Comprehensive analyses and characterization ofhaemophagocytic lymphohistiocytosis in Vietnamese children

Haemophagocytic lymphohistiocytosis (HLH) is a rapidly fatal

haematological disorder frequently associated with severe

infections and malignancies (Mathew et al, 2000; Henter et al,

2002; Gupta et al, 2009). The disease is characterized by

prolonged fever, jaundice, hepatosplenomegaly, pancytopenia

and coagulopathy (Henter et al, 2002). In past decades, a series

of studies have revealed that HLH results from the deregula-

tion of natural killer cells and cytotoxic T cell response, leading

to systemic proinflammatory cytokine injuries and macro-

phage activation with haemophagocytosis in bone marrow and

the lymphoid system (Ueda et al, 2006; Henter et al, 2007).

Current advances in molecular studies have further docu-

mented that a substantial percentage of HLH patients are

primarily associated with mutations of the SH2D1A (SAP),

PRF1, or UNC13D genes (Ohno et al, 2003; Henter et al, 2007;

Su, 2008). Furthermore, HLH may accompany autoimmune

disorders and malignancies, such as lymphomas (Gutierrez

et al, 2003; Krenova et al, 2007). Therefore, HLH encompasses

a diverse spectrum of diseases and their distribution may vary

among different ethnic populations or geographic regions.

Recently, a therapeutic protocol of HLH-2004 by the Histio-

cyte Society has been widely adopted with specific treatments

based on disease severity and particular genetic abnormalities

(Imashuku et al, 2000; Henter et al, 2007; Horne et al, 2008;

Su, 2008; Yamada et al, 2008).

Haemophagocytic lymphohistiocytosis is not well docu-

mented in Vietnamese children. Before 2000, the recognition

of this disease was hampered by the lack of appropriate

guidelines for diagnosis. Based on the guidelines for HLH-94

and HLH-2004 (Henter et al, 2002, 2007), HLH was increas-

ingly diagnosed and treated at the Children’s Hospital No. 1 of

Ho-Chi-Minh City (HCMC), Vietnam. The general outcome,

Lam T. My,1,2 Le B. Lien,1 Wen-Chuan

Hsieh,3 Toshihiko Imamura,4 Tran N. K.

Anh,1 Phan N. L. Anh,1 Nguyen T.

Hung,1 Fan-Chen Tseng,3 Chia-Yu Chi,3

Ngo T. H. Dao,5 Duong T. M. Le,5 Le

Q. Thinh,5 Tran T. Tung,5 Shinsaku

Imashuku,4 Tang C. Thuong1 and

Ih-Jen Su3,6

1Department of Clinical Haematology, Children

Hospital No. 1, 2Department of Paediatrics, The

University of Medicine and Pharmacy, Ho-Chi-

Minh City, Vietnam, 3Division of Infectious

Diseases, National Health Research Institutes,

Tainan, Taiwan, 4Department of Haematology,

Kyoto Prefecture University of Medicine, Kyoto,

Japan, 5Department of Laboratory Haematology,

Children’s Hospital No. 1, Ho-Chi-Minh City,

Vietnam, and 6Department of Pathology,

National Cheng Kung University College of

Medicine and Hospital, Tainan, Taiwan

Received 13 July 2009; accepted for

publication 1 September 2009

Correspondence: Dr Ih-Jen Su, Division of

Infectious Diseases, National Health Research

Institutes, 367 Sheng-Li Road, Tainan, Taiwan.

E-mail: suihjen@nhri.org.tw

Summary

Haemophagocytic lymphohistiocytosis (HLH) is a fatal haematological

disorder with diverse aetiology. This prospective study was undertaken to

characterize HLH cases in Vietnamese children. Clinical and laboratory data,

genetic analyses and outcome of the HLH patients were analysed. A total of

33 patients were enrolled from March 2007 to December 2008, with a median

age of 3 years. Mutations of the SH2D1A (SAP) and PRF1 genes were

detected in one patient, respectively. The virus association was high, up to

63Æ6% (21/33), including Epstein–Barr virus (19/33), cytomegalovirus (2/33)

and dengue virus (2/33). Five patients had malignant lymphoma and two had

autoimmune diseases. Twenty-eight patients were treated according to the

HLH-2004 protocol. The first response rate was 64Æ3% (18/28), with an early

death rate of 35Æ7% (10/28). High levels of interferon-c, interleukin-10, MIG

and interferon-inducible protein-10 (IP-10) were associated with early

mortality (P < 0Æ05). Reactivation among the responders was high (9/18) and

the uneventful resolution was low (3/18) after a median follow-up of

35 weeks. In conclusion, the majority of HLH cases are associated with virus

infections in Vietnamese children. Familial HLH is rare. The frequent

reactivation and high mortality demands a more appropriate therapeutic

regimen in tropical areas like Vietnam.

Keywords: haemophagocytic lymphohistiocytosis, Epstein–Barr virus, SAP,

cytokine storm, Vietnam.

research paper

First published online 27 October 2009ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310 doi:10.1111/j.1365-2141.2009.07957.x

however, was poor. The major challenge for the paediatricians

was, and continues to be, the diagnostic criteria and differen-

tial diagnosis of HLH, particularly in a tropical region where

dengue and other infectious diseases prevail (Mathew et al,

2000; Gurgey et al, 2005). In order to understand the potential

diagnostic and therapeutic problems and to characterize the

distribution of disease entities of HLH in Vietnam, a

prospective study on the diagnosis and treatment of HLH

was launched by collaboration between Children’s Hospital

No.1, HCMC, Vietnam and the Division of Infectious Diseases

of the National Health Research Institutes, Taiwan, with

consultation from Professor Shinsaku Imashuku, Kyoto, Japan.

Through this collaborative study, the clinical assessments,

cytokine profiles, virological etiologies, family studies and

genetic analyses and treatment outcome of HLH in Vietnamese

children were analysed.

Materials and methods

Patients and diagnosis

A prospective study on HLH was carried out from 1 March

2007 to 30 December 2008 at the Children’s Hospital No.1,

HCMC, Vietnam. The criteria for diagnosis of HLH were

based on the guideline of the HLH-2004 proposed by the

Haemophagocytic Lymphohistiocytosis Study Group (Henter

et al, 2007), and patients who fulfilled at least five of the

following signs or laboratory parameters were enrolled: fever

(higher than 38�C for more than 7 d), splenomegaly or

hepatomegaly, cytopenia (affecting ‡2 of three lineages in the

peripheral blood: haemoglobin (<90 g/l), leucopenia

(<5Æ0 · 109/l), neutropenia (<1Æ0 · 109/l), thrombocytopenia

(<100Æ0 · 109/l), hypertriglyceridemia (‡3 mmol/l) or hypo-

fibrinogenemia (£1 g/l), hyperferritinemia (‡500 lg/l) and

haemophagocytosis in bone marrow or lymph nodes. The

demographic, clinical and laboratory data of HLH patients

were recorded and collected beginning with the first hospital-

isation.

