SARS-CoV-2, the virus that causes COVID-19, likely does not directly infect the brain but can still inflict significant neurological damage, according to a new study from neuropathologists, neurologists, and neuro-radiologists at Columbia University Vagelos College of Physicians and Surgeons.
"There's been considerable debate about whether this virus infects the brain, but we were unable to find any signs of virus inside brain cells of more than 40 COVID-19 patients," says Dr James E Goldman, professor of pathology & cell biology (in psychiatry), who led the study with Dr Peter D Canoll, professor of pathology & cell biology, and Dr Kiran T Thakur, the Winifred Mercer Pitkin assistant professor of neurology.
"At the same time, we observed many pathological changes in these brains, which could explain why severely ill patients experience confusion and delirium and other serious neurological effects – and why those with mild cases may experience “brain fog” for weeks and months."
The study is the largest and most detailed COVID-19 brain autopsy report published to date, suggests that the neurological changes often seen in these patients may result from inflammation triggered by the virus in other parts of the body or in the brain's blood vessels.
The study examined the brains of 41 patients with COVID-19 who succumbed to the disease during their hospitalisation. The patients ranged in age from 38 to 97; about half had been intubated and all had lung damage caused by the virus. Many of the patients were of Hispanic ethnicity. There was a wide range of hospital length with some patients dying soon after arrival to the emergency room while others remained in the hospital for months. All of the patients had extensive clinical and laboratory investigations, and some had brain MRI and CT scans.
To detect any virus in the neurons and glia cells of the brain, the researchers used multiple methods including RNA in situ hybridization, which can detect viral RNA within intact cells; antibodies that can detect viral proteins within cells; and RT-PCR, a sensitive technique for detecting viral RNA.
Despite their intensive search, the researchers found no evidence of the virus in the patients' brain cells. Though they did detect very low levels of viral RNA by RT-PCR, this was likely due to virus in blood vessels or leptomeninges covering the brain.
"We've looked at more brains than other studies, and we've used more techniques to search for the virus. The bottom line is that we find no evidence of viral RNA or protein in brain cells," Goldman says. "Though there are some papers that claim to have found virus in neurons or glia, we think that those result from contamination, and any virus in the brain is contained within the brain's blood vessels." "If there's any virus present in the brain tissue, it has to be in very small amounts and does not correlate with the distribution or abundance of neuropathological findings," Canoll says.
The tests were conducted on more than two dozen brain regions, including the olfactory bulb, which was searched because some reports have speculated that the coronavirus can travel from the nasal cavity into the brain via the olfactory nerve. "Even there, we didn't find any viral protein or RNA," Goldman says, "though we found viral RNA and protein in the patients' nasal mucosa and in the olfactory mucosa high in the nasal cavity." (The latter finding appears in an unpublished study, currently on BioRxiv, led by Dr Jonathan Overdevest, assistant professor of otolaryngology, and Dr Stavros Lomvardas, professor of biochemistry & molecular biophysics and neuroscience.)
Despite the absence of virus in the brain, in every patient the researchers found significant brain pathology, which mostly fell into two categories.
"The first thing we noticed was a lot of areas with damage from a lack of oxygen," Goldman says. "They all had severe lung disease, so it's not surprising that there's hypoxic damage in the brain."
Some of these were large areas caused by strokes, but most were very small and only detectable with a microscope. Based on other features, the researchers believe these small areas of hypoxic damage were caused by blood clots, common in patients with severe COVID-19, that temporarily stopped the supply of oxygen to that area.
A more surprising finding, Goldman says, was the large number of activated microglia they found in the brains of most patients. Microglia are immune cells that reside in the brain and can be activated by pathogens.
"We found clusters of microglia attacking neurons, a process called 'neuronophagia,'" says Canoll. Since no virus was found in the brain, it's possible the microglia may have been activated by inflammatory cytokines, such as Interleukin-6, associated with SARS-CoV-2 infection.
"At the same time, hypoxia can induce the expression of 'eat me' signals on the surface of neurons, making hypoxic neurons more vulnerable to activated microglia," Canoll says, "so even without directly infecting brain cells, COVID-19 can cause damage to the brain."
The group found this pattern of pathology in one of their first autopsies, described by Dr Osama Al-Dalahmah, instructor in pathology & cell biology. Over the next few months, as the neuro-pathologists did many more COVID brain autopsies, they saw similar findings over and over again and realised that this is a prominent and common neuro-pathological finding in patients who die of COVID.
