International Journal of Laboratory Hematology The Official journal of the International Society for Laboratory Hematology
ORIGINAL ARTICLE
INTERNAT IONAL JOURNAL OF LABORATO RY HEMATO LOGY
Elevated serum adenosine deaminase levels in secondary hemophagocytic lymphohistiocytosis W. CHEN* , † , a , S. ZHANG † ,a , W. ZHANG † , X. YANG † , J. XU † , H. QIU † , X. ZHANG † , J. LI †
*Department of Hematology, the Affiliated jiangyin Hospital of Southeast University Medical College, Jiangyin, Jiangsu Province, China † Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, Nanjing, Jiangsu Province, China Correspondence: Hongxia Qiu, Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, 300 Guangzhou Road, Nanjing, Jiangsu 210029, China. Tel.: 86 25 68136091; Fax: 86 25 86306167; E-mail: [email protected] and Xiaoyan Zhang, Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, 300 Guangzhou Road, Nanjing, Jiangsu 210029, China. Tel.: 86 25 68135681; Fax: 86 25 86306167; E-mail: [email protected]
a
Weifeng Chen and Sujiang Zhang contributed equally to this work and should be considered as co-first author.
S U M M A RY Introduction: Hemophagocytic lymphohistiocytosis (HLH) is a severe hyperinflammatory condition caused by activated T cells and macrophages. Adenosine deaminase (ADA) plays an important role in immune regulation, especially in the proliferation, maturation, and differentiation of lymphocytes. Its role has been studied in inflammation and malignancy. Here, we present for the first time the alteration of ADA in secondary HLH (sHLH). Methods: Serum ADA levels were measured in 20 cases with lymphoma-associated hemophagocytic syndrome (LAHS), 15 with infection-associated hemophagocytic syndrome (IAHS), six with macrophage activation syndrome (MAS) as experimental group. Additionally, we enrolled 20 cases with lymphoma, 15 with infection, six with autoimmunity who were all not associated with HLH as conditional control group and 20 healthy subjects as blank group. We also demonstrated the ADA levels in 20 LAHS cases and 21 benign disease-associated HLH cases (IAHS and MAS). Results: Serum ADA levels were significantly higher in patients with LAHS, IAHS, and MAS compared to the healthy subjects (P < 0.001) and conditional control group (P < 0.05). Serum ADA levels of patients with LAHS were significantly higher than benign disease-associated HLH cases (P < 0.05). The optimum ADA cutoff point for LAHS was 89.25 U/L, with a sensitivity and specificity of 85.0% and 76.2%, respectively. Conclusion: Serum ADA levels were increased in sHLH suggesting a partial role of activated T-cell response in the disease pathophysiology. Serum ADA levels were particularly higher in LAHS and probably be a potential indicator of underlying lymphoma in sHLH patients.
doi:10.1111/ijlh.12334
Received 10 August 2014; accepted for publication 13 January 2015
544
© 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
W. CHEN ET AL. | ELEVATED ADA LEVELS IN SHLH
545
Keywords Adenosine deaminase, hemophagocytic lymphohistiocytosis, lymphoma, infection, autoimmune diseases
INTRODUCTION Hemophagocytic lymphohistiocytosis (HLH) represents a potentially fatal hyperinflammatory condition caused by activated T cells and macrophages [1]. It is characterized by prolonged fever, hepatosplenomegaly, cytopenia, liver dysfunction, coagulopathy, and central nervous system (CNS) involvement. It can be classified as two forms: familiar HLH (FHLH) and secondary HLH (sHLH). FHLH is an autosomal recessive inheritance disease and usually affects infants. sHLH generally refers to older children (or adults) who present without a family history or known genetic cause for their HLH and is associated with malignancy, infection, and autoimmunity [2]. According to the diagnostic criteria revised by the Histocyte Society in 2004 (HLH-2004) [3], two inflammatory markers – soluble CD25 (sCD25, also known as the a-chain of the IL-2 receptor) and serum ferritin (SF) – are valuable markers as they correlate with the disease activity [2]. It is suggested that delayed diagnosis, multiorgan failure, and CNS manifestation are poor prognostic indicators for HLH [4]. Early diagnosis is very difficult because of the diversity of clinical manifestation and laboratory examination. As soluble CD25 is not readily available in all institution, we should search some more simple and easy methods to aid the diagnosis of the disease. Adenosine deaminase (ADA) is a polymorphic enzyme involved in purine metabolism and catalyzes the deamination of both adenosine and 20 -deoxyadenosine to inosine and 20 -deoxyinosine, respectively [5]. ADA is widely distributed in tissues and body fluids. It is involved in the proliferation, maturation, and function of lymphocytes, especially for T cells [6]. ADA level varies during T-cell differentiation and has been considered as a nonspecific marker of T-cell activation [7]. ADA has been found to be increased in diseases characterized by T-cell proliferation and activation such as ulcerative colitis, tuberculosis, psoriasis, etc [8–10].
