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\begin{document}

\title{Secondary Hemophagocytic Lymphohistiocytosis During the Disease Course
of Multisystem Inflammatory Syndrome in Children}



\author[1]{Sibel Laçinel Gürlevik}%
\author[1]{Tekin Aksu}%
\author[1]{Seza Ozen}%
\author[1]{Selman Kesici}%
\author[1]{Fatma Gümrük}%
\author[1]{Yasemin Ozsurekci}%
\affil[1]{Hacettepe University Faculty of Medicine}%


\vspace{-1em}



  
  \date{June 17, 2024}


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\selectlanguage{english}
\begin{abstract}
Multisystem inflammatory syndrome in children (MIS-C) is a serious
disease with different and various clinical presentations. We may define
it as a ``game changer'' syndrome. Individual clinical entity is seen in
each patient and challenge clinicians. We have seen that patients with
perforated appendicitis findings on physical examinations who may
require urgent surgery recover with steroids or immunomodulators.
Treatment must be tailored to each patient individually. Here, we will
present three children with a diagnosis of MIS-C who have
hemophagocytosis in bone marrow aspiration, which are clinically
suggestive of secondary hemophagocytic lympho-histiocytosis (HLH).%
\end{abstract}%



\sloppy


\textbf{INTRODUCTION}

During coronavirus disease 2019 (COVID-19) pandemic millions of people
have been infected worldwide. Severe acute respiratory syndrome
coronavirus 2 (SARS-CoV-2) affects people of all ages. Although it is
known that children have a milder course of illness compared to adults,
it has been observed that a severe newly described clinical picture
which is called multisystem inflammatory syndrome in children (MIS-C)
has been reported (1-5). Herein we describe unique cases of MIS-C with
hemophagocytosis in bone marrow aspirate smears.

\textbf{CASE 1}

A four-year-old previously healthy boy referred with the diagnosis of
the Kawasaki Disease. He had no known exposure to SARS-CoV-2. He was
hospitalized with the complaint of fever for more than seven days and
severe abdominal pain. On admission he had rashes, conjunctival
injection, edema on the extremities, abdominal tenderness and
hepatosplenomegaly. On abdominal ultrasonography (USG),
hepatosplenomegaly, hydrops of gallbladder and free fluid in the peri
splenic area were reported. He had normal echocardiographic findings.
His SARS-CoV-2 RT-PCR test was negative for two times during 7-day
course of illness. Intravenous immunoglobulin (IVIg) (2gr/kg/day) and
acetylsalicylic acid were started. On the second day of hospitalization,
he had headache and nuchal rigidity. Lumbar puncture (LP) was performed.
Cerebrospinal fluid (CSF) biochemistry was within normal ranges for age
(CSF protein:40 mg/dL, CSF glucose: 58mg/dL, blood glucose level:72
mg/dL. No cell was detected on the microscopy and culture was negative.
Bone marrow aspiration (BMA) revealed hemophagocytosis with all normal
lineages. Later on, he was hypotensive and had respiratory distress.
Intensive Care Unit (ICU) admission was required, so he was referred to
our hospital.

Laboratory work-up was notable for thrombocytopenia and lymphopenia
(Table 1). Elevated cardiac enzymes; brain natriuretic peptide (BNP)
1405.5 pg/ml, troponin -I 109.5 ng/L, creatinine kinase myocardial band
(CK-MB) 5.2 \selectlanguage{greek}µ\selectlanguage{english}g/l were present. High C reactive protein (CRP) and
procalcitonin level were present. He had hypoalbuminemia,
hypofibrinogenemia, high D-dimer and ferritin level (Table 1) (Figure 1
a).

Echocardiographic findings were normal {[}Ejection fraction (EF):
70\%{]} with no coronary artery abnormalities. Bone marrow aspiration
smears from referring hospital were re-evaluated and revealed
hemophagocytic cells with no malignant cells (Figure 2 A\&B). His
SARS-CoV-2 IgG level was high; 8.9.

