Responsive polymer‐assisted 3D cryogel supports Huh7.5 as in vitro hepatitis C virus model and ectopic human hepatic tissue in athymic mice. Issue 3 (25th December 2020)

• Record Type: Journal Article
• Authors: Jayal, Priyanka; Behera, Padmanava; Mullick, Ranajoy; Ramachandra, Subbaraya G.; Das, Saumitra; Kumar, Ashok; Karande, Anjali
• Is Part Of: Biotechnology and bioengineering. Volume 118:Issue 3(2021)
• Page Start: 1286
• Page End: 1304
• Publication Date: 2020-12-25
• Subjects: ectopic human liver -- hepatitis C virus -- hepatospheres -- pH/temperature responsive polymers -- polymeric cryogel; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.27651
• Languages: English
• ISSNs: 0006-3592

Abstract

Abstract: The three‐dimensional (3D) cell culture models serve as the interface between conventional two‐dimensional (2D) monolayer culture and animal models. 3D culture offers the best possible model system to understand the pathophysiology of human pathogens such as hepatitis C virus (HCV), which lacks a small animal model, due to narrow host tropism and non‐permissiveness of murine hepatocytes. In this study, functionally robust spheroids of HCV permissive Huh7.5 cells were generated, assisted by the temperature or pH‐responsive polymers PNIPAAm and Eudragit respectively, followed by the long‐term growth of the multilayered 3D aggregates in poly(ethylene glycol) (PEG)–alginate–gelatin (PAG) cryogel. The human serum albumin (HSA), marker of hepatic viability was detected up to 600 ng/ml on 24th day of culture. The 3D spheroid culture exhibited a distinct morphology and transcript levels with the upregulation of hepato‐specific transcripts, nuclear factor 4α (HNF4α), transthyretin (TTr), albumin (Alb), phase I and phase II drug‐metabolizing genes. The two most important phase I enzymes CYP3A4 and CYP2D6, together responsible for 90% metabolism of drugs exhibited up to 9‐ and 12‐fold increment, respectively in transcripts. The 3D culture was highly permissive to HCV infection and supported higher multiplicity of infection compared to monolayer Huh7.5 culture. Quantitation of high levels of HSA (500–200 ng/ml) in circulation in mice for 32 days asserted integration with host vasculature and in vivo establishment of 3D culture implants as an ectopic human hepatic tissue in mice. The study demonstrates the 3D spheroid Huh7.5 culture as a model for HCV studies and screening potential for anti‐HCV drug candidates. Abstract : A novel strategy combining thermoresponsive polymer aided spheroid generation with 3D culture in cryogel as a 3D hepatic model for Hepatitis C virus infection studies has been described here. The article explores the potential of the 3D hepatic culture for efficient propagation of HCV and assessment of the efficacy of neutralizing anti‐HCV antibody in vitro as well as an ectopic human hepatic tissue in athymic mice.