1. Case information
The patient, a 24-year-old female, presented to a local hospital on May 26, 2022 for "epigastric pain for 5 days", and the patient presented with epigastric pain with anorexia, nausea, vomiting, abdominal distention, drowsiness, fatigue, and yellow urine 5 days ago. Liver function tests: TBil 129.5 μmol/L, DBil 104.7 μmol/L, ALT 2 035 U/L, AST 2 039 U/L; Coagulation function: prothrombin time (PT) 45.7 s, international standard ratio (INR) 3.61, prothrombin activity (PTA) 12.6%; Hepatitis B etiology: HBsAg 27.64 IU/mL, HBeAg negative, HBV DNA 2.82×104 IU/mL. FibroScan assay: liver stiffness was 11.6 kPa, and fat attenuation was 203 dB/m. CT scan of the abdomen showed that the surface of the liver was smooth and the density of the liver parenchyma was slightly reduced. After 3 days of antiviral treatment (entecavir), hepatoprotective enzyme yellowing (reduced glutathione, magnesium isoglycyrrhizate, ursodeoxycholic acid), hepatocyte regeneration (hepatocyte auxin), plasma supplementation and cryoprecipitate, and short-term hormone (methylprednisolone sodium succinate 40 mg for 2 consecutive days), he was admitted to our hospital because the TBil level rose to 138 μmol/L. The patient was in good health in the past, and his family and epidemiological history were unremarkable. Denial of previous risk behaviors of HBV infection, and no HBV infection among the patient's family members or daily contacts. Denial of drug abuse, blood transfusion, alcohol consumption, drug use, and travel and residence history in epidemic areas.
Physical examination: lethargy, moderate xanthochromia of the skin and sclera, right upper quadrant tenderness, no rebound tenderness, negative abdominal moving dullness, positive asterixis. Laboratory tests: TBil 182.7 μmol/L, ALT 1 023 U/L, AST 369 U/L, PT 29.2 s, INR 2.64. Tests for HAV, HCV, HEV, cytomegalovirus, Epstein-Barr virus, HIV antibodies, and antibodies associated with autoimmune liver disease were all negative. Continue the treatment of anti-HBV, strengthen anti-inflammatory and hepatoprotective treatment, control hepatic encephalopathy, and replenish plasma, cryoprecipitate and symptomatic treatment. After 17 days of treatment in our hospital, he was discharged after re-examination of all biochemical indicators. The dynamic monitoring of HBsAg and hypersensitive HBV DNA quantification in our hospital showed a continuous downward trend, and HBsAg negative conversion and seroconversion were observed. The anti-HBc test value was below the lower limit of the reference value at the time of admission and continued to increase during the course of the disease, during which anti-HBc-IgM was detected. The patient had acute onset, drowsiness within 2 weeks, and the TBil level increased > 17.1 μmol/L and INR >1.5, and was diagnosed with acute liver failure according to the 2018 guidelines for the diagnosis and treatment of liver failure. After 1 year of outpatient follow-up, liver function and coagulation function were normal. HBsAg negative, HBV DNA < 20 IU/mL, HBV RNA <50 copies/mL, anti-HBs 159.1 mIU/mL; FibroScan assay: liver stiffness 4.2 kPa, fat attenuation 150 dB/m; There was no abnormality on ultrasound of the liver. The dynamic changes of laboratory indexes and HBV serological and virological indexes are shown in Table 1 and 2. Combined with the epidemiological history, dynamic changes and clinical outcomes of HBV-related etiological indicators, and other causes of acute liver failure were ruled out, the cause was considered to be acute HBV infection.