Routine laboratory studies

The general laboratory tests after admission included blood

count, blood smear, Coombs’ test, aspartate transaminase

(AST), alanine transaminase (ALT), bilirubin, activated partial

thromboplastin time (APTT), prothrombin time (PT), fibrin-

ogen, ferritin, triglyceride, routine chest X-ray and abdominal

echography. To identify pathogens, blood culture and sero-

logical tests for hepatitis B virus (HBV), hepatitis C virus

(HCV), human immunodeficiency virus (HIV), Epstein–Barr

virus (EBV), dengue virus, cytomegalovirus (CMV) and

Salmonella typhi (Widal test) were performed. Bone marrow

aspiration was done on all patients. Biopsy of the lymph

nodes or liver was performed if necessary. The viral loads of

EBV and CMV infection were determined by real-time

polymerase chain reaction (Q-PCR) on the plasma samples

taken at diagnosis. The diagnosis of systemic lupus erythe-

matosus (SLE) or other autoimmune diseases was based on

the criteria from the American Rheumatism Association (Tan

et al, 1982). Patients with malignancy-associated HLH were

transferred to the National Cancer Hospital or the Hospital of

Blood and Transfusion of HCMC for further treatment.

Patients with immune disorder-associated HLH, including

SLE or juvenile idiopathic arthritis (JIA), at first were treated

according to the guidelines of HLH-2004 with ciclosporin and

dexamethasone. But after enough criteria for the positive

diagnosis of SLE or JIA were reached, they were further

treated pursuant to the guidelines for SLE or JIA. Patients

with infectious diseases-associated HLH, such as bacterial (e.g.

typhoid fever, tuberculosis) or parasitic (malaria), were

treated with specific antibiotics or anti-malarial drugs accord-

ing to the guidelines of the Ministry of Public Health,

Vietnam.

Cytokine and chemokine studies

Cytokine [including c-interferon (IFN-c), tumour necrosis

factor-a (TNF-a), interleukin (IL)-2, IL4, IL-6 and IL-10]

and chemokine [including IL-8, RANTES, MCP-1 (monocyte

chemoattractant protein-1), IP-10 (interferon-inducible

protein-10) and MIG (Monokine induced by IFN-c)] levels

in the plasma of HLH patients were measured at diagnosis

by the Becton-Dickinson Human Th1/Th2 Cytokine

Cytometric Bead Array kit-II (Becton-Dickinson Pharmin-

gen, San Diego, CA, USA) and the Becton-Dickinson Human

Chemokine Cytometric Bead Array kit (Becton-Dickinson

Pharmingen), respectively. Both cytokine and chemokine

assays were performed according to the manufacturer’s

instructions.

Genetic analyses

Mutation analyses for SH2D1A and PRF1. Genomic DNA was

isolated from PBMC according to the manufacture’s

instructions for the QIAamp DNA blood kit (QIAGEN,

Valencia, CA, USA). To detect mutations, we used two

primer pairs for SH2D1A: (exon 2: F 5¢-GGAAACTGT

GGTTGGGCAGATACAATATGG-3¢ and R 5¢-GGCTAAACA

GGATGGGACCAAAATTCTC-3¢; exon 3: F 5¢-GCTCCTCTT

GCAGGG AAATTCAGCCAACC-3¢ and R 5¢-GTCACCTCT

CATTTGACTTGCTGGCTACATC-3¢); and another two

primer pairs for PRF1 (exon 2: F5¢-TGTGCCCTGATAAT

CTGTG-3¢ and R 5¢-GCAGCCTCCAAGTTTGA-3¢; exon 3: F

5¢-TCCTAGTTCTGCCCACTTAC-3¢ and R 5¢-GGCTCCCAC

TGTGAGA-3¢) as described elsewhere (Sayos et al, 1998;

Goransdotter Ericson et al, 2001). For PRF1 gene analysis,

two primer pairs were used: F5¢-TTCCCCCATGTGCC CT

GATAATC-3¢ and R5¢-AGCAGCCTCCAAGTTTGATTGG-3¢for exon 2; F5¢-CCAGTCC TAGTTCTGCCCACTTAC-3¢ and

R5¢-GAACCCCTTCAGTCCAAGCATAC-3¢ for exon 3. For

each PCR reaction, 35 cycles were performed on 100 ng of

L. T. My et al

302 ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310

DNA in 50 ll of PCR reaction mixture, which consisted of

50 nmol/l KCl, 10 mmol/l Tris–HCl (pH 8Æ3), 1Æ5 mmol/l

MgCl2, 200 nmol/l dNTP, 0Æ2 lmol/l primers and 1Æ25 units of

Taq polymerase (Takara, Otsu, Shiga, Japan). PCR products

were analysed using 2% agarose gels and then sequenced and

analysed on an ABI Genetic Analyzer (Applied Biosystems Inc.,

Foster City, CA, USA).

Treatment and outcome

The initial therapy (initial period) comprised an 8-week

treatment with dexamethasone, ciclosporin A, etoposide and

broad-spectrum antibiotics in the same doses as described

in the HLH-2004 guidelines. For patients who experienced

disease reactivation, a continuation therapy (follow-up period)

of 9–40 weeks was given after the initial therapy. The

treatment consisted of dexamethasone (every second week,

10 mg/m2 for 3 d) and ciclosporin (ciclosporin A, 6 mg/kg per

day). Because we could not differentiate between primary HLH

and secondary HLH at hospitalisation, the subsequent therapy

was only used for patients who had not fulfilled the criteria for

response. Dexamethasone was used after 40 weeks of follow-

up. The outcome of patients was categorized into the following

groups: (A) early death group, for patients who died during

the initial therapy period (8 weeks); (B) first response group,

for patients who survived after the initial therapy period; (C)

transfer group, for patients who were excluded from study

because of underlying diseases (malignant disease, autoim-

mune disease, or infectious disease from special pathogens

such as malaria, tuberculosis and typhoid); (D) reactivation

group, for patients who have achieved remission but who

again developed at least three of the following signs:

fever, hepatosplenomegaly, platelet count <100 · 109/l, hyper-

triglyceridaemia, hypofibrinogenaemia, haemophagocytosis,

hyperferritinaemia. This latter group was divided into two

subgroups: the survival reactivation group (D1) for patients

who had reactivation and were followed continuously; and the

death reactivation group (D2) for those who had activation

and died. Finally, the resolution group (E) included patients

who were still alive after finishing the HLH-2004 therapy.