© 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
With this respect, we undertook this study to investigate whether serum ADA level alters in sHLH patients, especially in lymphoma-associated HLH (LAHS). To the best of our knowledge, we present here the first study to investigate the serum ADA level in sHLH patients. We consider that our study is important because it provides a new supportive marker for suspected sHLH.
M AT E R I A L S A N D M E T H O D S Patients and diagnosis Forty-one patients with sHLH attending at the Department of Hematology in First Affiliated Hospital of Nanjing Medical University between January 2009 and April 2012 were enrolled in the study, excluding patients younger than 16 years old. All patients met the HLH-2004 diagnostic criteria [3]. Patients were required to fulfill five of the following eight criteria: (i) fever; (ii) splenomegaly; (iii) cytopenia involving ≥2 of 3 lineages in the peripheral blood: hemoglobin 0.05 >0.05
20 8/12 40.05 17.71
20 7/13 50.00 13.03
7 4 4 5 15 7/8 43.00 20.27
6 4 3 7 15 7/8 46.40 20.16
7 3 5 6 4/2 37.50 16.48
5 4 6 6 3/3 38.50 9.59
2 4
>0.05 >0.05 >0.05
>0.05 >0.05 >0.05
>0.05 >0.05 >0.05
3 3
Data were shown in form of mean SD.
with a sensitivity and specificity of 85.0% and 76.2%, respectively (Figure 2).
DISCUSSION In our study, serum ADA levels in sHLH were significantly higher than healthy donors and patients in conditional control group. sHLH is a severe hyperinflammatory condition, which is characterized by prolonged fever, cytopenia, liver dysfunction, hepatosplenomegaly, and hemophagocytosis, caused by activated macrophages. The main pathophysiology of sHLH is hypercytokinemia resulting in hyperactivation of T lymphocytes and macrophages, and proliferation and infiltration of these cells into various organs [12]. The immune system is regulated by T-helper cells (Th), which is classified as type 1 Th-cells (Th1) and © 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
Table 2. ADA levels in sHLH groups and healthy control subjects Study groups
Serum ADA (U/L)
sHLH LAHS (n = 20) IAHS (n = 15) MAS (n = 6) Healthy subjects (n = 20)
115.12 150.29 91.33 57.40 12.36
69.17* 76.08* 44.38* 11.30* 4.07
*Compare with healthy subjects, P < 0.001. All data were shown in form of mean SD.
type 2 Th-cells (Th2). According to different cell types, the immune system will develop different immune responses. Th1 cell subset mainly secrets IL-2, IL-12, interferon (IFN)-c and tumor necrosis factor (TNF)-b and mediates cell immunity. Th2 cell subset mainly
548
W. CHEN ET AL. | ELEVATED ADA LEVELS IN SHLH
Figure 1. Compared ADA level between experimental group and conditional control group. The error bars represent standard deviation.
