Cytoprotective effect of low-dose tacrolimus on islets of Langerhans in cultures subjected to stimulation by acute rejection cytokines

Article information

Abstract

Introduction

The improvement in pancreatic islet transplantation results is due to immunosuppression protocols that include, among others, low-dose tacrolimus. Both antiinflammatory and anti-oxidant effects of tacrolimus could be useful in preventing primary rejection.

Aim

To evaluate in vitro islet low-dose tacrolimus response after pro-inflammatory stimulation.

Material and methods

Isolated rat islets were cultured in RPMI medium in the presence of IL-1 (50 UI/ml) plus IF-γ(1000UI/ml) and tacrolimus (5 ng/ml). The 24h production of lipoperoxide (LPO) and nitric oxide (NO) were measured as oxidative stress markers. Determination of apoptosis markers (nucleosome content and Bcl-2) was also performed.

Results

Oxidative stress (LPO 10.1±1.16 pmol/isletx24; NO 19.1±3.28 pmol/isletx24h) and apoptosis (nucleosome 0.24±0.04 UI/islet; Bcl-2 0.69±0.212 UI/islet) markers showed a very significant increase after cytokine stimulation (P<.01). Both effects improved by adding tacrolimus to the medium. Protective effect was complete when lipoperoxide (1.58 pmol/isletx24 h), nitric oxide (9.81 pmol/isletx24 h) and Bcl-2 (1.37±0.23 UI/islet) were determined.

Conclusion

In vitro cytoprotective effect of low-dose tacrolimus on isolated rat islets decreases both oxidative stress and apoptosis markers after stimulation of proinflammatory mediators.

Keywords:

Pancreatic islet transplantation

Apoptosis

Oxidative stress

Tacrolimus

Immunosuppression

Resumen

Introducción

La mejoría de los resultados en el trasplante de islotes pancreáticos se debe en gran parte a la introducción de nuevos protocolos de inmunosupresión que incluyen, entre otros, tacrolimus a bajas dosis. Este fármaco tiene efectos antioxidantes y antiapoptóticos que podrían ser de utilidad en la prevención del rechazo primario.

Objetivos

Evaluar la respuesta in vitro a tacrolimus a bajas dosis en islotes de rata estimulados con citocinas proinflamatorias implicadas en el rechazo primario de islotes.

Material y método

Se cultivaron islotes de rata en medio RPMI determinándose producción de lipoperóxido (LPO) y óxido nítrico (NO) y marcadores de apoptosis (nucleosomas y Bcl-2) en presencia de IL-1 (50UI/ml) e IF-γ(1000UI/ml) y adición de tacrolimus (FK-506; 5ng/ml).

Resultados

Tras la estimulación se apreció un aumento muy significativo (p < 0,01) de los marcadores de estrés oxidativo (LPO 10,1±1,16 pmol/islote x 24; NO 19,1±3,28 pmol/islote x 24 h) y apoptosis (nucleosomas 0,24±0,04; Bcl-2 0,69±0,212). Dichos efectos fueron contrarrestados de manera significativa tras añadir tacrolimus, siendo la reversión completa (p NS frente a controles) en el caso de la producción de lipoperóxidos (1,58 pmol/islote x 24h) y óxido nítrico (9,81 pmol/islote x 24h), así como en el descenso de Bcl-2 (1,37±0,23 UI/islote).

Conclusiones

El efecto citoprotector in vitro del tacrolimus a bajas dosis sobre islotes estimulados con citocinas proinflamatorias consigue aminorar la generación de estrés oxidativo y la activación de la apoptosis, habitualmente implicados en el rechazo en las primeras 48 h postimplante.

Palabras clave:

Trasplante de islotes pancreáticos

Apoptosis

Estrés oxidativo

Tacrolimus

Inmunosupresión

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