After obtaining informed consent, the activation level of peripheral blood immune cells and anti-HBV-specific immune response were dynamically monitored. Flow cytometry was used to detect the frequencies of T and B lymphocytes and the expression levels of immune indexes at three time points: when HBsAg was negative and seroconversion occurred (T1), 1 week after serological conversion (T2), and 1 year after serological conversion (T3), and peripheral blood mononuclear cells were stimulated with HBV epitope peptide culture in vitro, and the frequency of HBV-specific CD4+T and CD8+ T lymphocytes was detected 1 day later, and IL-2 secretion by HBV-specific T lymphocytes was detected 10 days later. Levels of TNF-α, IFN-γ. The results showed that compared with T1, the frequency of T lymphocytes and B lymphocytes decreased by 24% and 71%, respectively, at T2. The frequency of CD4+ T lymphocytes decreased by 12%, while the frequency of CD8+ T lymphocytes did not change significantly. HBsAg-specific CD4+T and CD8+ T lymphocytes decreased to undetected, and the frequency of HBcAg-specific CD4+T lymphocytes decreased by 79%, while the frequency of HBcAg-specific CD8+ T lymphocytes did not change significantly. The expression levels of programmed death receptor 1 (PD-1) in CD4+ and CD8+ T lymphocytes did not change significantly, the expression levels of CD95, granzyme B (GB) and human leukocyte class II antigen (HLA-DR) increased by 0.6, 1.2 and 0.2 times in CD8+ T lymphocytes, respectively, and the average fluorescence intensity of CD80 and programmed death receptor-ligand 1 (PD-L1) on the surface of B lymphocytes increased by 0.8 and 1.6 times, respectively. Compared with the T2 time point, the frequency of HBsAg-specific CD4+T and CD8+ T lymphocytes increased slightly at T3, the frequency of HBcAg-specific CD4+ T lymphocytes decreased to undetected, and the frequency of HBcAg-specific CD8+ T lymphocytes decreased by 29%. The levels of IFN-γ secreted by HBsAg and HBcAg-specific CD8+ T lymphocytes increased by 3.8 and 1.3 times, respectively, the expression levels of PD-1 in CD4+ and CD8+ T lymphocytes decreased by 46% and 50%, respectively, and the expression levels of HLA-DR in CD8+ T lymphocytes decreased by 45%. The average fluorescence intensity of CD80 and PD-L1 on the surface of B lymphocytes decreased by 43% and 32%, respectively.
2. Discussion
About one-third of adults infected with HBV have manifestations of acute hepatitis B (AHB), and nearly 1% of patients with AHB progress to acute liver failure, and the incidence of liver failure due to acute HBV infection may be underestimated because the patient's HBsAg or HBV DNA is below the lower limit of detection at presentation. Acute liver failure refers to liver failure characterized by grade II or higher hepatic encephalopathy within 2 weeks without a history of underlying liver disease. In this case, a young woman, the anti-HBc test value was lower than the lower limit of the reference value at the time of admission and continued to increase during the course of the disease, during which the anti-HBc-IgM was detected positive. HBV DNA decreased rapidly, with a decrease of more than 2 log10 IU/mL in 6 days, and rapid negative transorration and seroconversion of HBsAg were monitored. Because the patient has not been screened for hepatitis B before, the possibility of acute exacerbation of chronic hepatitis B cannot be ruled out, but the change from negative to positive anti-HBc during the course of the disease does not support chronic HBV infection. Combined with the patient's symptoms, signs, laboratory test results, epidemiological history, family history and personal history, according to the diagnostic criteria for AHB in the Chinese Standards for the Diagnosis and Treatment of Liver Diseases, the patient met the following four criteria: (1) recent onset of fatigue and gastrointestinal symptoms that could not be explained by other reasons; (2) abnormal liver biochemical examination; (3) HBsAg positive; (4) Acute HBV infection can be diagnosed if the convalescent serum HBsAg is negative and anti-HBs are positive. After ruling out other causes of acute liver failure, consider that acute liver failure is caused by acute hepatitis B virus infection.
During the course of the disease, it was observed that in the case of continuous increase in TBil and continuous decrease in ALT and AST, that is, "choline enzyme separation", APTT, PT and INR showed an improvement trend, and a decrease in blood ammonia and a significant increase in AFP were observed on the 4th day of admission to our hospital, indicating hepatocyte regeneration and improvement. After 1 year of follow-up, HBsAg was negative, and the anti-HBs remained at a high level. It is important to note that anti-HBc-IgM is only tested positive when HBsAg undergoes seroconversion, so multiple dynamic monitoring of quantitative changes in HBV etiology-related indicators can help confirm the diagnosis and avoid missing the diagnosis.