secrets IL-4, IL-5, and IL-9 and mediates humoral immunity [13]. Osugi et al. [14] and Tang et al. [15] have proposed that IFN-c, which is a Th1 type cytokine with immunomodulatory properties, was significantly higher during acute phase of sHLH in comparison with those after remission and healthy volunteers. IL-4, which is a Th2 type cytokine, however, was not significantly different between them. Janka [16] has shown that Th1 cytokines such as IFNc, IL-12, and IL-18, which correlate with disease activity were obviously decreased after disease remission. These all indicate that sHLH shows a Th1 polarization. Adenosine deaminase is an enzyme involved in purine catabolism and plays a role in proliferation,
Figure 2. ROC curve analysis of patients with LAHS vs. benign disease-associated HLH (IAHS and MAS).
maturation, and differentiation of lymphocytes, and its level increases during mitogenic and antigenic responses of these cells [8, 17]. Patients with deficiency of ADA develop severe defects in humoral and cellular immunity, which is called severe combined immunodeficiency (ADA-SCID) [18]. In the course of T lymphocyte activation and secretion of IFN-c, TNFa, and IL-6, ADA interacts with CD26 to produce a costimulatory signal. During this activation, ADA activity is regulated positively by IL-2 and IL-12 and negatively by IL-4 [19]. Elevated ADA level may be a part of the activated Th1 response [20] and has been reported in several diseases, which are characterized by T-cell activation [7–10].
Table 3. Compared laboratory findings between LAHS and benign disease-associated HLH (IAHS and MAS) Laboratory finding
LAHS
PLT (9109/L) Neu (9109/L) Hb (9109/L) SF (lg/L) FIB (g/L) TG (mM) ALT (U/L) AST (U/L) ADA (U/L)
48.95 1.62 95.50 35828.90 1.58 3.06 262.47 366.69 150.29
Benign disease-associated HLH
41.54 1.34 19.14 20889.11 1.03 1.83 244.66 306.18 76.08
56.43 2.19 99.81 32571.14 2.00 2.38 197.47 326.72 81.63
35.68 1.90 25.29 19225.79 1.01 1.46 150.43 296.78 40.71
P value >0.05 >0.05 >0.05 >0.05 >0.05 >0.05 >0.05 >0.05
ORIGINAL ARTICLE
INTERNAT IONAL JOURNAL OF LABORATO RY HEMATO LOGY
Elevated serum adenosine deaminase levels in secondary hemophagocytic lymphohistiocytosis W. CHEN* , † , a , S. ZHANG † ,a , W. ZHANG † , X. YANG † , J. XU † , H. QIU † , X. ZHANG † , J. LI †
*Department of Hematology, the Affiliated jiangyin Hospital of Southeast University Medical College, Jiangyin, Jiangsu Province, China † Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, Nanjing, Jiangsu Province, China Correspondence: Hongxia Qiu, Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, 300 Guangzhou Road, Nanjing, Jiangsu 210029, China. Tel.: 86 25 68136091; Fax: 86 25 86306167; E-mail: [email protected] and Xiaoyan Zhang, Department of Hematology, the First Affiliated Hospital of Nanjing Medical University, Jiangsu Province Hospital, 300 Guangzhou Road, Nanjing, Jiangsu 210029, China. Tel.: 86 25 68135681; Fax: 86 25 86306167; E-mail: [email protected]
a
Weifeng Chen and Sujiang Zhang contributed equally to this work and should be considered as co-first author.
S U M M A RY Introduction: Hemophagocytic lymphohistiocytosis (HLH) is a severe hyperinflammatory condition caused by activated T cells and macrophages. Adenosine deaminase (ADA) plays an important role in immune regulation, especially in the proliferation, maturation, and differentiation of lymphocytes. Its role has been studied in inflammation and malignancy. Here, we present for the first time the alteration of ADA in secondary HLH (sHLH). Methods: Serum ADA levels were measured in 20 cases with lymphoma-associated hemophagocytic syndrome (LAHS), 15 with infection-associated hemophagocytic syndrome (IAHS), six with macrophage activation syndrome (MAS) as experimental group. Additionally, we enrolled 20 cases with lymphoma, 15 with infection, six with autoimmunity who were all not associated with HLH as conditional control group and 20 healthy subjects as blank group. We also demonstrated the ADA levels in 20 LAHS cases and 21 benign disease-associated HLH cases (IAHS and MAS). Results: Serum ADA levels were significantly higher in patients with LAHS, IAHS, and MAS compared to the healthy subjects (P < 0.001) and conditional control group (P < 0.05). Serum ADA levels of patients with LAHS were significantly higher than benign disease-associated HLH cases (P < 0.05). The optimum ADA cutoff point for LAHS was 89.25 U/L, with a sensitivity and specificity of 85.0% and 76.2%, respectively. Conclusion: Serum ADA levels were increased in sHLH suggesting a partial role of activated T-cell response in the disease pathophysiology. Serum ADA levels were particularly higher in LAHS and probably be a potential indicator of underlying lymphoma in sHLH patients.
