Precision nutrition to reset virus-induced human metabolic reprogramming and dysregulation (HMRD) in long-COVID.

SARS-CoV-2, the etiological agent of COVID-19, is devoid of any metabolic capacity; therefore, it is critical for the viral pathogen to hijack host cellular metabolic machinery for its replication and propagation. This single-stranded RNA virus with a 29.9 kb genome encodes 14 open reading frames (ORFs) and initiates a plethora of virus-host protein-protein interactions in the human body. These extensive viral protein interactions with host-specific cellular targets could trigger severe human metabolic reprogramming/dysregulation (HMRD), a rewiring of sugar-, amino acid-, lipid-, and nucleotide-metabolism(s), as well as altered or impaired bioenergetics, immune dysfunction, and redox imbalance in the body. In the infectious process, the viral pathogen hijacks two major human receptors, angiotensin-converting enzyme (ACE)-2 and/or neuropilin (NRP)-1, for initial adhesion to cell surface; then utilizes two major host proteases, TMPRSS2 and/or furin, to gain cellular entry; and finally employs an endosomal enzyme, cathepsin L (CTSL) for fusogenic release of its viral genome. The virus-induced HMRD results in 5 possible infectious outcomes: asymptomatic, mild, moderate, severe to fatal episodes; while the symptomatic acute COVID-19 condition could manifest into 3 clinical phases: (i) hypoxia and hypoxemia (Warburg effect), (ii) hyperferritinemia ('cytokine storm'), and (iii) thrombocytosis (coagulopathy). The mean incubation period for COVID-19 onset was estimated to be 5.1 days, and most cases develop symptoms after 14 days. The mean viral clearance times were 24, 30, and 39 days for acute, severe, and ICU-admitted COVID-19 patients, respectively. However, about 25-70% of virus-free COVID-19 survivors continue to sustain virus-induced HMRD and exhibit a wide range of symptoms that are persistent, exacerbated, or new 'onset' clinical incidents, collectively termed as post-acute sequelae of COVID-19 (PASC) or long COVID. PASC patients experience several debilitating clinical condition(s) with >200 different and overlapping symptoms that may last for weeks to months. Chronic PASC is a cumulative outcome of at least 10 different HMRD-related pathophysiological mechanisms involving both virus-derived virulence factors and a multitude of innate host responses. Based on HMRD and virus-free clinical impairments of different human organs/systems, PASC patients can be categorized into 4 different clusters or sub-phenotypes: sub-phenotype-1 (33.8%) with cardiac and renal manifestations; sub-phenotype-2 (32.8%) with respiratory, sleep and anxiety disorders; sub-phenotype-3 (23.4%) with skeleto-muscular and nervous disorders; and sub-phenotype-4 (10.1%) with digestive and pulmonary dysfunctions. This narrative review elucidates the effects of viral hijack on host cellular machinery during SARS-CoV-2 infection, ensuing detrimental effect(s) of virus-induced HMRD on human metabolism, consequential symptomatic clinical implications, and damage to multiple organ systems; as well as chronic pathophysiological sequelae in virus-free PASC patients. We have also provided a few evidence-based, human randomized controlled trial (RCT)-tested, precision nutrients to reset HMRD for health recovery of PASC patients.

Phytonutrient Inhibitors of SARS-CoV-2/NSP5-Encoded Main Protease (Mpro) Autocleavage Enzyme Critical for COVID-19 Pathogenesis.

