Increased Renal Expression of Complement Components in Patients With Liver Diseases: Nonalcoholic Steatohepatitis, Alcohol-Associated, Viral Hepatitis, and Alcohol-Viral Combination

Abstract

Inflammatory liver diseases, including nonalcoholic steatohepatitis (NASH), alcohol-associated liver disease (ALD), hepatitis C virus (HCV), and ALD/HCV, account for nearly 2 million deaths annually. Despite increasing evidence that liver dysfunction impacts renal physiology, there is limited supportive clinical information, due to limited diagnosis of liver disease, complexity in liver disease etiology, and inadequacy of renal function tests. Human kidney biopsies with liver and renal pathology were obtained from patients with nonalcoholic fatty liver disease (NAFLD), NASH, ALD, HCV, and ALD/HCV (n = 5-7). Each liver disease showed renal pathology with at least 50% interstitial nephritis, 50% interstitial fibrosis, and renal dysfunction by estimated glomerular filtration rate (eGFR) (NAFLD 36.7 ± 21.4; NASH 32.7 ± 15.0; ALD 16.0 ± 11.0; HCV 27.6 ± 11.5; ALD/HCV 21.0 ± 11.2 mL/min/1.73m2). Transcriptomic analysis identified 55 genes with expression changes in a conserved direction in response to liver disease. Considering association with immune regulation, protein levels of alpha-2-macroglobulin (A2M), clusterin (CLU), complement C1q C chain (C1QC), CD163, and joining chain of multimeric IgA and IgM (JCHAIN) were further quantified by LC-MS/MS. C1QC demonstrated an increase in NASH, ALD, HCV, and ALD/HCV (42.9 ± 16.6; 38.8 ± 18.4; 39.0 ± 13.5; 40.1 ± 20.1 pmol/mg protein) relative to control (19.2 ± 10.4 pmol/mg protein; p ≤ 0.08). Renal expression changes identified in inflammatory liver diseases with interstitial pathology suggest the pathogenesis of liver associated renal dysfunction. This unique cohort overcomes diagnostic discrepancies and sample availability to provide insight for mechanistic investigations on the impact of liver dysfunction on renal physiology.

Renal Transporter Alterations in Patients with Chronic Liver Diseases: Nonalcoholic Steatohepatitis, Alcohol-Associated, Viral Hepatitis, and Alcohol-Viral Combination

“…Liquid chromatography-coupled mass spectrometry (LC-MS/MS) instrumental parameters for surrogate peptide quantification are consistent with previously published methods, apart from injection volume, which was increased to 10 µL (Frost et al, 2022). A binary solvent gradient (water + 0.1% formic acid aqueous; 90:10 ACN: water with 0.1% formic acid organic) was run on an Agilent UPLC system using a Acquity UPLC HSS C18 column (2.1 mm X 100 mm) with 1.8 µm particles (Waters, Inc.).…”

“…Renal biopsies were obtained from Banner University Medical Center as previously described by Frost et al (Frost et al, 2022). In brief, medical charts of all patients with available renal biopsies that contained International Classification of Disease 9 and 10 codes (ICD9, ICD10) for liver pathology were identified.…”

“…Control patients (n = 7) were uniquely classified throughout the chart review process and deemed normal upon renal pathology and abdominal imaging assessment. Demographics and biochemical function tests are available in previously published materials (Frost et al, 2022) and briefly summarized in Table 1.…”

“…Transcriptional expression was determined by Affymetrix Clariom D arrays according to published protocols established by Affymetrix. Detailed methods of RNA isolation and microarray expression profiling were previously published by Frost et al (Frost et al, 2022). In summary, formalinfixed paraffin-embedded (FFPE) renal needle biopsy specimens were gouged, and half of the sample proceeded with RNA isolation and quality control checks.…”

“…The remaining half of the gouged FFPE renal biopsy was deparaffinized and hydrated prior to heat-induced antigen retrieval (HIAR) as previously published (Frost et al, 2022). Of the 36 specimens, two biopsies did not have enough tissue leftover from transcriptomics to proceed with protein processing and were excluded from analysis (1 ALD and 1 ALD/HCV).…”