doi:10.1111/ijlh.12334
Received 10 August 2014; accepted for publication 13 January 2015
544
© 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
W. CHEN ET AL. | ELEVATED ADA LEVELS IN SHLH
545
Keywords Adenosine deaminase, hemophagocytic lymphohistiocytosis, lymphoma, infection, autoimmune diseases
INTRODUCTION Hemophagocytic lymphohistiocytosis (HLH) represents a potentially fatal hyperinflammatory condition caused by activated T cells and macrophages [1]. It is characterized by prolonged fever, hepatosplenomegaly, cytopenia, liver dysfunction, coagulopathy, and central nervous system (CNS) involvement. It can be classified as two forms: familiar HLH (FHLH) and secondary HLH (sHLH). FHLH is an autosomal recessive inheritance disease and usually affects infants. sHLH generally refers to older children (or adults) who present without a family history or known genetic cause for their HLH and is associated with malignancy, infection, and autoimmunity [2]. According to the diagnostic criteria revised by the Histocyte Society in 2004 (HLH-2004) [3], two inflammatory markers – soluble CD25 (sCD25, also known as the a-chain of the IL-2 receptor) and serum ferritin (SF) – are valuable markers as they correlate with the disease activity [2]. It is suggested that delayed diagnosis, multiorgan failure, and CNS manifestation are poor prognostic indicators for HLH [4]. Early diagnosis is very difficult because of the diversity of clinical manifestation and laboratory examination. As soluble CD25 is not readily available in all institution, we should search some more simple and easy methods to aid the diagnosis of the disease. Adenosine deaminase (ADA) is a polymorphic enzyme involved in purine metabolism and catalyzes the deamination of both adenosine and 20 -deoxyadenosine to inosine and 20 -deoxyinosine, respectively [5]. ADA is widely distributed in tissues and body fluids. It is involved in the proliferation, maturation, and function of lymphocytes, especially for T cells [6]. ADA level varies during T-cell differentiation and has been considered as a nonspecific marker of T-cell activation [7]. ADA has been found to be increased in diseases characterized by T-cell proliferation and activation such as ulcerative colitis, tuberculosis, psoriasis, etc [8–10].
© 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
With this respect, we undertook this study to investigate whether serum ADA level alters in sHLH patients, especially in lymphoma-associated HLH (LAHS). To the best of our knowledge, we present here the first study to investigate the serum ADA level in sHLH patients. We consider that our study is important because it provides a new supportive marker for suspected sHLH.
M AT E R I A L S A N D M E T H O D S Patients and diagnosis Forty-one patients with sHLH attending at the Department of Hematology in First Affiliated Hospital of Nanjing Medical University between January 2009 and April 2012 were enrolled in the study, excluding patients younger than 16 years old. All patients met the HLH-2004 diagnostic criteria [3]. Patients were required to fulfill five of the following eight criteria: (i) fever; (ii) splenomegaly; (iii) cytopenia involving ≥2 of 3 lineages in the peripheral blood: hemoglobin 0.05 >0.05
20 8/12 40.05 17.71
20 7/13 50.00 13.03
7 4 4 5 15 7/8 43.00 20.27
6 4 3 7 15 7/8 46.40 20.16
7 3 5 6 4/2 37.50 16.48
5 4 6 6 3/3 38.50 9.59
2 4
>0.05 >0.05 >0.05
>0.05 >0.05 >0.05
>0.05 >0.05 >0.05
3 3
Data were shown in form of mean SD.
with a sensitivity and specificity of 85.0% and 76.2%, respectively (Figure 2).