The genomic reshuffling, mutagenicity, and high transmission rate of the SARS-CoV-2 pathogen highlights an urgent need for effective antiviral interventions for COVID-19 control. Targeting the highly conserved viral genes and/or gene-encoded viral proteins such as main proteinase (Mpro), RNA-dependent RNA polymerase (RdRp) and helicases are plausible antiviral approaches to prevent replication and propagation of the SARS-CoV-2 infection. Coronaviruses (CoVs) are prone to extensive mutagenesis; however, any genetic alteration to its highly conserved Mpro enzyme is often detrimental to the viral pathogen. Therefore, inhibitors that target the Mpro enzyme could reduce the risk of mutation-mediated drug resistance and provide effective antiviral protection. Several existing antiviral drugs and dietary bioactives are currently repurposed to treat COVID-19. Dietary bioactives from three ayurvedic medicinal herbs, 18 ß-glycyrrhetinic acid (ΔG = 8.86 kcal/mol), Solanocapsine (ΔG = 8.59 kcal/mol), and Vasicoline (ΔG = 7.34 kcal/mol), showed high-affinity binding to Mpro enzyme than the native N3 inhibitor (ΔG = 5.41 kcal/mol). Flavonoids strongly inhibited SARS-CoV-2 Mpro with comparable or higher potency than the antiviral drug, remdesivir. Several tannin hydrolysates avidly bound to the receptor-binding domain and catalytic dyad (His41 and Cys145) of SARS-CoV-2 Mpro through H-bonding forces. Quercetin binding to Mpro altered the thermostability of the viral protein through redox-based mechanism and inhibited the viral enzymatic activity. Interaction of quercetin-derivatives with the Mpro seem to be influenced by the 7-OH group and the acetoxylation of sugar moiety on the ligand molecule. Based on pharmacokinetic and ADMET profiles, several phytonutrients could serve as a promising redox nutraceutical for COVID-19 management.

Plant-Derived Natural Non-Nucleoside Analog Inhibitors (NNAIs) against RNA-Dependent RNA Polymerase Complex (nsp7/nsp8/nsp12) of SARS-CoV-2.

The emergence of fast-spreading SARS-CoV-2 mutants has sparked a new phase of COVID-19 pandemic. There is a dire necessity for antivirals targeting highly conserved genomic domains on SARS-CoV-2 that are less prone to mutation. The nsp12, also known as the RNA-dependent RNA-polymerase (RdRp), the core component of 'SARS-CoV-2 replication-transcription complex', is a potential well-conserved druggable antiviral target. Several FDA-approved RdRp 'nucleotide analog inhibitors (NAIs)' such as remdesivir, have been repurposed to treat COVID-19 infections. The NAIs target RdRp protein translation and competitively block the nucleotide insertion into the RNA chain, resulting in the inhibition of viral replication. However, the replication proofreading function of nsp14-ExoN could provide resistance to SARS-CoV-2 against many NAIs. Conversely, the 'non-nucleoside analog inhibitors (NNAIs)' bind to allosteric sites on viral polymerase surface, change the redox state; thereby, exert antiviral activity by altering interactions between the enzyme substrate and active core catalytic site of the RdRp. NNAIs neither require metabolic activation (unlike NAIs) nor compete with intracellular pool of nucleotide triphosphates (NTPs) for anti-RdRp activity. The NNAIs from phytonutrient origin are potential antiviral candidates compared to their synthetic counterparts. Several in-silico studies reported the antiviral spectrum of natural phytonutrient-NNAIs such as Suramin, Silibinin (flavonolignan), Theaflavin (tea polyphenol), Baicalein (5,6,7-trihydroxyflavone), Corilagin (gallotannin), Hesperidin (citrus bioflavonoid), Lycorine (pyrrolidine alkaloid), with superior redox characteristics (free binding energy, hydrogen-bonds, etc.) than antiviral drugs (i.e. remdesivir, favipiravir). These phytonutrient-NNAIs also exert anti-inflammatory, antioxidant, immunomodulatory and cardioprotective functions, with multifunctional therapeutic benefits in the clinical management of COVID-19.

Lactoferrin for Mental Health: Neuro-Redox Regulation and Neuroprotective Effects across the Blood-Brain Barrier with Special Reference to Neuro-COVID-19.

