In chronic HBV infection, elevated reactive oxygen species levels derived from dysfunctional mitochondria can cause increased protein oxidation and DNA damage in exhausted virus-specific CD8 T cells. The aim of this study was to understand how these defects are mechanistically interconnected to further elucidate T cell exhaustion pathogenesis and, doing so, to devise novel T cell-based therapies. DNA damage and repair mechanisms, including parylation, CD38 expression, and telomere length were studied in HBV-specific CD8 T cells from chronic HBV patients. Correction of intracellular signalling alterations and improvement of antiviral T cell functions by the NAD precursor nicotinamide mononucleotide and by CD38 inhibition was assessed. Elevated DNA damage was associated with defective DNA repair processes, including NAD-dependent parylation, in HBV-specific CD8 cells of chronic HBV patients. NAD depletion was indicated by the overexpression of CD38, the major NAD consumer, and by the significant improvement of DNA repair mechanisms, and mitochondrial and proteostasis functions by NAD supplementation, which could also improve the HBV-specific antiviral CD8 T cell function. Our study delineates a model of CD8 T cell exhaustion whereby multiple interconnected intracellular defects, including telomere shortening, are causally related to NAD depletion suggesting similarities between T cell exhaustion and cell senescence. Correction of these deregulated intracellular functions by NAD supplementation can also restore antiviral CD8 T cell activity and thus represents a promising potential therapeutic strategy for chronic HBV infection. Correction of HBV-specific CD8 T cell dysfunction is believed to represent a rational strategy to cure chronic HBV infection, which however requires a deep understanding of HBV immune pathogenesis to identify the most important targets for functional T cell reconstitution strategies. This study identifies a central role played by NAD depletion in the intracellular vicious circle that maintains CD8 T cell exhaustion, showing that its replenishment can correct impaired intracellular mechanisms and reconstitute efficient antiviral CD8 T cell function, with implications for the design of novel immune anti-HBV therapies. As these intracellular defects are likely shared with other chronic virus infections where CD8 exhaustion can affect virus clearance, these results can likely also be of pathogenetic relevance for other infection models.

INTRODUCTION: Patients with type 2 diabetes mellitus (T2DM) are at increased risk of metabolic dysfunction-associated steatotic liver disease, advanced liver fibrosis, and metabolic dysfunction-associated steatohepatitis (MASH). We evaluated the prevalence and severity of metabolic dysfunction-associated steatotic liver disease among patients with T2DM at their first referral to diabetes clinics and assessed the effectiveness of the 2-tier screening approach by Fibrosis-4 (FIB-4) and vibration-controlled transient elastography (VCTE). METHODS: Consecutive patients with T2DM from 6 different diabetes clinics were prospectively enrolled. Liver stiffness measurement (LSM) was assessed by VCTE, whereas liver steatosis by controlled attenuation parameter (Fibroscan, Echosens, France). “At-risk MASH” was assessed by FibroScan-aspartate aminotransferase score. RESULTS: Eight hundred patients (median age: 59, 53–65 years; males: 485, 60.6%) met the inclusion criteria. Prevalence of liver steatosis (controlled attenuation parameter ≥ 248 db/m) was 73.6%. The proportion of patients at medium/high risk of advanced liver fibrosis (LSM ≥ 8.0 kPa) was 16.9%. Patients with “at-risk MASH” (FibroScan-aspartate aminotransferase > 0.67) were 12.0%. A 2-tier screening for advanced liver fibrosis by FIB-4 and VCTE would have led to 70 patients (8.8%) referred to liver clinics with a false-negative rate of 9.6% (n = 77; patients with FIB-4 < 1.3 and LSM ≥ 8.0 kPa). At multivariate analysis, overweight/obesity (odds ratio = 3.13, 95% confidence interval 1.23–7.97) and elevated alanine aminotransferase (odds ratio = 1.91, 95% confidence interval 1.17–3.10) were independently associated with LSM ≥ 8.0 kPa in patients with FIB-4 < 1.3. DISCUSSION: In diabetes clinics, the 2-tier screening using FIB-4 and VCTE is effective for the identification of patients with T2DM to be referred to hepatologists. VCTE referral may be considered for patients with overweight/obesity and elevated alanine aminotransferase classified as at low risk of advanced liver fibrosis by FIB-4.