DISCUSSION In our study, serum ADA levels in sHLH were significantly higher than healthy donors and patients in conditional control group. sHLH is a severe hyperinflammatory condition, which is characterized by prolonged fever, cytopenia, liver dysfunction, hepatosplenomegaly, and hemophagocytosis, caused by activated macrophages. The main pathophysiology of sHLH is hypercytokinemia resulting in hyperactivation of T lymphocytes and macrophages, and proliferation and infiltration of these cells into various organs [12]. The immune system is regulated by T-helper cells (Th), which is classified as type 1 Th-cells (Th1) and © 2015 John Wiley & Sons Ltd, Int. Jnl. Lab. Hem. 2015, 37, 544–550
Table 2. ADA levels in sHLH groups and healthy control subjects Study groups
Serum ADA (U/L)
sHLH LAHS (n = 20) IAHS (n = 15) MAS (n = 6) Healthy subjects (n = 20)
115.12 150.29 91.33 57.40 12.36
69.17* 76.08* 44.38* 11.30* 4.07
*Compare with healthy subjects, P < 0.001. All data were shown in form of mean SD.
type 2 Th-cells (Th2). According to different cell types, the immune system will develop different immune responses. Th1 cell subset mainly secrets IL-2, IL-12, interferon (IFN)-c and tumor necrosis factor (TNF)-b and mediates cell immunity. Th2 cell subset mainly
548
W. CHEN ET AL. | ELEVATED ADA LEVELS IN SHLH
Figure 1. Compared ADA level between experimental group and conditional control group. The error bars represent standard deviation.
secrets IL-4, IL-5, and IL-9 and mediates humoral immunity [13]. Osugi et al. [14] and Tang et al. [15] have proposed that IFN-c, which is a Th1 type cytokine with immunomodulatory properties, was significantly higher during acute phase of sHLH in comparison with those after remission and healthy volunteers. IL-4, which is a Th2 type cytokine, however, was not significantly different between them. Janka [16] has shown that Th1 cytokines such as IFNc, IL-12, and IL-18, which correlate with disease activity were obviously decreased after disease remission. These all indicate that sHLH shows a Th1 polarization. Adenosine deaminase is an enzyme involved in purine catabolism and plays a role in proliferation,
Figure 2. ROC curve analysis of patients with LAHS vs. benign disease-associated HLH (IAHS and MAS).
maturation, and differentiation of lymphocytes, and its level increases during mitogenic and antigenic responses of these cells [8, 17]. Patients with deficiency of ADA develop severe defects in humoral and cellular immunity, which is called severe combined immunodeficiency (ADA-SCID) [18]. In the course of T lymphocyte activation and secretion of IFN-c, TNFa, and IL-6, ADA interacts with CD26 to produce a costimulatory signal. During this activation, ADA activity is regulated positively by IL-2 and IL-12 and negatively by IL-4 [19]. Elevated ADA level may be a part of the activated Th1 response [20] and has been reported in several diseases, which are characterized by T-cell activation [7–10].
Table 3. Compared laboratory findings between LAHS and benign disease-associated HLH (IAHS and MAS) Laboratory finding
LAHS
PLT (9109/L) Neu (9109/L) Hb (9109/L) SF (lg/L) FIB (g/L) TG (mM) ALT (U/L) AST (U/L) ADA (U/L)
48.95 1.62 95.50 35828.90 1.58 3.06 262.47 366.69 150.29
Benign disease-associated HLH
41.54 1.34 19.14 20889.11 1.03 1.83 244.66 306.18 76.08
56.43 2.19 99.81 32571.14 2.00 2.38 197.47 326.72 81.63
35.68 1.90 25.29 19225.79 1.01 1.46 150.43 296.78 40.71
P value >0.05 >0.05 >0.05 >0.05 >0.05 >0.05 >0.05 >0.05