Overall mental health depends in part on the blood-brain barrier, which regulates nutrient transfer in-and-out of the brain and its central nervous system. Lactoferrin, an innate metal-transport protein, synthesized in the substantia nigra, particularly in dopaminergic neurons and activated microglia is vital for brain physiology. Lactoferrin rapidly crosses the blood-brain barrier via receptor-mediated transcytosis and accumulates in the brain capillary endothelial cells. Lactoferrin receptors are additionally present on glioma cells, brain micro-vessels, and neurons. As a regulator of neuro-redox, microglial lactoferrin is critical for protection/repair of neurons and healthy brain function. Iron imbalance and oxidative stress are common among patients with neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease, dementia, depression, and multiple sclerosis. As an endogenous iron-chelator, lactoferrin prevents iron accumulation and dopamine depletion in Parkinson's disease patients. Oral lactoferrin supplementation could modulate the p-Akt/PTEN pathway, reduce Aß deposition, and ameliorate cognitive decline in Alzheimer's disease. Novel lactoferrin-based nano-therapeutics have emerged as effective drug-delivery systems for clinical management of neurodegenerative disorders. Recent emergence of the Coronavirus disease-2019 (COVID-19) pandemic, initially considered a respiratory illness, demonstrated a broader virulence spectrum with the ability to cross the blood-brain barrier and inflict a plethora of neuropathological manifestations in the brain - the Neuro-COVID-19. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections are widely reported in Parkinson's disease, Alzheimer's disease, dementia, and multiple sclerosis patients with aggravated clinical outcomes. Lactoferrin, credited with several neuroprotective benefits in the brain could serve as a potential adjuvant in the clinical management of Neuro-COVID-19.

SARS-CoV-2 Infection Dysregulates Host Iron (Fe)-Redox Homeostasis (Fe-R-H): Role of Fe-Redox Regulators, Ferroptosis Inhibitors, Anticoagulants, and Iron-Chelators in COVID-19 Control.

Severe imbalance in iron metabolism among SARS-CoV-2 infected patients is prominent in every symptomatic (mild, moderate to severe) clinical phase of COVID-19. Phase-I - Hypoxia correlates with reduced O2 transport by erythrocytes, overexpression of HIF-1α, altered mitochondrial bioenergetics with host metabolic reprogramming (HMR). Phase-II - Hyperferritinemia results from an increased iron overload, which triggers a fulminant proinflammatory response - the acute cytokine release syndrome (CRS). Elevated cytokine levels (i.e. IL6, TNFα and CRP) strongly correlates with altered ferritin/TF ratios in COVID-19 patients. Phase-III - Thromboembolism is consequential to erythrocyte dysfunction with heme release, increased prothrombin time and elevated D-dimers, cumulatively linked to severe coagulopathies with life-threatening outcomes such as ARDS, and multi-organ failure. Taken together, Fe-R-H dysregulation is implicated in every symptomatic phase of COVID-19. Fe-R-H regulators such as lactoferrin (LF), hemoxygenase-1 (HO-1), erythropoietin (EPO) and hepcidin modulators are innate bio-replenishments that sequester iron, neutralize iron-mediated free radicals, reduce oxidative stress, and improve host defense by optimizing iron metabolism. Due to its pivotal role in 'cytokine storm', ferroptosis is a potential intervention target. Ferroptosis inhibitors such as ferrostatin-1, liproxstatin-1, quercetin, and melatonin could prevent mitochondrial lipid peroxidation, up-regulate antioxidant/GSH levels and abrogate iron overload-induced apoptosis through activation of Nrf2 and HO-1 signaling pathways. Iron chelators such as heparin, deferoxamine, caffeic acid, curcumin, α-lipoic acid, and phytic acid could protect against ferroptosis and restore mitochondrial function, iron-redox potential, and rebalance Fe-R-H status. Therefore, Fe-R-H restoration is a host biomarker-driven potential combat strategy for an effective clinical and post-recovery management of COVID-19.

COVID-19 during Pregnancy and Postpartum.

Coronavirus Disease 2019 (COVID-19) triggered by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) infection has been declared a pandemic by the World Health Organization (WHO) on March 11, 2020. Oxidative stress and its related metabolic syndromes are potential risk factors in the susceptibility to, and severity of COVID-19. In concert with the earliest reports of COVID-19, obstetricians started to diagnose and treat SARS-CoV-2 infections during pregnancy ("COVID-19-Pregnancy"). High metabolic demand to sustain normal fetal development increases the burden of oxidative stress in pregnancy. Intracellular redox changes intertwined with acute phase responses at the maternal-fetal interface could amplify during pregnancy. Interestingly, mother-to-fetus transmission of SARS-CoV-2 has not been detected in most of the COVID-19-Pregnancy cases. This relative absence of vertical transmission may be related to the presence of lactoferrin in the placenta, amniotic fluid, and lacteal secretions. However, the cytokine-storm induced during COVID-19-Pregnancy may cause severe inflammatory damage to the fetus, and if uncontrolled, may later result in autism spectrum-like disorders and brain development abnormalities in neonates. Considering this serious health threat to child growth and development, the prevention of COVID-19 during pregnancy should be considered a high priority. This review summarizes the intricate virulence factors of COVID-19 and elucidate its pathobiological spectrum during pregnancy and postpartum periods with a focus on the putative and complex roles of endogenous and exogenous lactoferrin in conferring immunological advantage to the host.