These features were compatible with MIS-C, methylprednisolone
(2mg/kg/day) and anakinra (4mg/kg/day) were administered. He was started
on antibiotics (vancomycin, ceftriaxone, and clarithromycin),
favipiravir and enoxaparin (1mg/kg/day). On the fifth day of the
treatment abnormal physical findings were resolved. He was well with
improved laboratory findings.

\textbf{CASE 2}

A three-year-old previously healthy girl admitted to our hospital with
fever of four days. Her father attended the funeral of relative passed
away due to COVID-19 one month ago, after which he had fever for two
days. Apart from this, there was no other exposure that would suggest
COVID-19. Her physical examination revealed 38.1\selectlanguage{ngerman}°C fever and tachycardia
(160/min). There were widespread maculopapular rashes on her body and
abdominal tenderness with no organomegaly. Her lips were red and
cracked. She had edema on her eyelids and extremities.

Laboratory findings showed marked thrombocytopenia, lymphopenia and
elevated liver enzymes (Table1). High CRP, procalcitonin and interleukin
6 (IL-6) levels were present. Ferritin and D-dimer level were high
(Table 1). Cardiac enzymes were in between normal ranges. Her SARS-CoV-2
RT-PCR was negative and SARS-CoV-2 IgG was positive; 9.1.
Echocardiographic findings were normal (EF: 77\%).

She was hospitalized with the diagnosis of MIS-C and started on IVIg
(2gr/kg/day), methylprednisolone (2mg/kg/day), anakinra (4mg/kg/day) and
enoxaparin (1mg/kg/day). Hemophagocytosis with increased number of free
histiocytes was present on BMA (Figure 2 C\&D). On the fifth day of the
treatment, she was well with improved laboratory findings.

\textbf{CASE 3}

A 17-year-old-boy with a sensorineural hearing loss and no chronic
systemic illness admitted to a hospital with a 7-day history of fever,
abdominal pain and vomiting. Lumbar puncture was performed due to severe
headache and CSF findings were normal. Later on, he had respiratory
distress, hypotension and admitted to ICU. Diminished left ventricular
function (EF:33\%) was present. Bilateral consolidations compatible with
COVID-19 pneumonia and pleural effusion were reported in the thoracic
computed tomography (CT). He had a history of contact with a relative
who had COVID-19 three weeks ago. During 10-day hospitalization
SARS-CoV-2 RT-PCR was negative for three times. He was referred to our
hospital for potential extracorporeal membrane oxygenation (ECMO)
requirement. On admission he was intubated and had invasive respiratory
support. He had 38.3°C fever with tachycardia (146/min). He was
hypotensive and started on inotropic treatment. He had edema on the
extremities, hepatomegaly and diminished respiratory sounds.

Marked laboratory findings are given in Table 1. Notably elevated
cardiac enzymes were present; BNP was 5704 pg/mL, troponin 5927 ng/L,
CK-MB 11.8 \selectlanguage{greek}µ\selectlanguage{english}g/L, myoglobin 559 \selectlanguage{greek}µ\selectlanguage{english}g/L. Left ventricular hypertropia and
minimal pericardial effusion were reported on echocardiography (EF:
45\%).

His SARS-CoV-2 RT-PCR was negative and SARS-CoV-2 IgG was positive; 8.9.
Plasma exchange applied. He was started on plasma exchange, IVIg,
steroid, anakinra, enoxaparin and antibiotics (ceftriaxone, teicoplanin
and clarithromycin). Bone marrow aspiration revealed hemophagocytosis
and increased number of free histiocytes (Figure 2 E\&F). After
two-month hospital stay, he was discharged very well.