Patients lost to follow-up after discharge were assigned to the

lost to follow-up group.

Statistical analysis

Categorical data analyses were performed for variables related

to clinical signs, aetiology, cytokine profiles and treatment.

Percentages of patients positive for a variable were compared

between the cases and control groups using the chi-square

test, or Fisher’s exact tests when expected counts were <5.

Continuous variables, such as biochemical laboratory data and

duration of activities, were described by median (range) value

and compared using the Wilcoxan–Mann–Whitney U-test.

Differences were considered significant if the P value was

<0Æ05.

Results

Demographic and clinical data

From March 2007 to December 2008, a total of 33 patients

with HLH were diagnosed and enrolled at the Children’s

Hospital No.1, HCMC (Table I). The median (range) age at

disease onset was 3 years (0Æ1–14 years); the 75th percentile

Table I. Demographic, clinical and laboratory findings of 33 patients

with HLH.

Variables Data

Patients’ characteristics

Age (years); median (range) 3 (0Æ1–14)

Male–female ratio 1Æ5 (20/13)

Birth weight (kg); median (range) 3Æ071 (2Æ3–3Æ8)

Positive previous illness; n (%) 8 (24Æ4)

Time to diagnosis (d); median (range) 14Æ0 (6–90)

Underlying diseases n (%)

Family history 1 (3Æ0)

Malignant haematology association 5 (15Æ1)

Immune disorders 2 (6Æ0)

Clinical features; n (%)

Fever 33 (100)

Duration of fever (d); median (range) 10Æ0 (3–75)

Hepatomegaly 31 (93Æ9)

Hepatosplenomegaly 24 (72Æ7)

Skin disorders 13 (39Æ4)

Haemorrhagic disorder 4 (12Æ1)

Central nervous system disorders 4 (12Æ1)

Laboratory data; median (range)

Haemoglobin (g/l) 82 (43–128)

White blood cell count (·109/l) 3Æ3 (0Æ8–15Æ3)

Neutrophil count (·109/l) 0Æ6 (0Æ1–5Æ9)

Platelet count (·109/l) 50Æ0 (7Æ0–449Æ0)

Aspartate aminotransferase (u/l) 173 (15Æ0–2920Æ0)

Alanine aminotransferase (u/l) 110 (14Æ0–1987Æ0)

Total bilirubin (lmol/l) 29Æ1 (3Æ4–273Æ6)

Triglyceride (mmol/l) 5Æ1 (1Æ6–10Æ9)

Fibrinogen (g/l) 2Æ8 (0Æ30–6Æ0)

Ferritin (lg/l) 7000 (43–88 000)

C-reactive protein (mg/l) 21Æ9 (0Æ5–62Æ8)

Coagulation study

APTT (s); median (range) 33Æ0 (22Æ2–120)

PT (s); median (range) 14Æ1 (10Æ8–120)

Increased PT (>15 s); n (%) 6/19 (31Æ6)

Involvement of viral infections; n (%)

EBV 19 (57Æ6)

CMV 2 (6Æ1)

EBV and CMV 2 (6Æ1)

EBV or CMV 21 (63Æ6)

Other studies; n (%)

Abnormal chest X ray 17 (51Æ5)

Abdominal echography 10 (30Æ3)

APTT, activated partial thromboplastin time; PT, prothrombin time;

EBV, Epstein–Barr virus; CMV, cytomegalovirus.

HLH in Vietnam

ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310 303

of age was 6 years; five patients were aged <1 year. The male–

female ratio was 1Æ5. The median (range) birth weight was

3Æ071 kg (2Æ3–3Æ8 kg). Two cases were diagnosed with auto-

immune disease. Only one case had a family history of sibling

death due to pneumonia (at 8 months); otherwise, no

familial histories were identified for the remaining 32

patients.

The median (range) duration of persistent fever was 10 d

(3–75 d). The duration between onset of symptoms and

diagnosis varied from 6 to 90 d with a median of 14 d.

Common clinical features at admission were hepatomegaly

(31/33; 93Æ9%), hepatosplenomegaly (24/33; 72Æ7%), skin

lesions (13/33; 39Æ4%) and neurological signs (4/33; 12Æ1%)

with lethargy in three and paresis of feet in one.

Laboratory data

At the time of diagnosis, the common laboratory findings

were hypertriglyceridaemia (25/29; 86Æ2%), thrombocyto-

penia (26/33; 78Æ8%), hyperferritinaemia (25/32; 78Æ1%)

and neutropenia (22/33; 66Æ7%). Haemophagocytosis was

detected in the bone marrow in all the patients. Other

abnormal tests were increased C-reactive protein (CRP; 24/

33; 72Æ7%), increased AST (15/28; 53Æ6%), ALT (15/28;

53Æ6%) and total bilirubin (14/29; 48Æ3%), pleural effusion

(7/33; 21Æ2%) and diffuse lung infiltrates (10/33; 30Æ3%)

detected by chest radiograph, and abdominal effusion

detected by ultrasound (10/33; 30Æ3%). Virus-associated

HLH was high (21/33; 63Æ6%). The serological tests for viral

infection revealed a high rate of EBV and CMV infection:

titres of EBV IgG higher than 640 (14/33; 42Æ4%), positive

test for EBV IgM (4/33; 12Æ1%), and titres of CMV IgG

higher than 640 (14/33; 42Æ4%). Two patients were diagnosed

with dengue haemorrhagic fever with positive IgM for

dengue virus (2/33; 6Æ0%). Real-time PCR for free EBV in

the plasma was interpreted as positive (19/33; 57Æ6%) if the

copy numbers were higher than 10 copies per ml; and real-

time PCR for free CMV in the plasma was positive (higher

than 10 copies per ml) (2/33; 6Æ0%). Two cases were both

real time PCR positive for EBV and CMV (2/33; 6%)

(Table I). Blood culture was positive in four cases (4/33;

12Æ1%): two cases were Acinetobacter and two cases coagulase-

negative Staphylococcus.

Associated underlying diseases

Haematological malignancy-associated HLH. Five patients had

associated malignant haematological diseases (5/33; 15Æ1%)

(Table I), including EBV-positive Hodgkin lymphoma; EBV-

positive peripheral T cell lymphoma (two cases) and

Langerhans cell histiocytosis. The remaining case was a

4-year-old male who had fulfilled the diagnosis for EBV-

associated HLH at the time of first hospitalisation, but after

12 months of follow-up developed reactivation and was

diagnosed with acute lymphoblastic leukaemia (ALL).

Immune disease-associated HLH. Two patients were

diagnosed with immune disease-associated HLH (2/33;

6Æ0%), (Table I). One was diagnosed with juvenile

rheumatoid arthritis (JRA). The other case concerned a

10-year-old female admitted with severe anaemia, acute

arthritis, positive Coombs’ test and positive anti-nuclear

antibody, who was diagnosed with SLE.