The outcome of HBV infection mainly depends on the interaction between the virus and host immunity, in which the viability and potency of HBV-specific T lymphocytes and the neutralizing antibodies secreted by B lymphocytes play an important role. Liver injury caused by HBV infection is mediated by an immune response rather than by a direct viral action. Typical AHB liver injury is thought to be mediated by a cellular immune response, with more HBV-specific cytotoxic T lymphocytes detected in patients with acute liver failure compared with patients with AHB. Humoral immunity mediated by intrahepatic HBcAg-specific B lymphocytes can also cause liver injury, and studies have found that a large number of high-affinity IgM and IgG antibodies are present in the liver of patients with AHB progression to acute liver failure and require liver transplantation, which target HBcAg to form antigen-antibody complexes and activate complement, resulting in large-scale necrosis of liver tissue. The above studies have shown that the body clears the virus through the immune response, but excessive immune activation can cause serious or even irreversible damage to the body.
Acute liver failure has a high mortality rate, with 75% of HBV-associated acute liver failure reported requiring liver transplantation or death. Although the patient developed acute liver failure, the prognosis was good. At 1 week of HBsAg seroconversion, the peripheral total PD-1+CD8+ T lymphocyte frequency and the PD-L1 expression level on B lymphocytes were higher than those after 1 year, while the levels of IFN-γ secreted by HBcAg and HBsAg-specific CD8+ T lymphocytes were lower than those after 1 year. Previous studies have found similar findings that in AHB patients, PD-1 expression of early HBV-specific CD8+ T lymphocytes is up-regulated, and blocking PD-1 in vitro can significantly enhance the proliferation of HBV-specific T lymphocytes and their IFN-γ production, indicating that PD-1 upregulation can reduce pathogenic CD8+ T lymphocyte response and liver injury. Other studies have shown that the high expression of PD-L1 on B lymphocytes inhibits the activity and proliferation of follicular helper T lymphocytes and the production of antibodies, thereby strongly inhibiting humoral immunity. The above studies have shown that PD-1 and PD-L1 can reduce liver injury by inhibiting T lymphocyte activity and B lymphocyte effect, respectively, suggesting that PD-1 and PD-L1 play a protective role in the course of acute liver failure to avoid further deterioration of liver failure.
HLA-DR is a marker of T lymphocyte activation. In this case, the patient observed that a higher frequency of HLA-DR+CD8+ T lymphocytes was detected in the peripheral blood during HBsAg seroconversion compared with 1 year later. Our team also found that the frequency of HLA-DR+CD8+ T lymphocytes was significantly increased before HBsAg negative conversion in patients with chronic hepatitis B. Sprengers et al. showed that HLA-DR was highly expressed in HBV-specific CD8+ T lymphocytes in both intrahepatic and peripheral blood of AHB patients, and decreased rapidly after HBsAg seroconversion. Therefore, the high expression of HLA-DR on CD8+ T lymphocytes may be associated with HBsAg negative transfection.
Unlike patients with functional cure of chronic hepatitis B, HBV DNA and HBV RNA tests were still positive at HBsAg serological conversion, and decreased after 1 week. However, HBV-specific CD8+ T lymphocytes remained high after 1 week of seroconversion, and the frequencies of CD95+CD8+ T lymphocytes and GB+CD8+ T lymphocytes were significantly higher than those during seroconversion. Therefore, it is hypothesized that CD8+ T lymphocytes may continue to clear the virus through CD95/CD95L-mediated apoptosis and GB-mediated cytotoxicity after seroconversion. In addition, HBV DNA and HBV RNA tests in peripheral blood were negative after 1 year, but HBV-specific CD8+ T lymphocytes and high levels of IFN-γ secretion could still be detected, suggesting that there is still HBV DNA in the liver and CD8+ T lymphocytes play an important role in the long-term control of the virus.
In summary, through the dynamic analysis of this case, the complex pathological mechanism of liver failure associated with acute HBV infection and the key immune events in the treatment process were revealed. Data from this patient suggest that CD8+ T lymphocytes and B lymphocytes play a key role in viral control and liver failure prognosis, and are involved in regulating and maintaining immune homeostasis, in addition to directly mediating antiviral effects. At the same time, dynamic monitoring of HBV etiology and immunology-related indicators plays an important role in clinical diagnosis and treatment decision-making, and further in-depth exploration of related research has important guiding significance for individualized precision diagnosis and treatment.
引证本文 Citation
QUAN Xufeng, XIONG Shu'e, ZHOU Wenqing, et al. Clinical cure of liver failure associated with acute HBV infection with HBsAg serological conversion: a report in 1 case[J]. Journal of Clinical Hepatobiliary Diseases, 2024, 40(7): 1446-1449