Liver transcriptomics highlights interleukin-32 as novel NAFLD-related cytokine and candidate biomarker.

OBJECTIVE: Efforts to manage non-alcoholic fatty liver disease (NAFLD) are limited by the incomplete understanding of the pathogenic mechanisms and the absence of accurate non-invasive biomarkers. The aim of this study was to identify novel NAFLD therapeutic targets andbiomarkers by conducting liver transcriptomic analysis in patients stratified by the presence of the PNPLA3 I148M genetic risk variant. DESIGN: We sequenced the hepatic transcriptome of 125 obese individuals. 'Severe NAFLD' was defined as the presence of steatohepatitis, NAFLD activity score ≥4 or fibrosis stage ≥2. The circulating levels of the most upregulated transcript, interleukin-32 (IL32), were measured by ELISA. RESULTS: Carriage of the PNPLA3 I148M variant correlated with the two major components of hepatic transcriptome variability and broadly influenced gene expression. In patients with severe NAFLD, there was an upregulation of inflammatory and lipid metabolism pathways. IL32 was the most robustly upregulated gene in the severe NAFLD group (adjusted p=1×10-6), and its expression correlated with steatosis severity, both in I148M variant carriers and non-carriers. In 77 severely obese, and in a replication cohort of 160 individuals evaluated at the hepatology service, circulating IL32 levels were associated with both NAFLD and severe NAFLD independently of aminotransferases (p<0.01 for both). A linear combination of IL32-ALT-AST showed a better performance than ALT-AST alone in NAFLD diagnosis (area under the curve=0.92 vs 0.81, p=5×10-5). CONCLUSION: Hepatic IL32 is overexpressed in NAFLD, correlates with hepatic fat and liver damage, and is detectable in the circulation, where it is independently associated with the presence and severity of NAFLD.

PNPLA3 I148M gene variant and chronic kidney disease in type 2 diabetic patients with NAFLD: Clinical and experimental findings.

BACKGROUND AND AIMS: Emerging evidence suggests an association between patatin-like phospholipase domain-containing protein-3 (PNPLA3) rs738409 (I148M protein variant) and risk of chronic kidney disease (CKD), but the mechanisms underpinning this association are poorly understood. METHODS: We studied 157 patients with type 2 diabetes (T2DM) who underwent ultrasonography and vibration-controlled transient elastography for diagnosing nonalcoholic fatty liver disease (NAFLD). CKD was defined as estimated glomerular filtration rate (e-GFR) <60 mL/min/1.73 m2 and/or abnormal albuminuria. We surveyed PNPLA3 mRNA expression in human tissues, using the liver as a positive control, and also measured PNPLA3 mRNA and protein expression levels in human cell lines represented in the kidney and the liver. RESULTS: In all, 112 patients had NAFLD and 43 had CKD. Patients homozygous for the I148M variant (n = 11) had lower e-GFR levels (60.6 ± 11.7 vs 77.8 ± 15.9 vs 83.5 ± 16.5 mL/min/1.73 m2 , P = .0001) and higher prevalence of CKD (63.6% vs 24.2% vs 25.0%, P = .028), compared to those with I/M (n = 66) and I/I (n = 80) PNPLA3 genotype. The association of I148M homozygosity with lower e-GFR levels (P < .0001) and higher risk of CKD (adjusted-odds ratio 6.65; 95% CI 1.65-26.8, P = .008) was independent of liver disease severity (as detected by liver stiffness ≥7kPa) and other risk factors. PNPLA3 mRNA expression was greatest in liver and renal cortex, and podocytes showed high PNPLA3 mRNA and protein levels, comparable to that of hepatocytes and hepatic stellate cells respectively. CONCLUSIONS: The PNPLA3 I148M variant was associated with CKD, independently of common renal risk factors and severity of NAFLD PNPLA3 expression levels were particularly high in renal podocytes.

NLRP3 promotes allergic responses to birch pollen extract in a model of intranasal sensitization.

Introduction & Objective: Allergic sensitization is an essential step in the development of allergic airway inflammation to birch pollen (BP); however, this process remains to be fully elucidated. Recent scientific advances have highlighted the importance of the allergen context. In this regard, microbial patterns (PAMPs) present on BP have attracted increasing interest. As these PAMPs are recognized by specialized pattern recognition receptors (PRRs), this study aims at investigating the roles of intracellular PRRs and the inflammasome regulator NLRP3. Methods: We established a physiologically relevant intranasal and adjuvant-free sensitization procedure to study BP-induced systemic and local lung inflammation. Results: Strikingly, BP-sensitized Nlrp3-deficient mice showed significantly lower IgE levels, Th2-associated cytokines, cell infiltration into the lung, mucin production and epithelial thickening than their wild-type counterparts, which appears to be independent of inflammasome formation. Intriguingly, bone-marrow chimera revealed that expression of NLRP3 in the hematopoietic system is required to trigger an allergic response. Conclusion: Overall, this study identifies NLRP3 as an important driver of BP-induced allergic immune responses.

Histone Deacetylase Inhibitors as a Therapeutic Strategy to Eliminate Neoplastic "Stromal" Cells from Giant Cell Tumors of Bone.

The neoplastic "stromal" cells in giant cell tumor of bone (GCTB) harbor a mutation in the H3F3A gene, which causes alterations in the epigenome. Current systemic targeted therapies, such as denosumab, do not affect the neoplastic cells, resulting in relapse upon treatment discontinuation. Therefore, this study examined whether targeting the epigenome could eliminate the neoplastic cells from GCTB. We established four novel cell lines of neoplastic "stromal" cells that expressed the H3F3A p.G34W mutation. These cell lines were used to perform an epigenetics compound screen (n = 128), which identified histone deacetylase (HDAC) inhibitors as key epigenetic regulators in the neoplastic cells. Transcriptome analysis revealed that the neoplastic cells expressed all HDAC isoforms, except for HDAC4. Therefore, five HDAC inhibitors targeting different HDAC subtypes were selected for further studies. All GCTB cell lines were very sensitive to HDAC inhibition in both 2D and 3D in vitro models, and inductions in histone acetylation, as well as apoptosis, were observed. Thus, HDAC inhibition may represent a promising therapeutic strategy to eliminate the neoplastic cells from GCTB lesions, which remains the paramount objective for GCTB patients who require life-long treatment with denosumab.

Novel Insights into the Genetic Landscape of Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD), the most common liver disorder worldwide, is epidemiologically associated with overweight, insulin resistance features and type 2 diabetes, and can progress to advanced liver fibrosis and hepatocellular carcinoma. Genetic factors play an important role in the development of NAFLD, which is a multifactorial disease. Several common naturally occurring variants modulating lipid and retinol metabolism in hepatocytes predispose to NAFLD development and progression, in particular those in PNPLA3, TM6SF2, MBOAT7, and HSD17B13. In addition, genetic variants that protect hepatic cells from oxidative stress modulate the susceptibility to progressive NAFLD. Although the molecular mechanisms linking these genetic variants with liver disease are not yet fully understood, hepatic fat has emerged as a major driver of the disease, while altered retinol metabolism and mitochondrial oxidative stress play a role in determining the development of advanced NAFLD.