Cytokine and chemokine profiles

Out of a total of 33 HLH patients, the plasma of 32 patients at

diagnosis was available for determining T-helper cell type 1/2

(Th1/Th2) cytokines and chemokines assay. Control plasma

was obtained from seven healthy subjects for comparison. The

mean plasma concentrations of cytokines and chemokines in

the HLH group and the healthy control group (n = 7) are

shown in Fig 1. Elevated blood levels of IL-6, IL-10, TNF-a and

IFN-c were observed in the HLH group (P < 0Æ05, P < 0Æ0001,

P < 0Æ01 and P < 0Æ01, respectively). Among the ten cases of

early death, eight patients (80%) had high IFN-c levels, above

500 pg/ml. IL-2 and IL-4 were consistently at low levels in HLH

patients and showed no significant difference with those in the

control group. Except for RANTES, most chemokine levels

measured (including IL-8, MIG, MCP-1 and IP-10) were all

significantly higher in the haemophagocytic syndrome patients

than in the healthy control group (Fig 1). The comparison of

cytokine and chemokine levels between the initial response

group and the early death group are described below.

Genetic analyses

SAP and perforin gene analysis. A boy aged 1 year and

4 months had a family history of sibling death resulting from

a similar illness and pneumonia. Mutation analyses of SH2D1A

revealed a smaller than normal band of transcript in the exon 2

primer set assay. Further sequencing revealed a 51 bp deletion

from the exon 2–5¢ splicing site of intron. This mutation is

expected to result in an abnormal SH2D1A expression and a

loss of proper SH2D1A function. This case had a high copy

number of EBV in the plasma and was therefore identified as a

typical case of X-linked lymphoproliferative disease (XLP).

Sequencing of the PCR products on the remaining 32 samples

revealed a wild type sequence of the SH2D1A gene. Mutation

analyses of PRF1 after sequencing of the PCR products revealed

a hetero single nucleotide substitution of 10 CT (R4C) in the

exon 2 of one case. However, no familial history was identified

in this case. No mutations of PRF1 were identified for the

remaining 32 cases.

Treatment outcome of the patients during the initialperiod

In the initial therapy pursuant to the HLH-2004 guidelines,

five patients were transferred to another specialist disease

hospital because of their underlying diseases, including three

L. T. My et al

304 ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310

patients with malignancy-associated HLH, one patient with

JRA-associated HLH, and one case with tuberculosis-associ-

ated HLH. The remaining 28 patients (28/33; 84Æ8%) with

HLH were treated according to the HLH-2004 guidelines

(Fig 2). During this period, 26 patients (26/28; 92Æ8%) were

treated as per the HLH-2004 guidelines, and two patients

received no treatment because of rapid fatality or early

resolution. All patients (26/26, 100%) received steroid treat-

ments, and most (25/26, 96Æ1%) received ciclosporin treat-

ments. However, the percentages of patients who received

etoposide and intravenous immunopglobulin (IVIG) were

suboptimal (5/26, 19Æ2% and 9/26, 34Æ6%, respectively) due to

the shortage of supply or the severity of the diagnosis of most

patients. The common supportive treatments were whole

blood cell (10/28, 35Æ7%), packed red cell (13/28, 46Æ4%),

fresh plasma (5/28, 17Æ8%) and platelet transfusions (2/28,

7Æ1%).

The rate of early death was high (10/28; 35Æ7%) (Fig 2); all

such deaths were attributable to superimposed bacterial

infections associated with HLH, with one case proving to be

Fig 2. Flow chart and outcomes of 33 patients with HLH in Viet-

namese children treated with HLH-2004 protocol. There was a high

rate of early death and frequent reactivation. Only three patients had

an uneventful resolution, including two with dengue infection.

(A)

(B)

Fig 1. Cytokine (A) and chemokine (B) levels in 32 haemophoagocytic syndrome (HPS) patients (the plasma in one patient was not available),

compared to seven healthy controls. Data were analysed by the Mann–Whitney test with GraphPad Prism 4.0 software (GraphPad Software Inc., San

Diego, CA, USA). *P < 0Æ05, ***P < 0Æ001.

HLH in Vietnam

ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310 305

lymphoma association. The percentage of first response was

64Æ3% (18/28).

The median (range) durations of hospitalisation of the early

death group and the first response group were 16 d (3–48 d)

and 24 d (7–57 d), respectively. The median number of days

for hospitalisation for treatment with dexamethasone and

ciclosporin of the early death group was significantly lower

(P = 0Æ01) than that of the first response group (Table II).

Other factors that showed significant differences between the

early death group and first response group included the levels

of cytokines and chemokines. High levels of IFN-c, IL-10,

LIG and IP-10 were associated with a poor prognosis in HLH

patients (P = 0Æ03, P = 0Æ04, P = 0Æ02 and P = 0Æ01, respec-

tively). No significant statistical difference between the two

groups were revealed for other factors, including age, ferritin-

emia and levels of TNF-a, IL-6, IL-8 and MCP-1. The EBV

association and viral loads also showed no significant differ-

ences between the two groups (Table II).

Table II. Comparison of the clinical and labo-

ratory characteristics of the early death group

and the first response group in HLH patients.

Early death

(n = 10)

First response

(n = 18) P-value

Characteristics

Age (year) 1Æ3 (0Æ2–8Æ0) 3Æ3 (0Æ4–13Æ3) 0Æ09

Male per cent (%) 6 (60%) 10 (55Æ6%) 1Æ0Malnutrition (%) 4 (40%) 4 (22Æ2%) 0Æ4Birth weight (kg) 3Æ1 (2Æ3–3Æ5) 3Æ2 (2Æ7–3Æ8) 0Æ4Duration of fever (d) 11 (4–31) 10 (4–30) 0Æ4Duration of diagnosis (d) 12 (6–38) 13 (5–32) 0Æ8

Laboratory data

Hb (g/l) 83 (50–124) 88 (43–128) 0Æ7White blood cell count

(1Æ0 · 109/l)

2Æ6 (0Æ8–15Æ3) 3Æ1 (1Æ3–9Æ8) 0Æ8

Neutrophil count (1Æ0 · 109/l) 0Æ5 (0Æ1–1Æ3) 0Æ5 (0Æ1–2Æ1) 0Æ6Platelet count (1Æ0 · 109/l) 26Æ5 (7Æ0–22Æ0) 54Æ0 (20Æ0–193Æ0) 0Æ6Ferritin (lg/l) 20 200 (1899–88 000) 6100 (43–49 600) 0Æ2Triglyceride (mmol/l) 6Æ1 (3Æ8–10Æ9) 5Æ0 (1Æ6–8Æ0) 0Æ06

