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Biology subjects

Pulitano, C.

Publications and source records attributed to Pulitano, C..

2 recordsLinked to original sources

Development and Validation of a Continuous Real-Time Optical Sensor for Indocyanine Green Clearance Measurement During Ex-Vivo Perfusion of Human Livers

Liver transplantation remains the only curative treatment for end-stage liver failure, yet its impact is constrained by organ shortages and graft non-utilisation. Machine perfusion (MP) enables ex-vivo assessment of donated livers; however, existing viability criteria rely on intermittent sampling, reducing temporal resolution and accuracy. Indocyanine green (ICG), a clinically validated dye cleared exclusively by hepatocytes, provides a continuous index of hepatic function beyond initial injury. Accordingly, we present a non-invasive, clamp-on optical sensor that enables continuous, real-time quantification of ICG clearance during MP. The sensor consists of a clamp-on module with an 808nm laser and phototransistor connected to a microcontroller-based unit and computer for real-time plotting. The raw phototransistor signal was linearised to a unitless absorbance signal proportional to perfusate ICG; bi-exponential fitting yielded plasma disappearance rate (PDRbi, %/min) and the 15-minute residual fraction (R15). Across 10 whole and 3 split human livers (45 boluses; 13 paired with spectrophotometry), the sensor closely matched spectrophotometric measurements (mean R2 = 0.994; range 0.983-0.999). The sensor resolved expected physiological trends: ICG clearance increased with temperature (PDRbi: 8.2%/min (subnormothermic MP) to 22.6%/min (normothermic MP) (n=4); 9.3%/min (32{degrees}C) to 11.9%/min (36{degrees}C) (n=1)). The sensors continuous signal traces also revealed early mixing dynamics and medication-related effects that are missed by intermittent sampling. This optical sensor enables accurate, real-time monitoring of ICG clearance during ex-vivo perfusion. The ex-vivo setting is uniquely positioned to validate ICG clearance models and enhance clinical interpretation.

bioengineering↗

Harnessing the power of whole human liver ex situ normothermic perfusion for preclinical AAV vector evaluation

Developing clinically predictive model systems for evaluating gene transfer and gene editing technologies has become increasingly important in the era of personalized medicine. Liver-directed gene therapies present a unique challenge due to the complexity of the human liver. In this work, we describe the application of whole human liver explants in an ex situ normothermic perfusion system to evaluate a set of fourteen natural and bioengineered adeno-associated viral (AAV) vectors directly in human liver, in the presence and absence of neutralizing human sera. Under non-neutralizing conditions, the recently developed AAV variants, AAV-SYD12 and AAV-LK03, emerged as the most functional variants in terms of cellular uptake and transgene expression. However, when assessed in the presence of human plasma containing anti-AAV neutralizing antibodies (NAbs), vectors of human origin, specifically those derived from AAV2/AAV3b, were extensively neutralized, whereas AAV8-derived variants performed efficiently. This study establishes the use of normothermic liver perfusion as an invaluable preclinical model for evaluating liver-targeted gene therapies and providing guidance for making essential decisions that promote the most effective translational programs.

molecular biology↗