CoVID-19 patients, iron deficiency is strongly associated with increased inflammation and longer stay in hospitals 74 .
Health beneficial compounds, including minerals, antioxidants, phytochemicals, vitamins, and minerals present in fruits and vegetables, can exert antioxidative, anti-inflammatory and antiviral effects during various non-infectious and infectious disease 71 . Alliin, an S-allyl cysteine sulfoxide compound present in garlic has shown to have inhibitory action on 3CL pro , a protease that plays a vital role in SARS-CoV-2 replication 75 . Salvianolic acid A and curcumin have the potential to bind to 3CL pro with greater affinity 76 . Resveratrol acts as an anti-inflammatory molecule that inhibits the NFκB pathway and thereby reduces circulatory cytokines, such as IL-6 and TNF-α levels, which are observed in severe SARS-CoV-2 infection 77 . Sea cucumber (Stichopus japonicus) derived sulphated polysaccharide showed significant anti-viral activity against SARS-CoV-2 infection 78 . Omega-3 polyunsaturated fatty acids, including eicosapentaenoic acid and docosahexaenoic acid have been shown to exhibit anti-inflammatory effects by downregulation of the NF-κB pathway 71,79,80 . Free fatty acids such as oleic acid, arachidonic acid and linoleic acid have shown antiviral activity at micromolar concentrations 81 . Dietary fibre intake alters the intestinal microflora and enhances relative proportion of SCFAs, which exhibit anti-inflammatory properties through fatty acid receptors like G-protein-coupled receptor (GPCR) 41 and 43 82-84 .

Probiotics: suppressors of respiratory tract infections and inflammation
Probiotics are living microorganisms that provide health benefits to the host upon administration at appropriate doses 85 . Probiotics exert a wide range of beneficial effects such as host microbiome balancing, stimulation of immune system, enhancement of intestinal barrier function or inhibiting pathogens by direct interactions 40,46,86,87 (Table 1)