Fibrinogen (g/l) 2Æ2 (0Æ7–4Æ0) 2Æ9 (0Æ3–6Æ0) 0Æ3Prothrombin time (s) 15Æ9 (11Æ9–24Æ6) 13Æ9 (10Æ8–120) 0Æ4Activated partial thromboplastin

time (s)

34Æ5 (27Æ4–120) 31Æ3 (24Æ4–120) 0Æ6

Aspartate aminotransferase (u/l) 375Æ5 (31Æ0–2920Æ0) 109Æ0 (15–2182) 0Æ2Analine aminotransferase (u/l) 141Æ0 (14Æ0–943Æ0) 98Æ0 (14Æ0–1987Æ0) 0Æ9

EBV association

Positive EBV–PCR 7 (70Æ0%) 10 (55Æ6%) 0Æ6Copies of EBV (·103/ml) 1Æ95 (0Æ091–85Æ200) 0Æ250 (0Æ001–6Æ37) 0Æ3C-reactive protein (mg/l) 37 (4Æ3–250) 21Æ3 (0Æ5–318) 0Æ8

Cytokine/chemokine profiles

TNF (pg/ml) 3Æ7 (0–9Æ3) 2Æ2 (0–7Æ8) 0Æ07

IFN (pg/ml) 631Æ0 (30Æ6–10 000) 24Æ8 (0–2844Æ0) 0Æ03

IL-6 (pg/ml) 73Æ9 (2Æ3–5571Æ8) 6Æ4 (0Æ0–65Æ3) 0Æ2IL-10 (pg/ml) 1117Æ0 (51Æ8–9003Æ0) 32Æ1 (0Æ0–3218Æ0) 0Æ04

IL-8 (pg/ml) 2397Æ0 (13Æ5–10 000*) 80Æ0 (12Æ3–391Æ6) 0Æ1MIG (pg/ml) 10 000Æ0 (3025Æ0–10 000*) 5863Æ4 (662Æ0–10 710Æ0) 0Æ02

MCP-1 (pg/ml) 232Æ2 (7Æ2–10 000*) 99Æ9 (14Æ0–749Æ6) 0Æ1IP-10 (pg/ml) 5763Æ0 (831Æ8–10 880Æ0) 1218Æ0 (71Æ8–6124Æ0) 0Æ01

Duration of initial

hospitalisation (d)

16 (3–48) 24 (7–57) 0Æ01

Duration of HLH-2004 treatment in the initial hospitalisation (d)

Dexamethasone 15 (0–23) 22 (0–46) 0Æ01

CSA 14 (0–21) 22 (0–46)

EBV, Epstein–Barr virus; PCR, polymerase chain reaction; TNF, tumour necrosis factor; IFN,

interferon; IL, interleukin; MIG, monokine induced by IFN-c; MCP-1, monocyte chemoattr-

actant protein-1; IP-10, interferon-inducible protein-10; CSA, ciclosporin A.

*Upper limitation of cytokines and chemokines is 10 000 pg/ml. When the cytokine or

chemokine level is above 10 000 pg/ml, then the values are simply reported as 10 000 pg/ml.

L. T. My et al

306 ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310

Treatment outcome of the patients in the continuationperiod

There were 18 patients during the follow-up period, which

ranged from 9 weeks to more than 40 weeks. During this

period, four patients were lost to follow up (4/18; 22Æ2%). Two

other patients were transferred because of the development of

underlying diseases: one had acute lymphoblastic leukaemia

and the other had SLE. The rate of reactivation was high (9/18;

50Æ0%) including 22Æ2% (4/18) for the death reactivation

group and 27Æ8% (5/18) for the survival reactivation group

(Fig 2). The percentage of patients with resolution was low

(3/18; 16Æ7%). The median (range) follow-up duration for the

death reactivation group was 17 weeks (10–35 weeks), for the

survival reactivation group 36 weeks (33–87 weeks) and

72 weeks (55–82 weeks) for the resolution group.

Discussion

This study characterized, for the first time, childhood HLH in

HCMC, Vietnam. Considering there were 33 HLH cases in a

single institution in 1 year and 9 months, the projected

incidence of childhood HLH in Vietnam was relatively high,

as compared to that in Taiwan, Canada, Turkey or Thailand

(Veerakul et al, 2002; Chen et al, 2004; Gurgey et al, 2005).

Recently, a nationwide survey of HLH in Japan showed that

the mean number of 160 patients per year between 2001 and

2005 (Ishii et al, 2007). Our studies revealed a high prevalence

of EBV infection. Two HLH cases were associated with dengue

infection, due to the prevalence of dengue virus infection in

this tropical region. Familial HLH is rare in Vietnamese

children, with only one case found during this study of XLP

(1/33; 3%), a situation similar to that of Taiwan (Chen et al,

2004; Lee et al, 2009).

The diagnosis of HLH in tropical regions is complicated by

common clinical and laboratory findings which may present in

other tropical infections, such as severe malaria, tuberculosis,

typhoid fever or HIV infection (Mathew et al, 2000; Veerakul

et al, 2002). In this study, the time between the onset of

symptoms and the diagnosis varied from 6 to 90 d (median

14 d). The delay of diagnosis or prolonged duration of

diagnosis (14 d) again reflected the difficulties of the paedi-

atricians to diagnose HLH when facing these disorders. In

practice, persistent fever, hepatosplenomegaly, hyperferritina-

emia and hypertriglyceridaemia and cytopenia, together with

the progressive course, represent a good index for suspicion of

an HLH diagnosis and prompt the physician to study the

haemophagocytosis in bone marrow in order to establish the

diagnosis. Since early recognition of HLH and immediate

application of appropriate regimens to HLH patients have

been shown to improve or to arrest the disease progression

(Imashuku et al, 2004; Lee et al, 2005; Tang et al, 2008;

Yamada et al, 2008), the delayed diagnosis of our HLH

patients may partly explain the poor outcome of treatments in

this cohort.

Besides delayed diagnosis, this study attempted to identify

other potential prognostic factors, such as cytokine levels, virus

association and genetic factors. The cytokine and chemokine

studies revealed a consistent elevation of several cytokines and

chemokines, though not IL-2 and IL-4, which were usually not

elevated in our HLH patients. These findings indicated that a

cytokine storm occurred in most of our HLH patients and

were responsible for the clinical manifestations and abnormal

laboratory data of HLH (Ishii et al, 1991; Fujiwara et al, 1993;

Osugi et al, 1997). The high levels of IFN-c, IL-6, IP-10 and

MIG in HLH patients with poor prognosis or early death

support the observations of previous reports (Ohga et al, 1993;

Imashuku et al, 1994; Tang et al, 2008). Hypercytokinemia is

responsible for the systemic problems of HLH, such as

pancytopenia, depletion of the immune system, diffuse respi-

ratory infiltrates of lungs and liver damage (Imashuku, 2002).

