By Karina Toledo | Agência FAPESP – After analyzing laboratory tests pertaining to almost 179,000 Brazilians, 33,266 of whom tested positive for SARS-CoV-2, a group of researchers identified different clinical profiles influenced by the patient’s biological sex and age, as well as the severity of the disease.
The study was supported by FAPESP, and the findings are reported in an article available from medRxiv, which publishes preprints (scientific papers that have not yet been peer-reviewed).
According to the authors, the findings will be useful to health workers on the front line of the fight against the COVID-19 pandemic.
“The coronavirus can unleash a broad spectrum of clinical manifestations, ranging from asymptomatic or mild disease to severe disease and death. Laboratory parameters also vary considerably according to the patient’s age and biological sex, and physicians often struggle to interpret test results in search of significant alterations. We hope our study will help them in this process,” Helder Nakaya, a professor at the University of São Paulo’s School of Pharmaceutical Sciences (FCF-USP), told Agência FAPESP.
The group led by Nakaya used bioinformatics methods to analyze more than 200 laboratory parameters for the thousands of patients in the sample, including complete blood cell count, electrolytes, metabolites, arterial blood gases, liver enzymes, hormones, and inflammation biomarkers. These are standard tests in the clinical investigation of patients with suspected COVID-19 or other infectious diseases.
The data used in the study came from COVID-19 Data Sharing/BR, an open-access repository established by FAPESP and the University of São Paulo (USP) to hold anonymized data on patients tested for the novel coronavirus by partner healthcare institutions. At the time of the study, the repository held data from the laboratories of Fleury Group throughout Brazil and two leading private hospitals in São Paulo City, Hospital Sírio-Libanês and Albert Einstein Jewish Hospital (HIAE) (read more at agencia.fapesp.br/33633 and agencia.fapesp.br/33522).
Other health institutions are joining the group, and will deposit data from their patients in the repository. At the moment, there are six additional institutions: the Hospital das Clínicas, the general hospital run by USP’s Medical School (HC-FMUSP); Beneficência Portuguesa de São Paulo (BP), one of the largest private hospitals in Latin America; SPDM, the Paulista Association for the Development of Medicine; Hospital Infantil Sabará, a leading children’s hospital in São Paulo, and its research arm Instituto PENSI; and Real Hospital Português de Beneficência in Recife.
“This is the largest cohort of COVID-19 patients with systematically analyzed laboratory data so far,” Nakaya said. “Previous research has looked at relationships between the disease and many of these parameters, especially cytokines [molecules that trigger the inflammatory process] and C-reactive protein [CRP, a key biomarker of systemic inflammation], but the papers published hitherto reported on studies of only a few parameters in many individuals or many parameters in only a few individuals.”
The first step, he explained, was to separate the different groups of patients to be analyzed, mainly in terms of age, sex, and diagnostic test result. The next was to clean up the data from the various institutions that contribute to the repository in order to eliminate redundancies and discrepancies (e.g. different names for the same lab test). After processing the results to form a single harmonized database, a sequence of analyses was performed to produce profiles of the patient groups and compare them with controls (individuals who took the same tests but were not diagnosed as having COVID-19).
Outcome data (hospitalization, death, mild symptoms, recovery) was not available for most of the sample, but part of the data sent by Sírio-Libanês was identified as pertaining to patients in intensive care, enabling the researchers to investigate the laboratory profile of patients on mechanical ventilators and compare it with that of non-hospitalized patients.
Initially considered a respiratory tract infection, COVID-19 has turned out to be a systemic disease that can be associated with gastrointestinal, hepatic, cardiovascular and neurological disorders, and can progress to acute respiratory distress syndrome, multiple organ failure, and death.
“These extrapulmonary manifestations are associated with alterations in circulating levels of several biochemical parameters, such as bilirubin, urea, creatinine, myoglobin and coagulation factors. Little is known about the influence of age and sex on the pattern of these parameters,” said Bruno Andrade, a physician and researcher affiliated with Oswaldo Cruz Foundation (FIOCRUZ) in Salvador, Bahia, and a co-author of the article.
According to Andrade, observational studies based on the analysis of hospitalizations and deaths suggest that the risk of severe COVID-19 is greatest for older males. “But biological confirmation is required for this correlation, and the physiopathological mechanism has yet to be fully elucidated,” he said.
The results of the analyses reported in the article showed that for male subjects aged 13-60 infection by SARS-CoV-2 induced more alterations in laboratory parameters than for female subjects in the same age group. In older subjects, several parameters were altered for both men and women.
Levels of CPR and ferritin, among other indicators of systemic inflammation, were significantly altered in COVID-19 patients, especially men over 60. This suggests the high frequency of complications and death documented for this subpopulation in other research may be directly associated with dysregulated systemic inflammation.
Alterations in liver function tests (AST, ALT, GGT) were frequent in several age groups except young women. According to the authors, this suggests liver dysfunction is common among COVID-19 patients. “This observation is important because of the liver’s key role in the organism,” Andrade said. “It coordinates the production of several proteins and other molecules that regulate inflammation and coagulation, for example. Alterations in the liver may be a decisive factor in the exacerbated systemic inflammation associated with the most unfavorable clinical outcomes.”
Analyzing levels of white blood cells, the researchers found that low levels of basophils and eosinophils (immune cells with a key role in fighting viral infection) were more frequent in older COVID-19 patients of both sexes. Men diagnosed with the disease had higher levels of neutrophils (also important immune cells), and these tended to rise with age. According to Andrade, a high neutrophil count is a marker of acute systemic inflammation.
Finally, in intensive-care patients, the group found major alterations in d-dimer and other tests for blood clots, higher neutrophil counts, and higher levels of markers of systemic inflammation (such as CPR) and damage to cells and tissues (such as lactate dehydrogenase).
“These findings suggest a clear association between disease severity and exacerbated inflammatory processes that may lead to clotting,” Andrade said. “The hypothesis that coagulopathy triggers inflammation is significant since it means treatment can be optimized by including anti-inflammatory medications, but more research is needed to test this idea directly.”
Researchers affiliated with USP, FIOCRUZ and the Federal University of São Paulo (UNIFESP) participated in the study coordinated by Nakaya, which was funded by FAPESP via several projects (2018/14933-2, 2018/21934-5, 2017/27131-9, 2013/08216-2, 2019/27139-5 and 2019/13880-5).
“The paper is the first to analyze data from COVID-19 Data Sharing/BR,” Nakaya said. “We submitted it for publication about a month after the repository began operating. This shows it’s possible to make rapid use of big datasets shared by several centers.”
The open-access repository holds three types of data: demographics (gender, year of birth, region of residence); clinical examinations and laboratory tests; and patient transfers and outcome, when available – for instance, sent to emergency, recovery, or death.
According to Fátima Nunes, computer scientist, professor in USP’s School of Arts, Sciences and Humanities (EACH), and head of its São Paulo information technology center (CETI-SP), since the repository was launched on July 1 it has registered almost 6,500 visits and more than 2,500 downloads of data in all four categories. At a later stage, it will also hold X-rays, CT scans, and other images.
“It’s a powerful resource and most valuable for research on the disease. I’m not aware of any other initiatives of the kind in Brazil,” Andrade said.
The article “In-depth analysis of laboratory parameters reveals the interplay between sex, age and systemic inflammation in individuals with COVID-19” can be read at: www.medrxiv.org/content/10.1101/2020.08.07.20170043v2.