The Pattern of Liver Dysfunction in Patients with COVID-19: A Retrospective Study

Background. Information about liver dysfunction in patients with COVID-19 is scarce. We aimed to explored the pattern and risk factors of liver dysfunction in patients with COVID-19. Methods. In this retrospective study, we included all consecutive con�rmed patients with COVID-19 in Fuyang Second People’s Hospital between January 20 and February 25, 2020 and collected clinical characteristics until discharge. The pattern and risk factors of liver dysfunction, viral shedding and outcome were analyzed. Results. Totally, 146 patients were analyzed. The median age was 44.9 years and 54.1% were men, 43.8% patients presented liver dysfunction (22.6% on admission, 21.2% during hospitalization). The percentage of elevated ALT (15.1% on admission and 24.7% during hospitalization) were signi�cantly higher than ALP (2.1% on admission and 3.4% during hospitalization) (P < 0.001). Four clinical types were identi�ed, type 1 (persistent normal liver function, 56.2%), type 2 (normal liver function on admission developed to liver dysfunction during hospitalization, 21.2%), type 3 (liver dysfunction on admission restored to normal on discharge, 13.0%) and type 4 (persistent liver dysfunction, 9.6%). The median duration of viral shedding was 12.0 (type 1), 15.0 (type 2), 14.0 (type 3) and 18.0 (type 4) days (P < 0.001). Prolonged viral shedding and severity were potential risk factors associated with liver dysfunction.


Background
][3][4][5][6] SARS-CoV-2 shares 82% genome sequence similarity to SARS-CoV-2 and 50% genome sequence homology to Middle East respiratory syndrome coronavirus (MERS-CoV),and all three coronaviruses can cause severe respiratory symptoms.Liver injury has been reported in patients with SARS or MERS-CoV. 7,8tudies have con rmed that SARS-CoV-2 enters cells primarily through angiotensin converting enzyme2 (ACE2), the high expression of ACE2 in alveolar type II cells makes the lung become the main target organ. 9,10A number of existing clinical studies showed that some patients with COVID-19 had different degrees of liver dysfunction [11][12][13][14][15][16] and the liver biopsy from a dead COVID-19 case showed moderate microvascular steatosis and mild lobular activity in addition to severe lung injury. 17The objective of this retrospective study was to explored the pattern of liver dysfunction, risk factors and outcomes in patients with COVID-19, so as to provide reference for clinical decision-making.

Study design
All consecutive patients with COVID-19 admitted to the Fuyang Second People's Hospital (FYSPH) in Anhui Province of China between January 20, 2020 and February 25, 2020 were retrospective enrolled.
The including criteria was SARS-CoV-2 positive using real-time reverse-transcriptase polymerase chain reaction (RT-PCR) assay by local municipal Center for Diseases Prevention and Control (CDC) as described previously.The excluding criteria was other primary pathogens infection, such as bacteria, fungi, other respiratory virus, mycoplasma, or chlamydia, and age below 18 years old.(Figure 1).Clinical outcomes were followed up until discharge.COVID-19 was diagnosed based on the guidance for corona virus disease 2019 issued by National Health Commission of China 18 .The criteria of discharge included all the following conditions: body temperatures remained normal over 3 days, the symptoms of respiratory improved obviously, pulmonary imaging shows remarkable absorption of in ammation, and repeated tests for SARS-CoV-2 at least 24 hours apart con rmed viral clearance.The study was approved by the Ethics Committees of FYSPH (20200303006).Written informed consent was waived in view of the designated hospital for new emerging infectious diseases.

Data collection
All the patients' information, such as epidemiological, demographic (age, sex, etc.), smoking and drinking history , chronic liver disease and comorbidity, laboratory ndings, treatment and outcome data, were collected from electronic medical records.Thereafter, alanine aminotransferase (ALT), aspartate transaminase (AST), total bilirubin (TBIL), alkaline phosphatase (ALP) were collected until the last followup.All the data in source documents were con rmed independently by at least two researchers.

De nitions
Liver dysfunction was de ned as ALT level over 50 U/L (the upper limit of normal, ULN) or AST level over 40 U/L (ULN) or TBIL level over 26μmol/L (ULN) or ALP level over 125 U/L (ULN), according to Fuyang Second People's Hospital laboratory department of the normal reference value.Comorbidity was de ned as having at least one of the followings: hypertension, diabetes, cardiovascular disease, asthma, chronic lung disease and malignancy for over 6 months.We de ned the degree of severity of COVID-19 on admission based on the guidance for corona virus disease 2019 issued by National Health Commission of China 18 .Viral shedding was calculated by date of SARS-CoV-2 negative minus date of illness onset.