Therefore, anti-inflammatory or immunoregulatory agents

targeted to the master genes of inflammation, such as NFKB1

and AP1, may ameliorate the outcome of HLH patients

(Chuang et al, 2007a; Su, 2008). Agonists of peroxisome

proliferator-activated receptors (PPAR) have been recently

found to exert such remarkable results with minimal side

effects in an animal model of virus-associated haemophago-

cytic syndrome and could represent potential therapeutic

agents (Chuang et al, 2007a).

Haemophagocytic lymphohistiocytosis can be classified into

genetic (primary) HLH and acquired (secondary) HLH

(Henter et al, 2007). Genetic HLH can be further divided into

familial HLH (FHLH) and immune deficiency HLH, including

Chediak–Higashi syndrome (CHS), Griscelli syndrome (GS)

and XLP (Henter et al, 1998; Janka, 2007). Both genetic

subgroups are associated with impaired NK cell function

(Sullivan et al, 1998; Kogawa et al, 2002; Henter et al, 2007).

This study did not include assessment of NK function or NK

cells due to the unavailability of flow cytometry in the hospital.

The prevalence of genetic HLH appeared to be low (2/33; 6%)

in this study, based on the genetic mutation analyses of

SH2D1A and PRF1 genes. The analysis of UNC13D gene,

however, was not performed in this study due to the large

UNC13D gene of 32 exons for such a big sample size of 33

cases. There was only one case of XLP involving a family

history, specifically a sibling death due to pneumonia at

8 months. XLP has been now recognized as a primary

immunological disorder in infants and children (Mischler

et al, 2007). The only case of a mutation of the PRF1 gene in

this study showed no family history. Other candidate genes

associated with HLH were not explored here and whether

other ‘acquired’ HLH may have underlying genetic or immu-

nological disorders remains to be clarified.

Although we detected a high association with EBV infection

(19/33, 57Æ6%), no prognostic significance of the EBV

association, for either reactivation or mortality, could be

identified in this study, similar to the report from Taiwan (Su

et al, 1994; Chen et al, 2004; Lee et al, 2009). EBV infects T

cells in acquired HLH and the EBV latent membrane protein 1

HLH in Vietnam

ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310 307

(LMP1) can transcriptionally inhibit the expression of

SH2D1A, leading to sustained T cell activation and the release

of Th1 cytokines, such as TNF-a and IFN-c, via the

TRAF2,5/nuclear factor jB (NF-jB) pathway (Chuang et al,

2005, 2007b). Interestingly, two HLH cases were noted to have

positive serology tests for dengue virus at the convalescent

stage of dengue haemorrhage fever. Both patients had a good

response to treatment with dexamethasone and ciclosporin

and recovered thereafter. In Thailand, Veerakul et al (2002)

also reported three paediatric HLH patients with positive

dengue virus detection. Several reports of adult dengue

haemorrhagic fever-associated HLH have been reported from

Taiwan (Lu et al, 2005), Thailand (Veerakul et al, 2002;

Srichaikul et al, 2008) and Japan (Nakamura et al, 2009).

Further studies are needed to clarify the mechanism of HLH

association with dengue virus infection.

For malignancy-associated HLH, four patients were identi-

fied with haematological malignancies, including two cases of

peripheral T cell lymphoma, one case of Hodgkin lymphoma

and one case of Langerhans cell histiocytosis (LCH). Interest-

ingly, EBV association was detected by in situ EBV-encoded

RNA 1 (EBER-1) hybridization in both cases of peripheral T

cell lymphoma and in the Hodgkin lymphoma case. Besides

malignancy-associated HLH, this study noted two patients

with autoimmune disease-associated HLH: an 11-year-old

female with SLE and a 13-year-old male with JRA. Both

patients had unusually high levels of hyperferritinemia (1130

and 5200 lg/l) and both are still alive, receiving therapy for

autoimmune diseases. This study also noted a high rate of

pleural and abdominal effusion, probably due to the conse-

quences of severe liver damage, immune reactions, or infiltra-

tion of the lymphatic system (Akcam et al, 2007). Our study

also noted the presence of neurological disorders in three HLH

cases. The development of central nervous system involvement

in HLH, which has received increasing attention in recent

years, possibly involves infiltration of the brain by lympho-

histiocytes, or therapeutic events (Akiyoshi et al, 2006; Horne

et al, 2008).

The high percentage of early deaths and reactivation deaths

in our HLH patients (14/28, 50%) drove us to clarify and

compare the potential prognostic factors between the early

response group and the early death group. Consistent with

previous reports (Tang et al, 2008), we noted that high serum

concentrations of IFN-c, IL-10, MIG and IP-10 were associ-

ated with early death (P = 0Æ03; P = 0Æ04; P = 0Æ02 and

P = 0Æ01, respectively). Therefore, measuring the levels of

these specific cytokine and chemokine profiles may provide

predictive markers of disease activity and severity for thera-

peutic consideration. Finally, some of our patients received

suboptimal therapy in this study due to the shortage of

medications, such as etoposide and IVIG, during the early

phase of the study, which partly explains the poor outcome of

the treatments. The high rate of recurrence and of reactivation

death in this study suggests that a more aggressive regimen or

bone marrow transplantation may be needed in the future.

In conclusion, this prospective, collaborative study provides

a comprehensive analysis and characterization of HLH cases in

Vietnamese children. Early diagnosis of HLH, measurements

of specific cytokine profiles, and a more appropriate thera-

peutic regimen may be needed in the future.

Acknowledgement

This project is sponsored by grants from National Health

Research Institutes, Taiwan.

References

Akcam, M., Artan, R., Yilmaz, A. & Akkaya, B. (2007) An unusual

cause of ascites: hemophagocytic lymphohistiocytosis. Indian Pedi-

atrics, 44, 371–374.

Akiyoshi, K., Hamada, Y., Yamada, H., Kojo, M. & Izumi, T. (2006)

Acute necrotizing encephalopathy associated with hemophagocytic

syndrome. Pediatric Neurology, 34, 315–318.

Chen, C.J., Huang, Y.C., Jaing, T.H., Hung, I.J., Yang, C.P., Chang,

L.Y. & Lin, T.Y. (2004) Hemophagocytic syndrome: a review of 18

pediatric cases. Journal of Microbiology, Immunology, and Infection,

37, 157–163.

Chuang, H.C., Lay, J.D., Hsieh, W.C., Wang, H.C., Chang, Y.,

Chuang, S.E. & Su, I.J. (2005) Epstein-Barr virus LMP1 inhibits

the expression of SAP gene and upregulates Th1 cytokines in the

pathogenesis of hemophagocytic syndrome. Blood, 106, 3090–

3096.