Is butyrate an alternative to dexamethasone?
Dexamethasone is a synthetic corticosteroid that acts as an anti-inflammatory agent, widely affecting innate and acquired immune system via glucocorticoid receptor 118,119 . Low dose dexamethasone treatment significantly supresses neutrophil infiltration and subsequent pulmonary inflammation and significantly improves lung function in early phase of ARDS 120 . Lower respiratory tract transcriptomic profiling of patients with CoVID-19 associated ARDS shows dysregulated immunoregulation and inflammation. This dysregulated immune response can be modulated by dexamethasone 121 . A short course of dexamethasone significantly reduces CRP levels and accelerates recovery 122 . Dexamethasone treatment in CoVID-19 patients who were receiving mechanical ventilation support results in lower mortality rate 123 . Severe CoVID-19 cases have been brought to remission state after 6 mg once a day intravenous administration of dexamethasone 124 .
Dexamethasone is indicated as a therapeutic option for immune thrombocytopenic purpura associated with CoVID-19 124,125 . Administration of dexamethasone before 30 hours of ARDS onset can significantly reduce the period of mechanical ventilation and mortality 12 . Dexamethasone provides an excellent protective effect against hypoxia associated with CoVID-19 118 . Intravenous dexamethasone treatment for CoVID-19 patients along with standard care significantly decreases the number of ventilator dependent days over 28 days 11 . High dose pulse therapy of dexamethasone increased the survival rate in CoVID-19 patients presented with hyperinflammation 126 . However, dexamethasone, a broad spectrum immunosuppressant, inhibits lymphocytes function and prevents macrophage mediated removal of apoptotic cells, which leads to reduced viral shedding and increases subsequent viremia in mild to moderately ill CoVID-19 patients 124,127,128 . Prolonged use of corticosteroids is associated with serious adverse effects such as short-term hyperglycaemia, cataracts, glaucoma, hypertension, psychological effects, weight gain, increased risk of secondary infections and osteoporosis 13,129 . Use of such corticosteroids may induce gut dysbiosis 14 .
Intestinal microflora widely affects host health and alterations in the gut microbiome is correlated with several disease including respiratory disease 130 . Commensal gut microbiome and its metabolites can modulate host immunity and can also impact on pro inflammatory and immune-regulatory response 131 . Increased production of microbiome metabolite SCFAs may improve health condition 132 . Depletion of SCFA production makes mice more susceptible for allergic lung inflammation. Biological effects exerted by SCFAs is dependent mainly on two mechanisms: SCFA mediated (i) activation of GPCRs and (ii) inhibition of HDAC. SCFAs, via HDAC inhibition, positively impacts the functions and numbers of T-helper 1 cells, T-regulatory cells, and Th17 effector cells resulting in reduced inflammatory response in airway diseases 130 . The short chain fatty acid, butyrate or butyric acid is produced in the colon by anaerobic bacteria such as Roseburia intestinalis, Faecalibacterium prausnitzii, Clostridium butyricum, Megasphaera elsdenii, Mitsuokella multiacida, Eubacterium spp., Fusobacterium spp., Butyrivibrio spp. and Eubacterium hallii 133 . Butyrate concentration in the colon can reach from 10 to 20 mM and serves as major source of energy for colonocytes. Sodium butyrate supplementation enhances the abundance of beneficial bacteria such as Coprococcus, Lachnospiraceae, Ruminococcus, Bifidobacteriaceae and Actinobacteria improving intestinal barrier integrity in obese mice 134 .
Primarily, butyric acid exhibits anti-inflammatory and tissue protective function in the large intestine 135 . Butyric acid is a potential inhibitor of pro-inflammatory molecule NF-κB 135-137 ( Figure 1). Tight junction protein expression in intestinal epithelial cells is also influenced by butyrate mediated regulation 138   Free fatty acid receptors 2/3 and GPCR 109A or by directly binding to HDAC active sites. Inhibition of NF-κB activity by butyrate attenuates inflammation and oxidative stress associated with various pathologies including CoVID-19. Butyrate also activates the transcription factor B lymphocyte-induced maturation protein-1 (BLIMP-1) and enhances the production of anti-inflammatory cytokines.
Butyrate also reduces the infiltration of proinflammatory Th9 cells and eosinophils into lungs 146 . Mice treated with butyrate exhibited a significant reduction of inflammatory infiltrates in the airways, tissue, and vascular disruption, and subsequently less haemorrhaging in the lungs induced by influenza infection 82 . HDAC inhibitor sodium butyrate can suppress ACE2 expression in gut epithelial cells which can help in reducing gastrointestinal symptoms associated with CoVID-19 147 .
Pancreatitis and associated fibrosis induced by L-Arginine can be attenuated by sodium butyrate, which reduces collagen deposition and nitric oxide along with inhibition of profibrotic pancreatic stellate cells 148 . Butyric acid ameliorates bleomycin induced pulmonary fibrosis by attenuating leukocytes infiltration, oxidative stress and NF-κB activation 149 .
Consequently, based on the evidence presented, the potential anti-inflammatory and tissue protective effects of butyric acid on lungs and gut, along with its ability to modulate gut microbiome diversity, enhancing production of endogenous butyric acid could be a better preventive approach to manage CoVID-19 over dexamethasone (Figure 2). However, there is a need Figure 2. A. SARS-CoV-2 transmitted through aerosols reach the lungs via respiratory tract and enters the host cell by binding to its receptor, ACE2 present on the surface of pneumocytes. Followed by endosome mediated internalization, SARS-CoV-2 causes cell injury and subsequent hyperinflammation and cytokine storm, resulting in fibrosis of lungs. These cytokines reach the gut via blood and lymphatic vessels that instigates local inflammation in gut, ushering to leaky gut and gut dysbiosis, resulting in diarrhoea and malabsorption together with reduced production of short chain fatty acids. B. Dexamethasone a synthetic broad-spectrum immunosuppressant can inhibit cytokine storm associated with CoVID-19. As an alternative, oral administration of probiotics or gut microbiome metabolite, SCFAs may ameliorate gut inflammation, restore gut integrity, and gut microbiome. This enhances the production of endogenous SCFAs and reaches the lungs via blood and lymphatic vessels, and may inhibit hyperinflammation and cytokine storm along with induction of anti-inflammatory cytokines production which recovers the lung from injury and the acute respiratory distresses associated with CoVID-19.
for more detailed studies and clinical trials to determine the potency and long-term effect of butyric acid in the preventive management of seriously ill CoVID-19 patients.

Conclusion
Seriously ill CoVID-19 patients are succumbing to respiratory distress syndrome due to significant hyperinflammation and cytokine storm. A broad-spectrum immunosuppressant, dexamethasone, is widely used to treat such cases. However, the prolonged use of this corticosteroid leads to severe adverse events and disrupted immune responses. There are growing number of advanced research studies in search of an alternative to dexamethasone for the better management of critical CoVID-19 patients. Hence, this review extensively searched for evidence to show the anti-inflammatory properties of butyric acid or butyrate and its associated molecular pathways involved in preventing SARS-CoV-2 infected patients from cytokine storm and hyperinflammation. It has been observed that the SARS-CoV-2 infection significantly decreases butyric acid producing bacteria in the host gut. Further, previous research shows that a histone deacetylase inhibitor, butyric acid has proven to be antiinflammatory in lung inflammation including inflammation associated with respiratory viral infection. Therefore, based on the various positive reports, we presume that butyric acid possesses potent anti-inflammatory activity, making it a suitable alternative candidate for the preventive management of primary and secondary complications related to CoVID-19.

Data availabilty
No data is associated with this article.

Open Peer Review
Authors mentioned the usefulness of probiotics in management of tissue inflammation, some more clear idea on mechanism to mitigate the inflammation, particularly the viral load is demanded.
Chemistry of butyrate and dexamethasone and its structure activity relationship in preventing the cytokine storm is necessary.

Yes
Some of the molecules like PGE2, ROS-IKK pathway has not been described in Figure 1 legend.