Clues toward the Cause of Long COVID

I previously shared some of the amazing data from the paper "Muscle abnormalities worsen after post-exertional malaise in long COVID" by Appelman et al. 

The paper will undoubtable become a classic in the field of Long COVID, providing a fascinating series of clues that the researchers followed past several dead-ends to their interesting implications. 

First, and most importantly, the researchers confirmed beyond a shadow of doubt that Post-Exertional Malaise (PEM) is a real disease, with myriad muscle and metabolic abnormalities in the Long COVID patients following intense exercise.  Metabolomics provided additional key findings, including the first clue: a possible blockage of glycolysis in Long COVID (see diagrams in previous post).  

A Clue: Glycolysis Blockage

In the glycolysis pathway, the phosphoenolpyruvate (PEP) levels of Long COVID patients were increased, while pyruvate levels were decreased, indicating a disruption or imbalance in enzyme activities within the pathway.   This could be due to a decreased activity of the enzyme pyruvate kinase (PK), which converts PEP to pyruvate in the final step of glycolysis.  Reduced PK activity would result in a buildup of PEP.  With less PEP being converted to pyruvate, the downstream levels of pyruvate would be lower.  

There are several interconnected regulatory pathways that can reduce the activity of pyruvate kinase (PK), the two most relevant being Oxidative Stress and Hypoxia.  Increased levels of reactive oxygen species (ROS) or oxidative stress can lead to the oxidation and inactivation of PK.  Under hypoxic conditions (low oxygen levels), the transcription factor HIF-1 (Hypoxia-Inducible Factor 1) can be activated, which can lead to the downregulation of PK expression and activity.  This is part of the cellular adaptation to hypoxia, where glycolysis is regulated to favor the production of metabolic intermediates for other pathways.

Hypoxia?

In the context of Long Covid and post-exertional malaise (PEM), hypoxia from microclots has been suggested to increase lactic acid production. However, in this study the metabolomics showed decreased* lactic acid, because glycolysis was shut down at PEP by loss of PK activity, not at pyruvate by loss of pyruvate dehydrogenase (PDH).  

The researchers looked for but did not find decreased muscle oxygen perfusion or any differences in microvasculature.  The researchers noted decreased oxygen utilization, but this could be due to anything that disrupts metabolism and does not indicate hypoxia as a specific issue.  

They noted amyloid plaques in the extracellular matrix; the plaques were not blocking the microcapillaries and it is unclear what role they play in the pathophysiology of Long COVID: are they a cause of PEM, or a consequence?  

Their observation that Long COVID patients' muscle force was not dependent on The Citric Acid (TCA) cycle enzyme succinate dehydrogenase (SDH) can also be explained by impaired metabolism upstream of TCA Cycle, i.e. in glycolysis.

Oxidative Stress

Therefore, it seems likely that the regulatory pathway most likely to contribute to reduced pyruvate kinase (PK) expression and activity is the oxidative stress pathway.  

Long Covid patients have been reported to exhibit higher levels of oxidative stress markers, such as lipid peroxidation products and decreased antioxidant levels, compared to healthy individuals or those who have recovered from acute COVID-19 infection.  Physical exertion and exercise can lead to an acute increase in reactive oxygen species (ROS) production, potentially exacerbating oxidative stress in individuals with Long Covid and triggering PEM symptoms.

Some studies have suggested that Long Covid patients may experience mitochondrial dysfunction, which can further contribute to increased ROS generation and oxidative stress.

Next Steps

The paper concluded with these results, but the logical next step would be to use metabolomics to assess free radical concentrations.  One theory is that COVID spike proteins form "pores", or holes in the mitochondrial membranes, disrupting the mitochondria and releasing free radicals into the cell.  

The researchers did look at COVID nucleocapsid protein, but found it in both the control and Long COVID groups in equal concentrations, suggesting that remnant viral protein doesn't explain the pathophysiology of Long COVID.  But maybe remnant virus affects the Long COVID patients differently?  

The researchers noted immune cell infiltration into muscle tissue, which could be in response to a signal from excess free radicals, or could be due to some other reason like persistent COVID infection/expression.  

Lactate?

This study seems to indicate the lactate is not an important variable for the pathophysiology of Long COVID.  However, the details about how lactate was measured limit these conclusions. The researchers measured lactate from three different sources:  metabolomic blood (venous) and muscle lactate measured before and after PEM, and capillary (i.e. finger prick) lactate measured during exercise.  Venous lactate measured one week after PEM induction showed a slightly increased level in Long COVID patients, but none of the other metabolomic lactate measurements showed any difference.  The capillary lactate also showed a slightly elevated blood lactate level before exercise in the Long COVID patients, but the difference was not significantly different.  

However, the baseline measurements were not taken in a fasted condition, and because eating normally raises resting lactate it cannot be determined from this study if fasted lactate might show other differences that are important to Long COVID.