Chuang, H.C., Lay, J.D., Hsieh, W.C. & Su, I.J. (2007a) Pathogenesis

and mechanism of disease progression from hemophagocytic lym-

phohistiocytosis to Epstein–Barr virus-associated T-cell lymphoma:

nuclear factor-kappa B pathway as a potential therapeutic target.

Cancer Science, 98, 1281–1287.

Chuang, H.C., Lay, J.D., Chuang, S.E., Hsieh, W.C., Chang, Y. & Su,

I.J. (2007b) Epstein–Barr virus (EBV) latent membrane protein-1

down-regulates tumor necrosis factor-alpha (TNF-alpha) receptor-1

and confers resistance to TNF-alpha-induced apoptosis in T cells:

implication for the progression to T-cell lymphoma in EBV-asso-

ciated hemophagocytic syndrome. American Journal of Pathology,

170, 1607–1617.

Fujiwara, F., Hibi, S. & Imashuku, S. (1993) Hypercytokinemia in

hemophagocytic syndrome. The American Journal of Pediatric

Hematology/Oncology, 15, 92–98.

Goransdotter Ericson, K., Fadeel, B., Nilsson-Ardnor, S., Soderhall, C.,

Samuelsson, A., Janka, G., Schneider, M., Gurgey, A., Yalman, N.,

Revesz, T., Egeler, R., Jahnukainen, K., Storm-Mathiesen, I., Har-

aldsson, A., Poole, J., de Saint Basile, G., Nordenskjold, M. &

Henter, J. (2001) Spectrum of perforin gene mutations in familial

hemophagocytic lymphohistiocytosis. American Journal of Human

Genetics, 68, 590–597.

Gupta, A.A., Tyrrell, P., Valani, R., Benseler, S., Abdelhaleem, M. &

Weitzman, S. (2009) Experience with hemophagocytic lymphohist-

iocytosis/macrophage activation syndrome at a single institution.

Journal of Pediatric Hematology/Oncology, 31, 81–84.

Gurgey, A., Secmeer, G., Tavil, B., Ceyhan, M., Kuskonmaz, B., Cengiz,

B., Ozen, H., Kara, A., Cetin, M. & Gumruk, F. (2005) Secondary

hemophagocytic lymphohistiocytosis in Turkish children. Pediatric

Infectious Disease Journal, 24, 1116–1117.

L. T. My et al

308 ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310

Gutierrez, A., Solano, C., Ferrandez, A., Marugan, I., Terol, M.J., Benet,

I., Tormo, M., Bea, M.D. & Rodriguez, J. (2003) Peripheral T-cell

lymphoma associated consecutively with hemophagocytic lym-

phohistiocytosis and hypereosinophilic syndrome. European Journal

of Haematology, 71, 303–306.

Henter, J.I., Arico, M., Elinder, G., Imashuku, S. & Janka, G. (1998)

Familial hemophagocytic lymphohistiocytosis. Primary hemophag-

ocytic lymphohistiocytosis. Hematology/Oncology Clinics of North

America, 12, 417–433.

Henter, J.I., Samuelsson-Horne, A., Arico, M., Egeler, R.M., Elinder,

G., Filipovich, A.H., Gadner, H., Imashuku, S., Komp, D., Ladisch,

S., Webb, D. & Janka, G. (2002) Treatment of hemophagocytic

lymphohistiocytosis with HLH-94 immunochemotherapy and bone

marrow transplantation. Blood, 100, 2367–2373.

Henter, J.I., Horne, A., Arico, M., Egeler, R.M., Filipovich, A.H.,

Imashuku, S., Ladisch, S., McClain, K., Webb, D., Winiarski, J. &

Janka, G. (2007) HLH-2004: diagnostic and therapeutic guidelines

for hemophagocytic lymphohistiocytosis. Pediatric Blood & Cancer,

48, 124–131.

Horne, A., Trottestam, H., Arico, M., Egeler, R.M., Filipovich, A.H.,

Gadner, H., Imashuku, S., Ladisch, S., Webb, D., Janka, G. &

Henter, J.I. (2008) Frequency and spectrum of central nervous

system involvement in 193 children with haemophagocytic lym-

phohistiocytosis. British Journal of Haematology, 140, 327–335.

Imashuku, S. (2002) Clinical features and treatment strategies of

Epstein–Barr virus-associated hemophagocytic lymphohistiocytosis.

Critical Reviews in Oncology/Hematology, 44, 259–272.

Imashuku, S., Hibi, S., Fujiwara, F., Ikushima, S. & Todo, S. (1994)

Haemophagocytic lymphohistiocytosis, interferon-gamma-naemia

and Epstein–Barr virus involvement. British Journal of Haematology,

88, 656–658.

Imashuku, S., Tabata, Y., Teramura, T. & Hibi, S. (2000) Treatment

strategies for Epstein–Barr virus-associated hemophagocytic lym-

phohistiocytosis (EBV-HLH). Leukaemia & Lymphoma, 39, 37–49.

Imashuku, S., Teramura, T., Tauchi, H., Ishida, Y., Otoh, Y., Sawada,

M., Tanaka, H., Watanabe, A., Tabata, Y., Morimoto, A., Hibi, S. &

Henter, J.I. (2004) Longitudinal follow-up of patients with Epstein-

Barr virus-associated hemophagocytic lymphohistiocytosis. Hae-

matologica, 89, 183–188.

Ishii, E., Ohga, S., Aoki, T., Yamada, S., Sako, M., Tasaka, H., Kuwano,

A., Sasaki, M., Tsunematsu, Y. & Ueda, K. (1991) Prognosis of

children with virus-associated hemophagocytic syndrome and

malignant histiocytosis: correlation with levels of serum interleukin-

1 and tumor necrosis factor. Acta Haematologica, 85, 93–99.

Ishii, E., Ohga, S., Imashuku, S., Yasukawa, M., Tsuda, H., Miura, I.,

Yamamoto, K., Horiuchi, H., Takada, K., Ohshima, K., Nakamura,

S., Kinukawa, N., Oshimi, K. & Kawa, K. (2007) Nationwide survey

of hemophagocytic lymphohistiocytosis in Japan. International

Journal of Hematology, 86, 58–65.

Janka, G.E. (2007) Familial and acquired hemophagocytic lymphoh-

istiocytosis. European Journal of Pediatrics, 166, 95–109.

Kogawa, K., Lee, S.M., Villanueva, J., Marmer, D., Sumegi, J. &

Filipovich, A.H. (2002) Perforin expression in cytotoxic lympho-

cytes from patients with hemophagocytic lymphohistiocytosis and

their family members. Blood, 99, 61–66.

