Pharmacodynamic monitoring by residual gene expression of the nuclear factor of activated T cell-regulated genes in lung transplant recipients and its correlation with tacrolimus blood levels.

Introduction: Trough blood levels (C0) of tacrolimus are used to adjust drug dosage, but they do not consistently correlate with clinical outcomes. Measurement of residual gene expression of nuclear factor of activated T cell (NFAT)-regulated genes (NFAT-RGE) has been proposed as a pharmacodynamic biomarker to assess the degree of immunosuppression in certain solid organ transplantations, but little is known regarding lung transplant recipients (LTR). Our primary objective is to correlate tacrolimus blood levels with NFAT-RGE. Methods: NFAT-RGE and tacrolimus C0 and peak (C1.5) levels were determined in 42 patients at three, six and 12 months post-transplantation. Results: Tacrolimus C0 did not exhibit a correlation with NFAT-RGE, whereas C1.5 did. Besides, over 20% of measurements indicated high levels of immunosuppression based on the below 30% NFAT-RGE threshold observed in many studies. Among those measurements within the therapeutic range, 19% had an NFAT-RGE<30%. Conclusion: Consequently, a subset of patients within the tacrolimus therapeutic range may be more susceptible to infection or cancer, potentially benefiting from NFAT-RGE and tacrolimus peak level monitoring to tailor their dosage. Further quantitative risk assessment studies are needed to elucidate the relationship between NFAT-RGE and the risk of infection, cancer, or rejection.

Lymphangioleiomyomatosis: Searching for potential biomarkers.

Background: Vascular endothelial growth factor-D (VEGF-D) is the most commonly used biomarker for diagnosing lymphangioleiomyomatosis (LAM). However, lung biopsy is often necessary as well; therefore, defining new biomarkers for LAM is crucial. The aim of this study was to describe the diagnostic accuracy of a variety of biomarkers. Methods: We assessed 13 analytes in serum related to extracellular matrix remodeling, lymphatic involvement and angiogenesis in a cohort of patients with LAM, comparing them with patients with other cystic lung diseases (OCLD) and healthy women. A scoring method based on the cut-point of each VEGF-D and metalloproteinase-2 (MMP-2) was used to evaluate the diagnostic performance of the marker combination. Results: A total of 97 subjects were recruited: 59 (61%) LAM patients, 18 (19%) OCLD patients, and 20 (20%) healthy female controls. MMP-2 was the only extracellular matrix remodeling biomarker able to differentiate LAM patients from OCLD and healthy patients. Serum MMP-2 was higher in LAM patients [median 578 (465-832) ng/ml] than in patients with OCLD and healthy controls [medians 360 (314-546) and 427 (365-513) ng/ml, respectively (p < 0.0001)]. The area under ROC curve (AUC) of MMP-2 was 0.785 and that of VEGF-D 0.815 (p = 0.6214). The sensitivity/specificity profiles of each biomarker (54/92% for MMP-2, 59/95% for VEGF-D) yielded a composite score (-6.36 + 0.0059 × VEGF-D + 0.0069 × MMP-2) with higher accuracy than each component alone (AUC 0.88 and sensitivity/specificity 79/87%). Conclusion: Combining MMP-2 and VEGF-D may increase diagnostic accuracy for LAM.

Pilot Trial of Extended Hypothermic Lung Preservation to Analyze Ischemia-reperfusion Injury in Pigs / Ensayo piloto de conservación pulmonar hipotérmica prolongada para analizar la lesión de isquemia-reperfusión en cerdos

Background: In lung transplantation (LT), the length of ischemia time is controversial as it was arbitrarily stablished. We ought to explore the impact of extended cold-ischemia time (CIT) on ischemia-reperfusion injury in an experimental model. Methods: Experimental, randomized pilot trial of parallel groups and final blind analysis using a swine model of LT. Donor animals (n=8) were submitted to organ procurement. Lungs were subjected to 6h (n=4) or 12h (n=4) aerobic hypothermic preservation. The left lung was transplanted and re-perfused for 4h. Lung biopsies were obtained at (i) the beginning of CIT, (ii) the end of CIT, (iii) 30min after reperfusion, and (iv) 4h after reperfusion. Lung-grafts were histologically assessed by microscopic lung injury score and wet-to-dry ratio. Inflammatory response was measured by determination of inflammatory cytokines. Caspase-3 activity was determined as apoptosis marker. Results: We observed no differences on lung injury score or wet-to-dry ratio any given time between lungs subjected to 6h-CIT or 12h-CIT. IL-1β and IL6 showed an upward trend during reperfusion in both groups. TNF-α was peaked within 30min of reperfusion. IFN-γ was hardly detected. Caspase-3 immunoexpression was graded semiquantitatively by the percentage of stained cells. Twenty percent of apoptotic cells were observed 30min after reperfusion. (AU)