Post Exertional Malaise in Long Covid

 Severe exercise-induced myopathy has been found in long COVID post-exertional malaise (PEM).  

Just look at these long COVID patients (in red) pushing close to 20 mmol/L lactate on the exercise bike!  That is serious dedication for a group of patients who know what the consequences will be.

After the exercise test, blood metabolomics show elevated glycolysis, but decreased pyruvate and TCA cycle metabolites.

Muscle biopsy metabolomics show decreased purine synthesis and TCA cycle.

Key:

Appelman, B., Charlton, B.T., Goulding, R.P. et al. Muscle abnormalities worsen after post-exertional malaise in long COVID. Nat Commun 15, 17 (2024). https://doi.org/10.1038/s41467-023-44432-3

Lactic Acid in Health and Disease

 Introduction

Lactic acid is produced for energy when mitochondria can't use oxygen for aerobic respiration. 

Lactic acidosis develops when you have too much lactic acid in your body. Athletes monitor their blood lactate levels as a way to pace their training.  Chronic resting lactate levels greater than 2 mmol/L represent hyperlactatemia, whereas lactic acidosis is generally defined as a serum lactate concentration above 4 mmol/L.   High levels of lactic acid can cause muscle soreness, fibromyalgia-like symptoms, and anxiety.

Overproduction or Under-removal?

The body naturally produces and consumes lactate: although anaerobic exercise can raise blood lactate levels over 10 mmol/L, resting lactate in health adults is usually between 1 and 2 mmol/liter and is constantly produced and consumed.  High levels of lactic acid can be caused by overproduction or under-removal of lactate.

Resting lactate is usually between 0.5-1.5 in a large (10,000 participant) study of healthy adults.  More info.

Although many organs consume lactate, the liver and the kidney represent the major sites of lactate uptake and clearance as they metabolize approximately 53% and 30% of daily lactate production, respectively. Lactate is metabolized by two main mechanisms: First, lactate can be used as a substrate to regenerate glucose by gluconeogenesis, a process that is exclusive to liver and the kidney. Second, at least 50% of circulating lactate is removed and metabolized by means of oxidation during resting conditions. Unlike gluconeogenesis, which is restricted to liver and kidney, oxidation can take place in many organs, including the heart, brain, and skeletal muscle.

Supplements that affect Lactic Acid

Biotin (B7) is a cofactor required for gluconeogensis.  Aspirin can increase lactate levels by interfering with LDH lactate dehydrogenase, the enzyme responsible for turning lactate into pyruvate for gluconeogensis.   B1 deficiency impairs Citric Acid cycle and leads to accumulation of pyruvate and lactate.  Organic acids involved in the Citric Acid cycle may may provide substrates to better metabolize lactate and may reduce the acidifying side effects of elevated lactate (i.e. reduce Potential Renal Acid Load (NRAL)).  Nutrient deficiencies of CoQ10 and lipoic acid have also been associated with elevated lactic acid levels in both urine and blood.   Magnesium in muscles helps to decrease contractions and lactate buildup.  

Exercise

Slow exercise (i.e. HR below 100) can lower blood lactate and blood glucose levels in healthy adults.

Mitochondrial Dysfunction in Disease

Many diseases, included Type-2 diabetes, Chronic Fatigue Syndrome, and Long Covid, are associated with mitochondrial dysfunction and increased resting lactate levels.  Post-exertional malaise (PEM) is a major symptom of ME/CFS and Long Covid and may be caused by elevated lactate levels.  Although we don’t know the cure, #StopRestPace is the best treatment because only resting and then reducing/pacing activity can lower lactate levels.  

Lactic acid is being investigated as a possible metric of Long Covid and PEM severity.  If it plays a causal role in these diseases, methods that reduce lactic acid buildup may be promising treatments.  

Research

Elevated blood lactate in resting conditions correlate with post-exertional malaise severity in patients with Myalgic encephalomyelitis/Chronic fatigue syndrome. https://www.nature.com/articles/s41598-019-55473-4

-Patients having ≥1 lactate measurement ≥2 mmol/L defined elevated lactate group. The study included 123 patients. Elevated (n = 55; 44.7%) and normal (n = 68; 55.3%) lactate groups were comparable except for PEM, which was more severe in the elevated lactate group.

Decreased Fatty Acid Oxidation and Altered Lactate Production during Exercise in Patients with Post-acute COVID-19 Syndroms.  https://www.atsjournals.org/doi/full/10.1164/rccm.202108-1903LE

-The transition from fat oxidation to glucose oxidation occurs prematurely, suggesting metabolic reprogramming and dysfunctional mitochondria.

Figure 1 from paper.  Long Covid patients show greater increases in lactic acid at low intensity exercise compared to controls.  Blue and purple are Long Covid patients (with and without comorbidities (like diabetes), respectively) and grey and black are matched controls (with and without comorbidities, respectively).