\textbf{DISCUSSION}

Hemophagocytic lymphohistiocytosis (HLH) is an acute and rapidly
progressive systemic inflammatory disorder characterized by cytopenia,
excessive cytokine production and hyperferritinemia. Hemophagocytic
lymphohistioctytosis can develop by genetic defects in lymphocyte
cytotoxicity which is depicted as primary or familial HLH. Infections,
malignancies, rheumatologic diseases or immune compromising and/or
activating therapies may also trigger HLH (6).

Patients with HLH have dramatically elevated levels of numerous serum
proinflammatory cytokines. This clinical condition is thought to be a
cytokine storm syndrome. Besides typical laboratory findings, red blood
cells, platelets, or white blood cells can be observed within the
cytoplasm of the macrophages of bone marrow specimens (7).

Fever and organomegaly are the most common presenting symptoms.
Cytopenia, especially anemia and thrombocytopenia, hypertriglyceridemia,
hypofibrinogenemia and hyperferritinemia are typical laboratory
findings. Recent reports suggest that the cytokine storm caused by
SARS-CoV-2, has significant similarities with the clinical and
laboratory findings of HLH. Case series have raised concerns about a
hyperinflammatory process associated with COVID-19 in children, defined
as MIS-C (1-5). Most children had fever lasting more than four days, and
common presenting symptoms such as rashes, conjunctival injection,
gastrointestinal symptoms and lethargy. Cardiac manifestations
especially left ventricular failure and organomegaly may occur. Mostly
high level of inflammatory markers, such as CRP, ESR, procalcitonin and
ferritin occur. Many patients have lymphopenia and thrombocytopenia.
Although clinical and laboratory findings of MIS-C resemble HLH, there
are a few studies reporting that MIS-C is immunologically different from
HLH (8, 9). Both HLH and MIS-C are conditions which patients deteriorate
rapidly require rapid diagnosis and appropriate treatment (7). The
treatment can be lifesaving.

Despite appropriate treatment, cytopenia, marked elevation of
inflammatory markers and additionally elevated ferritin, triglyceride
and D-dimer were seen in these three cases suggesting HLH occurrence.
Bone marrow evaluation was performed and hemophagocytosis was observed.
Hemophagocytosis has been reported for adults in association with severe
COVID-19 disease however there are no reported data for children
demonstrated by bone marrow aspiration (10).

Being a newly described entity, every patient may have different
presentation and clinical course therefore treatment must be tailored to
each patient individually. These cases highlight that MIS-C and HLH have
similar findings, while deeper thrombocytopenia, very high ferritin and
d-Dimer levels should suggest HLH.

\textbf{CONFLICT OF INTERES}

The authors declare that there is no conflict of interest.

\textbf{AUTHOR CONTRIBUTIONS}

Sibel Lacinel Gurlevik, Tekin Aksu, Seza Ozen, Selman Kesici, Fatma
Gumruk, Yasemin Ozsurekci contributed to clinical care and primary
manuscript composition.

\textbf{REFERENCES}

\begin{enumerate}
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\item
  Riphagen S, Gomez X, Gonzales-Martinez C, Wilkinson N, Theocaris P.
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\item
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\item
  Lee P Y, Day-Lewis M, Henderson L A, et al. Distinct clinical and
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\item
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\item
  Gadfred-Cato S, Bryant B, Leung J, et al. COVID-19 Associated
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\item
  Al-Samkari H, Berliner N. Hemophagocytic Lymphohistiocytosis. Annu Rev
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\item
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  Carter MJ, Fish M, Jennings A, et al. Peripheral immunophenotypes in
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\end{enumerate}

\textbf{Figure Legend:}

\textbf{FIGURE 1.}

Laboratory findings of cases during hospitalization period.

\textbf{A.} Case 1

\textbf{B.} Case 2

\textbf{C.} Case 3

(Arrows show bone marrow aspiration day.)

\textbf{FIGURE 2.}

Hemophagocytosis on bone marrow aspirates.

\textbf{A} \& \textbf{B:} Case 1

\textbf{C} \&\textbf{D} : Case 2

\textbf{E} \&\textbf{F} Case 3

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