Elsevier

Transplant Immunology

Volume 32, Issue 3, June 2015, Pages 188-194
Transplant Immunology

Elevated level of HSPA1L mRNA correlates with graft-versus-host disease

https://doi.org/10.1016/j.trim.2015.02.002Get rights and content

Highlights

  • HSPA1L mRNA expression at Day + 28 is predictive of chronic GVHD.

  • Lower HSPA1L mRNA is expressed at the time of skin acute GVHD onset.

  • HSPA1L mRNA expression is significantly varied between grade I and grade II–III skin histopathological acute GVHD.

  • HSPA1B and HSPA1L mRNAs are both up-regulated in patients with severe acute GVHD (grades II–III).

Abstract

Graft-versus-host disease (GVHD) can be a fatal complication of allogeneic stem cell transplantation (allo-HSCT). GVHD can be classified as acute (aGVHD: up to 100 days) or chronic (cGVHD: after 100 days) based on the time-point of disease occurrence. At present there are a limited number of biomarkers available for use in the clinic. Thus, the aim of this research was to evaluate the biomarker potential of the extensively studied Heat Shock Protein 70 family members (HSPA1A/HSPA1B and HSPA1L) at the messenger RNA (mRNA) level in acute and cGVHD patient cohorts. In the skin biopsies, HSPA1L mRNA expression was lower in patients with severe aGVHD (grades II–III) when compared to those with none or low grade aGVHD (grades 0–I) and normal controls. In whole blood, HSPA1L mRNA expression level was significantly (p = 0.008) up-regulated at 28 days post-transplant in cGVHD patients with a significant area under the curve (AUC = 0.773). In addition, HSPA1B expression in whole blood was significantly higher at 3 months post-transplant in both the aGVHD grade II–III (p = 0.012) and cGVHD (p = 0.027) patients. Our initial results in this small cohort show that quantifying HSPA1L mRNA expression in the whole blood of allo-HSCT patients at day 28 post-allo-HSCT may be a useful predictive biomarker for cGVHD.

Introduction

Allogeneic-HSCT is used to cure numerous malignant and non-malignant haematological disorders [43]. More than 25,000 allo-HSCTs are performed annually world-wide [33]. However, GVHD still remains a major risk factor for allo-HSCT; directly contributing to a transplant related mortality rate of 26% for matched sibling and 35% for matched unrelated donor transplants [34]. Despite all the improvements achieved to date with respect to treatment and patient after-care, there are a limited number of validated biomarkers for GVHD, in particular for cGVHD [23], [32].

Heat Shock Proteins (HSPs) are highly conserved and ubiquitously present in all organisms [50]. Various physiological roles have been attributed to circulating HSPs such as (i) molecular chaperones [14], (ii) down-regulation of autoimmune responses [3], (iii) down-regulation of T cell responses [47] and (iv) cytoprotection [25], [27]. As molecular chaperones, they play a role in protein folding, repair and maintenance of cellular homeostasis [14], [50]. Under normal conditions HSPs comprise only 5% of the total intracellular proteins; however, under stress, the level elevates to approximately 15% [45]. Heat Shock Protein 70 family (HSPA) is induced by different stimuli such as: heat [41], metal ions and oxidative stress [51], infection [30], inflammation [40] and anti-cancer drugs [24]. Investigations have exhibited HSPs on the surface of cells [28], in sera [37] and in secretions from viable cells [19].

Heat Shock Proteins are characterised according to their molecular weight and their role has been investigated in a variety of diseases [29], [52]. The HSPA family comprises of thirteen members [10]; two members in this investigation belong to the heat inducible group; HSPA1A (HSP70-1) and HSPA1B (HSP70-2), and the third constitutively expressed member; HSPA1L (HSP70-hom). The two heat inducible members are collectively known as HSP0i [22]. The exons of HSPA1A and HSPA1B vary by only six single bases [44], which generates a difference of two amino-acids in the protein sequences [22].

Heat Shock Proteins have been shown to be associated with responses to inflammation and allograft rejection [20], [48]. HSPAs have also exhibited crucial functions during stress and in prediction of allo-HSCT outcome [12]. HSPA1B protein has been shown to be correlated to severity of graft-versus-host reactivity (GVHR) in the in vitro skin explant model [20].

Section snippets

Objective

The overall objective of this study was to test whether there was a significant differential HSPA1L, HSPA1B and/or HSPA1A mRNA expression level in clinical skin biopsies and whole blood from patients with or without aGVHD and cGVHD that could predict disease incidence and/or severity prior to GVHD onset.

Ethics and consent

The study was approved by the Newcastle and North Tyneside Research Ethics Committee I, and all patients and normal volunteers gave their written informed consent for the collection of both skin and whole blood specimens as well as for molecular testing. The investigations were conducted in accordance with the Helsinki Declaration, and consents were taken prospectively by trained personnel. Both aGVHD and cGVHD were assessed by clinicians according to the NIH consensus. All the clinical data

Patients with histopathological skin aGVHD grades II–III had significantly lower HSPA1L mRNA levels

Quantitative RT-PCR was used to investigate HSPA1L, HSPA1B and HSPA1A mRNA expression levels in clinical skin biopsies obtained from allo-HSCT patients with and without aGVHD at various time-points post-transplant at the onset of aGVHD. The histopathological grading was used to analyse the HSPA expressions. Interestingly, HSPA1L expression was significantly lower in patients with grade II–III aGVHD when compared to those without aGVHD (p  0.05) and normal controls (p  0.01). As expected, there

Discussion

Even though the field of allo-HSCT has advanced progressively, GVHD is still a major post-transplant complication. There are numerous risk factors that predispose patients to GVHD. However, the incidence of prior aGVHD can increase the likelihood to cGVHD [4], [5], [7]. In this investigation we have looked at both acute and cGVHD patients with regard to HSPA1L, HSPA1B and HSPA1A expression in clinical skin biopsies and in whole blood samples. The potential role of HSP0i in initiation and GVHD

Funding

This investigation was supported by the European Commission Marie Curie Research Training Network MCRTN-TRANSNET (Contract no. 512253), FP6 EC Stemdiagnostics (Contract no. 037703) and the FP7 Marie Curie Initial Training Network CELLEurope (Contract No: 315963).

Disclosure

The authors declare no conflicts of interest.

Author's contributions

SA: performed the research and data analysis and wrote the manuscript.

BT, JN, SA, KBK, XNW and AD: designed the study and commented on the manuscript.

AD, JN and KFP: contributed to data interpretation.

KFP: contributed to statistical analysis.

MC: contributed clinical samples for the study.

Acknowledgements

The authors thank the patients who have donated samples for this study. The authors also thank E. Douglas and L.J. Ambler for their technical support.

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