Statistical analysis
Continuous variables were expressed as medians and interquartile ranges and were compared using the Mann-Whitney test.Categorical variables were presented as numbers (percentage) and compared by the chi-square test or the Fisher exact test.Correlation analyses were performed by Pearson's method.
Comparisons among multiple groups were performed using one-way ANOVA and pairwise comparisons were performed using the LSD test.A P-value < 0.05 was considered as signi cant for all statistical tests.The statistical analyses were performed using SPSS (version 22.0; SPSS, Chicago, IL).

The dynamic change in four types of liver dysfunction
According to clinical course of liver dysfunction, four clinical types were identi ed, type 1 (persistent normal liver function, 56.2%), type 2 (normal liver function on admission developed to liver dysfunction during hospitalization, 21.2%), type 3 (liver dysfunction on admission restored to normal on discharge, 13.0%) and type 4 (persistent liver dysfunction, 9.6%) (Figure 1,Table 2).

Discussion
COVID-19 has developed into a pandemic, although some literature had reported 14-53% patients with COVID-19 had elevated ALT and AST, 2,3,12,15 little data analyzed other liver enzymes (such as ALP) and the dynamic change of liver function from admission until discharge.In this retrospective study, we clari ed four clinical types, and concluded that the liver injury was common but slight in non-critical patients.In 146 enrolled patients, 43.8% patients presented liver dysfunction (22.6% on admission, 21.2% during hospitalization).Brie y, the percentage of elevated ALT (15.1% on admission and 24.7% during hospitalization) were signi cantly higher than ALP (2.1% on admission and 3.4% during hospitalization) (P < 0.001).
Next, we analyzed the potential mechanisms associated with liver dysfunction in COVID-19 patients.First, SARS-CoV-2 directly damage the hepatocyte.Zhang et al consider the liver injury could have been caused by SARS-CoV-2 infection based on SARS-CoV-2 RNA has been detected in stool and blood samples, 19 Chai et al found that the speci c expression of ACE2 in cholangiocytes was 20 times higher than hepatocyte, 20 while Xu et al analyzed the pathological of liver tissue from a patient who died from COVID-19 did not observed viral inclusions in the liver. 17In our study, ALP, a diagnostic biomarker for cholangiocyte injury, was not observed signi cant elevated and the incidence was scarce.Therefore, liver injury in COVID-19 patients was not directly caused by SARS-CoV-2 infection.Second, SARS-CoV-2mediated immune injury.2][23] In the present study, viral shedding was signi cant longer in patients with liver dysfunction than normal liver function which implied that prolonged viral shedding caused more intense cytokine storm, and further damage of multiple organs, including the liver.We speculate that liver dysfunction of type 3 might induced by this mechanism.Third, drug-induce liver injury.Some studied showed that about majority of COVID-19 patients had received nonsteroidal anti-in ammatory drugs, antibiotics and antiviral agents (e.g., acetaminophen, moxi oxacin, oseltamivir, arbidol, lopinavir/ritonavir, etc), all of above drugs had explicit liver injury effect, such as type 2 and type 4, however, it is even di cult to evaluate exact liver damage particularly in combination therapy. 2,3,11,12,14,22urth, underlying of chronic liver disease.Guan et al found that non-severe patients with COVID-19, even though they have basic liver diseases (such as viral hepatitis, etc.), seldom showed liver dysfunction which is consistent with our results. 13Nevertheless, we should pay more attention to the potential liver injury in those with chronic hepatitis B, obesity, hyperlipidemia and metabolic syndrome.
This study has several limitations.First, the number of severe or critical patients was small, so the pattern of liver dysfunction in these kinds of patients was not obtained.Second, due to the limitations of local test condition, many cytokines (e.g., IL-6, TNF-α, ferritin, percentage of T lymphocytes) could not be detected.Last, this is a single-center study, and more patients are needed to further study.

Conclusion
our study summarized four clinical types of liver dysfunction in patients with COVID-19, and speculated that the mechanism of liver dysfunction might due to SARS-CoV-2-mediated immune injury on hepatocyte, besides DILI and underlying chronic liver disease.Further research should focus on the causes of liver dysfunction, treatment and outcome of COVID-19.All data are presented as n (%) or † median (IQR).Comparison between two groups was performed using a (Chi-square test) or b (Manm-Whitney U test) or c (Fisher's exact test) as appropriate.ALT, alanine aminotransferase; AST, aspartate transaminase; TBIL, total bilirubin; ALP, alkaline phosphatase.* indicated significant difference between ALT or AST and ALP (P <0.001).

Fever
All data are presented as n (%) or † median (IQR).Comparison between four types was performed using a (Chi-square test) or b (ANOVA) as appropriate.

Figure 1 Flow
Figure 1

Figure 3 Dynamic
Figure 3

Table 1
Baseline characteristics and liver function of the Study Patients with COVID-19 on admission

Table 2
Features and risk factors of four clinical types in patients with COVID-19