The activated microglia were found predominantly in the lower brain stem, which regulates heart and breathing rhythms, as well as levels of consciousness, and in the hippocampus, which is involved in memory and mood.
"We know the microglia activity will lead to loss of neurons, and that loss is permanent," Goldman says. "Is there enough loss of neurons in the hippocampus to cause memory problems? Or in other parts of the brain that help direct our attention? It's possible, but we really don't know at this point."
Goldman says that more research is needed to understand the reasons why some post-COVID-19 patients continue to experience symptoms.
The researchers are now examining autopsies on patients who died several months after recovering from COVID-19 to learn more. They are also examining the brains from patients who were critically ill with acute respiratory distress syndrome (ARDS) before the COVID-19 pandemic to see how much of COVID-19 brain pathology is a result of the severe lung disease.
Study details
COVID-19 neuropathology at Columbia University Irving Medical Center/New York Presbyterian Hospital
Kiran T Thakur, Emily Happy Miller, Michael D Glendinning, Osama Al-Dalahmah, Matei A Banu, Amelia K Boehme, Alexandra L Boubour, Samuel S Bruce, Alexander M Chong, Jan Claassen, Phyllis L Faust, Gunnar Hargus, Richard A Hickman, Sachin Jambawalikar, Alexander G Khandji, Carla Y Kim, Robyn S Klein, Angela Lignelli-Dipple, Chun-Chieh Lin, Yang Liu, Michael L Miller, Gul Moonis, Anna S Nordvig, Jonathan B Overdevest, Morgan L Prust, Serge Przedborski, William H Roth, Allison Soung, Kurenai Tanji, Andrew F Teich, Dritan Agalliu, Anne-Catrin Uhlemann, James E Goldman, Peter Canoll
Published in Brain on 15 April 2021
Abstract
Many patients with SARS-CoV-2 infection develop neurological signs and symptoms, though, to date, little evidence exists that primary infection of the brain is a significant contributing factor. We present the clinical, neuropathological, and molecular findings of 41 consecutive patients with SARS-CoV-2 infections who died and underwent autopsy in our medical center. The mean age was 74 years (38–97 years), 27 patients (66%) were male and 34 (83%) were of Hispanic/Latinx ethnicity. Twenty-four patients (59%) were admitted to the intensive care unit (ICU). Hospital-associated complications were common, including 8 (20%) with deep vein thrombosis/pulmonary embolism (DVT/PE), 7 (17%) patients with acute kidney injury requiring dialysis, and 10 (24%) with positive blood cultures during admission. Eight (20%) patients died within 24 hours of hospital admission, while 11 (27%) died more than 4 weeks after hospital admission. Neuropathological examination of 20–30 areas from each brain revealed hypoxic/ischemic changes in all brains, both global and focal; large and small infarcts, many of which appeared hemorrhagic; and microglial activation with microglial nodules accompanied by neuronophagia, most prominently in the brainstem. We observed sparse T lymphocyte accumulation in either perivascular regions or in the brain parenchyma. Many brains contained atherosclerosis of large arteries and arteriolosclerosis, though none had evidence of vasculitis. Eighteen (44%) contained pathologies of neurodegenerative diseases, not unexpected given the age range of our patients. We examined multiple fresh frozen and fixed tissues from 28 brains for the presence of viral RNA and protein, using quantitative reverse-transcriptase PCR (qRT-PCR), RNAscope, and immunocytochemistry with primers, probes, and antibodies directed against the spike and nucleocapsid regions. qRT-PCR revealed low to very low, but detectable, viral RNA levels in the majority of brains, although they were far lower than those in nasal epithelia. RNAscope and immunocytochemistry failed to detect viral RNA or protein in brains. Our findings indicate that the levels of detectable virus in COVID-19 brains are very low and do not correlate with the histopathological alterations. These findings suggest that microglial activation, microglial nodules and neuronophagia, observed in the majority of brains, do not result from direct viral infection of brain parenchyma, but rather likely from systemic inflammation, perhaps with synergistic contribution from hypoxia/ischemia. Further studies are needed to define whether these pathologies, if present in patients who survive COVID-19, might contribute to chronic neurological problems.