Increase in Proportion of Th-17 Lymphocytes in Blood Heralds Overt Acute Graft Versus Host Disease in HSCT Patients.

IL-17 is involved in chronic inflammation and autoimmunity, this cytokine producing cells are well characterized in the mouse model. In man, situation is more complex due to the genetic variability, co-morbidities and environmental factors influencing the immune response. In patients post HSCT alloreactivity with the presence of aGvHD is a major factor affecting the outcome of transplantation. Understanding of the role of cells involved in regulation of the immune system is crucial for the treatment tailoring to favour tolerance but not making the recipient more prone to opportunistic infections and leukaemia relapse.

In this study 27 patients post HSCT were investigated for the presence of regulatory cells and those producing IFNgamma and IL-17 in blood. Blood was collected at the time of haematological recovery or when the first clinical symptoms of aGvHD became apparent and then in one week intervals until +60 day post transplantation. PBMC were stimulated for 4 hrs with BD Leukocyte Activation Cocktail in the presence of Golgi Stop (BD, Erembodegen, Belgium) and then stained extracellularly with anti-CD4, permeabilized with Fixation/Permeabilization Concentrate and Diluent (eBioscience, San Diego, CA) and finally stained with anti-IFNgamma (BD), anti-IL-17 (eBioscience) anti FoxP3 (eBioscience). CD4+ cells subpopulations were analysed according to the expression of the stained features.

It was found:

FoxP3+CD4+ cells were in higher proportions in pts with aGvHD (results from all time-points taken together) (9.93%±0.61 vs 8.2%±0.50, n=98, p=0.040, U Mann-Whitney test)

IFNgamma producing CD4+ lymphocytes were in higher proportions (0,34 vs 0.14, ns) in blood samples taken from patients lacking as compared to those having aGvHD.

CD4+IL-17+ lymphocytes proportions increased from 1.39%±0.42 to 5.33%±2.45 (p=0.04, Wilcoxon Test for pairs) one week before aGvHD.

Notably, at the time of full blown aGvHD the proportions of CD4+IL-17+ cells were lower as compared to the results of previous measurements (0.74%±0.29, p=0.008, Wilcoxon Test for pairs). Taking all results together the proportion of CD4+IL17+ lymphocytes were lower in patients having as compared to those lacking aGVHD (0.93%±0.27 vs 1,53%±0,41, p=0.05, U Mann-Whitney test).

It appears that:

FoxP3 positive cells expand during aGvHD likely as a response to alloreactive stimulation.

INFgamma+ CD4+ cells benefit the course post HSCT making the patients less susceptible to aGvHD.

CD4+IL17+ cells are likely involved at the early stage of aGvHD patho-mechanism heralding the clinical manifestation of this complication, but then they disappear from blood, possibly being marginalized in the inflamed tissues.