COVID-19 during Pregnancy and Postpartum.

As the COVID-19 pandemic intensified the global health crisis, the containment of SARS-CoV-2 infection in pregnancies, and the inherent risk of vertical transmission of virus from mother-to-fetus (or neonate) poses a major concern. Most COVID-19-Pregnancy patients showed mild to moderate COVID-19 pneumonia with no pregnancy loss and no congenital transmission of the virus; however, an increase in hypoxia-induced preterm deliveries was apparent. Also, the breastmilk of several mothers with COVID-19 tested negative for the virus. Taken together, the natural barrier function during pregnancy and postpartum seems to deter the SARS-CoV-2 transmission from mother-to-child. This clinical observation warrants to explore the maternal-fetal interface and identify the innate defense factors for prevention and control of COVID-19-Pregnancy. Lactoferrin (LF) is a potent antiviral iron-binding protein present in the maternal-fetal interface. In concert with immune co-factors, maternal-LF modulates chemokine release and lymphocyte migration and amplify host defense during pregnancy. LF levels during pregnancy may resolve hypertension via down-regulation of ACE2; consequently, may limit the membrane receptor access to SARS-CoV-2 for cellular entry. Furthermore, an LF-derived peptide (LRPVAA) has been shown to block ACE receptor activity in vitro. LF may also reduce viral docking and entry into host cells and limit the early phase of COVID-19 infection. An in-depth understanding of LF and other soluble mammalian milk-derived innate antiviral factors may provide insights to reduce co-morbidities and vertical transmission of SARS-CoV-2 infection and may lead to the development of effective nutraceutical supplements.

Inflammatory responses improve with milk ribonuclease-enriched lactoferrin supplementation in postmenopausal women.

OBJECTIVE AND DESIGN: A 6-month, randomized clinical study was conducted to evaluate the effect of a ribonuclease-enriched lactoferrin (R-ELF) supplement on the circulating cytokine levels and bone health of postmenopausal women. SUBJECTS: Thirty-eight healthy postmenopausal women, aged 45-60 years, were randomized into placebo and R-ELF groups. TREATMENT: The R-ELF group was supplemented with R-ELF (2 × 125 mg/day) and calcium (100% RDA), while the placebo group received only the calcium supplement. METHODS: Serum levels of receptor activator for NF-κB ligand (RANKL), C-reactive protein (CRP) and various pro- and anti-inflammatory cytokines were determined by ELISA. RESULTS: Pro-inflammatory cytokines IL-6 and TNF-α decreased significantly (-44 and -10%, respectively) while anti-inflammatory IL-10 increased (140%) due to R-ELF supplementation at the end of study. RANKL and CRP were modestly reduced (-50%) relative to their placebo levels, although RANKL elevated initially. CONCLUSIONS: R-ELF supplementation showed beneficial effects towards improvement of inflammatory status in postmenopausal women.

Activated lactoferrin and fluconazole synergism against Candida albicans and Candida glabrata vaginal isolates.