Antecedentes: En el trasplante de pulmón (TP), la duración del tiempo de isquemia es controvertida, ya que se estableció de forma arbitraria. Sería útil explorar el impacto del tiempo de isquemia fría (TIF) prolongado sobre la lesión de isquemia-reperfusión en un modelo experimental. Métodos: Ensayo piloto experimental aleatorizado de grupos paralelos y análisis ciego final utilizando un modelo de TP en cerdos. Se extrajeron los órganos de los animales donantes (n=8). Los pulmones se conservaron durante 6 horas (n=4) o 12 horas (n=4) en hipotermia aeróbica. El pulmón izquierdo se trasplantó y reperfundió durante 4 horas. Se obtuvieron biopsias de pulmón (i) al comienzo del TIF, (ii) al final del TIF, (iii) 30 minutos después de la reperfusión y (iv) 4 horas después de la reperfusión. Los injertos de pulmón se evaluaron histológicamente mediante la puntuación de daño histológico pulmonar y la relación de peso húmedo y peso seco. La respuesta inflamatoria se valoró mediante la determinación de citoquinas inflamatorias. Se determinó la actividad de caspasa-3 como marcador de apoptosis. Resultados: No observamos diferencias en la puntuación de daño histológico pulmonar o en la relación de peso húmedo y peso seco en un momento dado entre los pulmones sometidos a 6 h-TIF o 12 h-TIF. Las IL-1β e IL-6 mostraron una tendencia ascendente durante la reperfusión en ambos grupos. El TNF-α alcanzó su punto máximo dentro de los 30 minutos posteriores a la reperfusión. Apenas se detectó IFN-γ. La inmunoexpresión de caspasa-3 se clasificó semicuantitativamente por el porcentaje de células teñidas. Se observó un 20% de células apoptóticas 30 minutos después de la reperfusión. (AU)

Pilot Trial of Extended Hypothermic Lung Preservation to Analyze Ischemia-reperfusion Injury in Pigs.

BACKGROUND: In lung transplantation (LT), the length of ischemia time is controversial as it was arbitrarily stablished. We ought to explore the impact of extended cold-ischemia time (CIT) on ischemia-reperfusion injury in an experimental model. METHODS: Experimental, randomized pilot trial of parallel groups and final blind analysis using a swine model of LT. Donor animals (n=8) were submitted to organ procurement. Lungs were subjected to 6h (n=4) or 12h (n=4) aerobic hypothermic preservation. The left lung was transplanted and re-perfused for 4h. Lung biopsies were obtained at (i) the beginning of CIT, (ii) the end of CIT, (iii) 30min after reperfusion, and (iv) 4h after reperfusion. Lung-grafts were histologically assessed by microscopic lung injury score and wet-to-dry ratio. Inflammatory response was measured by determination of inflammatory cytokines. Caspase-3 activity was determined as apoptosis marker. RESULTS: We observed no differences on lung injury score or wet-to-dry ratio any given time between lungs subjected to 6h-CIT or 12h-CIT. IL-1ß and IL6 showed an upward trend during reperfusion in both groups. TNF-α was peaked within 30min of reperfusion. IFN-γ was hardly detected. Caspase-3 immunoexpression was graded semiquantitatively by the percentage of stained cells. Twenty percent of apoptotic cells were observed 30min after reperfusion. CONCLUSIONS: We observed that 6 and 12h of CIT were equivalent in terms of microscopic lung injury, inflammatory profile and apoptosis in a LT swine model. The extent of lung injury measured by microscopic lung injury score, proinflammatory cytokines and caspase-3 determination was mild.

Pilot Trial of Extended Hypothermic Lung Preservation to Analyze Ischemia-reperfusion Injury in Pigs.

BACKGROUND: In lung transplantation (LT), the length of ischemia time is controversial as it was arbitrarily stablished. We ought to explore the impact of extended cold-ischemia time (CIT) on ischemia-reperfusion injury in an experimental model. METHODS: Experimental, randomized pilot trial of parallel groups and final blind analysis using a swine model of LT. Donor animals (n=8) were submitted to organ procurement. Lungs were subjected to 6h (n=4) or 12h (n=4) aerobic hypothermic preservation. The left lung was transplanted and re-perfused for 4h. Lung biopsies were obtained at (i) the beginning of CIT, (ii) the end of CIT, (iii) 30min after reperfusion, and (iv) 4h after reperfusion. Lung-grafts were histologically assessed by microscopic lung injury score and wet-to-dry ratio. Inflammatory response was measured by determination of inflammatory cytokines. Caspase-3 activity was determined as apoptosis marker. RESULTS: We observed no differences on lung injury score or wet-to-dry ratio any given time between lungs subjected to 6h-CIT or 12h-CIT. IL-1ß and IL6 showed an upward trend during reperfusion in both groups. TNF-α was peaked within 30min of reperfusion. IFN-γ was hardly detected. Caspase-3 immunoexpression was graded semiquantitatively by the percentage of stained cells. Twenty percent of apoptotic cells were observed 30min after reperfusion. CONCLUSIONS: We observed that 6 and 12h of CIT were equivalent in terms of microscopic lung injury, inflammatory profile and apoptosis in a LT swine model. The extent of lung injury measured by microscopic lung injury score, proinflammatory cytokines and caspase-3 determination was mild.