Krenova, Z., Sterba, J., Blatny, J., Kren, L. & Slany, J. (2007) A case of

anaplastic large cell lymphoma-induced hemophagocytic lymphoh-

istiocytosis in an adolescent female. Pediatric Blood & Cancer, 49,

1056.

Lee, J.S., Kang, J.H., Lee, G.K. & Park, H.J. (2005) Successful treatment

of Epstein–Barr virus-associated hemophagocytic lymphohistio-

cytosis with HLH-94 protocol. Journal of Korean Medical Science, 20,

209–214.

Lee, W.I., Chen, S.H., Hung, I.J., Yang, C.P., Jaing, T.H., Chen, C.J., Li,

S.P. & Huang, J.L. (2009) Clinical aspects, immunologic assessment,

and genetic analysis in Taiwanese children with hemophagocytic

lymphohistiocytosis. Pediatric Infectious Disease Journal, 28, 30–34.

Lu, P.L., Hsiao, H.H., Tsai, J.J., Chen, T.C., Feng, M.C., Chen, T.P. &

Lin, S.F. (2005) Dengue virus-associated hemophagocytic syndrome

and dyserythropoiesis: a case report. The Kaohsiung Journal of

Medical Sciences, 21, 34–39.

Mathew, L.G., Cherian, T., Sudarshanam, A., Korah, I., Kumar, N.K. &

Raghupathy, P. (2000) Hemophagocytic lymphohistiocytosis: a case

series. Indian Pediatrics, 37, 526–531.

Mischler, M., Fleming, G.M., Shanley, T.P., Madden, L., Levine, J.,

Castle, V., Filipovich, A.H. & Cornell, T.T. (2007) Epstein–Barr

virus-induced hemophagocytic lymphohistiocytosis and X-linked

lymphoproliferative disease: a mimicker of sepsis in the pediatric

intensive care unit. Pediatrics, 119, e1212–e1218.

Nakamura, I., Nakamura-Uchiyama, F., Komiya, N. & Ohnishi, K.

(2009) [A case of dengue fever with viral-associated hemophagocytic

syndrome]. Kansenshogaku Zasshi, 83, 60–63.

Ohga, S., Matsuzaki, A., Nishizaki, M., Nagashima, T., Kai, T., Suda,

M. & Ueda, K. (1993) Inflammatory cytokines in virus-associated

hemophagocytic syndrome. Interferon-gamma as a sensitive indi-

cator of disease activity. The American Journal of Pediatric Hema-

tology/Oncology, 15, 291–298.

Ohno, T., Ueda, Y., Nagai, K., Takahashi, T., Konaka, Y., Takamatsu,

T., Suzuki, T., Sasada, M. & Uchiyama, T. (2003) The serum cyto-

kine profiles of lymphoma-associated hemophagocytic syndrome: a

comparative analysis of B-cell and T-cell/natural killer cell lym-

phomas. International Journal of Hematology, 77, 286–294.

Osugi, Y., Hara, J., Tagawa, S., Takai, K., Hosoi, G., Matsuda, Y., Ohta,

H., Fujisaki, H., Kobayashi, M., Sakata, N., Kawa-Ha, K., Okada, S.

& Tawa, A. (1997) Cytokine production regulating Th1 and Th2

cytokines in hemophagocytic lymphohistiocytosis. Blood, 89, 4100–

4103.

Sayos, J., Wu, C., Morra, M., Wang, N., Zhang, X., Allen, D., van

Schaik, S., Notarangelo, L., Geha, R., Roncarolo, M.G., Oettgen, H.,

De Vries, J.E., Aversa, G. & Terhorst, C. (1998) The X-linked lym-

phoproliferative-disease gene product SAP regulates signals induced

through the co-receptor SLAM. Nature, 395, 462–469.

Srichaikul, T., Punyagupta, S., Kanchanapoom, T., Chanokovat, C.,

Likittanasombat, K. & Leelasiri, A. (2008) Hemophagocytic

syndrome in Dengue hemorrhagic fever with severe multiorgan

complications. Journal of the Medical Association of Thailand, 91,

104–109.

Su, I.J. (2008) Perspectives on the pathogenesis and therapy of he-

mophagocytic syndrome. Journal of the Formosan Medical Associa-

tion, 107, 277–280.

Su, I.J., Chen, R.L., Lin, D.T., Lin, K.S. & Chen, C.C. (1994) Epstein–

Barr virus (EBV) infects T lymphocytes in childhood EBV-associated

hemophagocytic syndrome in Taiwan. American Journal of Patho-

logy, 144, 1219–1225.

Sullivan, K.E., Delaat, C.A., Douglas, S.D. & Filipovich, A.H. (1998)

Defective natural killer cell function in patients with hemophago-

cytic lymphohistiocytosis and in first degree relatives. Pediatric

Research, 44, 465–468.

HLH in Vietnam

ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310 309

Tan, E.M., Cohen, A.S., Fries, J.F., Masi, A.T., McShane, D.J., Roth-

field, N.F., Schaller, J.G., Talal, N. & Winchester, R.J. (1982) The

1982 revised criteria for the classification of systemic lupus erythe-

matosus. Arthritis and Rheumatism, 25, 1271–1277.

Tang, Y., Xu, X., Song, H., Yang, S., Shi, S., Wei, J., Pan, B., Zhao, F.,

Liao, C. & Luo, C. (2008) Early diagnostic and prognostic signifi-

cance of a specific Th1/Th2 cytokine pattern in children with

haemophagocytic syndrome. British Journal of Haematology, 143,

84–91.

Ueda, I., Ishii, E., Morimoto, A., Ohga, S., Sako, M. & Imashuku, S.

(2006) Correlation between phenotypic heterogeneity and gene

mutational characteristics in familial hemophagocytic lymphohisti-

ocytosis (FHL). Pediatric Blood & Cancer, 46, 482–488.

Veerakul, G., Sanpakit, K., Tanphaichitr, V.S., Mahasandana, C. &

Jirarattanasopa, N. (2002) Secondary hemophagocytic lymphohist-

iocytosis in children: an analysis of etiology and outcome. Journal of

the Medical Association of Thailand, 85(Suppl. 2), S530–S541.

Yamada, K., Yamamoto, Y., Uchiyama, A., Ito, R., Aoki, Y., Uchida, Y.,

Nagasawa, H., Kimura, H., Ichiyama, T., Fukao, T. & Kohno, Y.

(2008) Successful treatment of neonatal herpes simplex-type 1

infection complicated by hemophagocytic lymphohistiocytosis and

acute liver failure. Tohoku Journal of Experimental Medicine, 214, 1–5.

L. T. My et al

310 ª 2009 Blackwell Publishing Ltd, British Journal of Haematology, 148, 301–310