OBJECTIVE: To evaluate the fungistatic activity of activated lactoferrin (ALF), fluconazole (FCN) individually and in combination against Candida vaginal isolates as well as to measure the time to recovery from the fungistatic effects after exposure in vitro to threshold minimal inhibitory concentrations (MIC). STUDY DESIGN: Fungistasis patterns for ALF (2.5 mg/mL) and FCN (0.25 mg/mL) were tested at threshold MIC against vaginal isolates of C albicans (n = 5) and C glabrata (n = 5) grown in Sabouraud's dextrose broth against 10(5) yeast inoculum at 37 degrees C for 48 hours by microscale optical density (OD) assay according to the following criteria: "Total stasis" indicates that an agent elicited no change or a change in turbidity <0.1 OD unit for >48 hours (complete growth inhibition), "stasis recovery" (SR) is the time point at which turbidity of a previous stasis system shows an upward growth trend for >0.1 OD unit (recovery from growth inhibition), and "partial stasis" (PS) is proliferation after stasis recovery, measured as a percentage relative to growth control at any time (incomplete growth inhibition). RESULTS: For ALF (2.5 mg/mL), the mean SR time was 15.6 +/- 2 hours for C albicans (n = 5) and 27.5 +/- 2 hours for C glabrata (n = 5). The SR patterns for FCN were strain dependent and showed a wide range of deviation for both Candida species; accordingly, the values were 15.8 +/- 9 hours for C albicans and 25.5 +/- 12 hours for C glabrata. After 48 hours exposure to C albicans, ALF and FCN elicited a mean PS of 27.5 +/- 2% and 24.8 +/- 7%, respectively. The PS values at 48 hours showed a marked variation between C glabrata isolates, 29.1 +/- 24% for ALF and 21.5 +/- 38% for FCN. However, a combination of ALF and FCN at their threshold MIC showed significant drug synergism, causing total stasis of both species of Candida isolates. Thus, no SR for any Candida isolate was detected at or beyond 48 hours. Conversely, native lactoferrin failed to demonstrate such potent synergism with FCN against either Candida species. CONCLUSION: The combination of ALF and FCN at the threshold MIC elicited potent synergism, leading to total fungistasis of C albicans and C glabrata vaginal pathogens. ALF is a new class of fungistatic agent with a mode of action distinct from that of azoles.

Activated lactoferrin's ability to inhibit Candida growth and block yeast adhesion to the vaginal epithelial monolayer.

OBJECTIVE: To study in vitro growth-inhibitory effects of activated lactoferrin (ALF) against vaginal isolates of Candida species and to measure the ability of ALF to block interactions of Candida albicans and Candida glabrata to the vaginal epithelial (VE) monolayer. STUDY DESIGN: In vitro effects of ALF on growth of C albicans and C glabrata in Sabouraud dextrose (SD) broth were measured as change in broth turbidity by microscale optical density assay. ALF was tested at 5 and 2.5 mg/mL concentrations against 105 yeast cell inoculum at 370 degrees C for 96 hours and compared with native lactoferrin and control (growth in broth without ALF). VE cells were isolated from human vaginal tissue biopsies to establish a functional monolayer for yeast interaction studies. ALF effects on Candida interactions with the VE monolayer were tested using 3H-thymidine-labeled yeast. Prophylactic (treatment prior to yeast inoculation onto VE) and therapeutic (treatment to detach VE-adherent yeast) potential of ALF (5 mg/mL) was evaluated against vaginal isolates of C albicans strain NTRL809A and C glabrata strain NTRL131G. RESULTS: Growth of Candida species indicated that a 105 yeast inoculum in SD broth proliferated to a stationary growth equilibrium (approximately 10(9) yeast cell density) in 18 hours (approximately 2 hours of generation time). ALF (5 mg/mL) elicited >96 hours of total stasis (100% growth inhibition) and was significantly effective against both Candida species (p < 0.0001). At 2.5 mg/mL dilution, ALF sustained total stasis activity to an average of 18 hours and 24 hours for C albicans (n = 5) and C glabrata (n = 5), respectively. Interaction studies indicated avid binding of C albicans (70 - 140 x 10(3) yeast) and C glabrata (50 - 75 x 10(3) yeast) per square centimeter of VE monolayer. ALF-treated VE showed significant blockade (p < 0.05) of yeast adhesion by 33% and 58% with C albicans and C glabrata, respectively. ALF treatment of yeast-VE complexes resulted in significant detachment (p < 0.05) of C albicans and C glabrata, by 58% and 51%, respectively. CONCLUSION: ALF is a natural fungistatic agent with potent yeast adhesion-blocking and detachment properties and is effective against the vaginal pathogens C albicans and C glabrata.