Fulminant Necrotizing Enterocolitis and Multiple Organ Dysfunction in a Toddler with Mitochondrial DNA Depletion Syndrome-13

Abstract

Necrotizing enterocolitis (NEC) is exceptional after the neonatal period. A toddler with encephalopathy, mitochondrial myopathy, and hypertrophic cardiomyopathy developed fatal NEC and multiple organ dysfunction within 48 hours of the introduction of enteral feeding. She was subsequently found to have pathogenic mutations in FBXL4 , a cause of mitochondrial DNA depletion syndrome-13. Intestinal dysmotility in the context of deficient mitochondrial respiration may have contributed to the development of NEC. Current paradigms call for early introduction of enteral nutrition to reinstate energy homeostasis. Enteral feeding should be administered with caution during metabolic crises of patients with mitochondrial DNA depletion syndromes.

Introduction

The heterogeneous clinical manifestations of metabolic diseases, often presenting as an acute critical illness, represent a diagnostic challenge. ¹ Here, we report a 19-month-old girl who died of fulminant necrotizing enterocolitis (NEC) and was found to have biallelic FBXL4 mutations, a cause of mitochondrial DNA depletion syndrome-13 (MTDPS13). ^{2 3 4 5} To our knowledge, this is the first report of NEC in mitochondrial DNA depletion syndromes, an increasingly recognized cause of mitochondrial diseases. The best-documented targets of mitochondrial disorders are postmitotic tissues with high rates of energy utilization, such as the brain, heart, and skeletal muscles. Tissues with significant cell turnover, such as intestinal mucosa, are often spared. ^{6 7} Enteral nutrition is therefore encouraged to prevent catabolism as in most critical illnesses. ⁸

The development of NEC in toddlers is exceptional, and we discuss pathogenic mechanisms that may have led to intestinal necrosis in this patient. We compare this patient with previous reports, according to CARE guidelines. ⁹

Case Report

This girl was born at term to nonconsanguineous French Canadian parents. Antenatal ultrasounds showed mega cisterna magna with subependymal hemorrhage of the right thalamocaudate region, which was confirmed after birth by cerebral ultrasound. Her birth weight was 2.6 kg (third centile). Feeding difficulties were present from birth. At 6 weeks of age, she was admitted to the pediatric intensive care unit (PICU) for respiratory distress and feeding problems. Metabolic acidosis was noted (capillary pH 7.24; pCO ₂ 19 mm Hg; bicarbonates 11 mmol/L), with increased lactate in serum (14 mmol/L, normal 0.6–3.2 mmol/L) and cerebrospinal fluid (14 mmol/L), mild hyperammonemia (104 μmol/L, normal 5–55 μmol/L), and neutropenia (0.2 × 10 ⁹ /L, normal 1.5–5.5 × 10 ⁹ /L). Broad-spectrum antibiotics were administered until blood cultures were confirmed to be sterile. She received sodium bicarbonate, L-carnitine, sodium benzoate/phenylacetate, coenzyme Q10, thiamine, riboflavin, and biotin. She had a severe neurodevelopmental delay, axial hypotonia, and muscular weakness. Diagnostic investigations performed from birth to 18 months of age are presented in Table 1 . Of note, quadriceps muscle biopsy revealed myopathy with structurally abnormal mitochondria and deficiencies of multiple respiratory chain complexes. Cerebral imaging ( Figs. 1 and 2 ) showed mild cerebral dysplasia and signal abnormalities in the basal ganglia. She was hospitalized nine times for feeding difficulties. From birth to 4 months of age, maternal milk was given every 3 hours (bottle/gastric feeding tube). A regular formula milk (0.67 cal/mL) was then started providing 780 mL/d (95 cal/kg/d). Gastrostomy was performed at 8 months due to severe gastroesophageal reflux and failure to thrive (weight <3rd; length 5th–15th; head <3rd). From 8 to 12 months, an enriched (0.8 cal/mL) milk Eq. (3 × 150 mL; 40 mL/h at night) was given (810 mL/d; 85 cal/kg/d). Thereafter, until admission, she was fed by gavage (Nutren Jr 1 cal/mL; 150 mL three times a day plus nocturnal gavages for a total of 700 mL/d; 95 cal/kg/d). Weight gain remained slow: birth (2.6 kg), 5 months (5.3 kg), 12 months (7.6 kg), and 19 months (9.3 kg). She also presented persistent metabolic acidosis, hypocapnia, and significant hyperoxia ( Fig. 3 ).

Table 1. Diagnostic evaluation from birth to 18 months.

Encephalomyopathy
Cerebral magnetic resonance imaging	Fig. 1
Quadriceps muscle biopsy (age 6 wk)	Histology : Nonspecific myopathy
	Ultrastructure : Abnormal pleomorphic mitochondria including some megamitochondria. Discrete increases of glycogen and of lipid droplets
	Biochemical testing : Reduced combined oxidative phosphorylation (low enzymatic activity ratios of CI/CII: 0.19 [Reference: 0.26], CIII/CII: 5.73 [6.10], CIV/CII: 3.24 [6.40], and CII + CIII/citrate synthase: 0.3 [0.5]). The baseline expression of CI/CII and CIV/CII were decreased on blue native polyacrylamide gel electrophoresis
Creatine kinase	Intermittent elevation (80–850 U/L)
Electrophysiology	Myopathic electromyographic pattern; normal nerve conduction
Hepatic
Liver function tests	Intermittent increase of serum alanine aminotransferase (11–642 U/L). Absence of cholestasis
Liver biopsy	Slight portal fibrosis without steatosis
Heart
Electrocardiogram	Left ventricular hypertrophy at 4 d; biventricular hypertrophy at 2 wk
Cardiac ultrasound	Normal systolic function; nonprogressive aortic dilatation (Z score, +4); hypertrophic cardiomyopathy at 6 mo with right ventricle diastolic dysfunction at 9 mo
Hematology	Episodic neutropenia and lymphopenia; normal marrow biopsy
Renal	No renal disease identified
Ophthalmology	Small cataract at 14 mo
Genetic	Homozygous frameshift mutation in FBXL4, exon 8: c.1641_1642_delTG, reference sequence NM_012160.4; (p.Cys547Ter)

Fig. 1.

Brain magnetic resonance imaging at 46 days of life. ( A ) Presence of a persistent falcine sinus (white arrow, sagittal T1) and a mega cisterna magna (white star); the latter is considered a normal variant, although previously reported in FBXL4 deficiency. ( B ) Discrete cerebellar dysplasia was present (black arrow, coronal T2), as previously reported in FBXL4 deficiency. ( C ) Nonspecific hyperintensities within basal ganglia were also noted (axial T2).

Fig. 2.

Cerebral computed tomography performed at 18 months of age. ( A ) Since the head circumference was small, dilatation of subarachnoid spaces and ventricles suggested cerebral atrophy; ( B ) hypodensities within basal ganglia were also observed (white arrow).

Fig. 3.

Evolution of the patient acid–base profile. Aging correlated weakly with ( A ) higher capillary pH ( R  = +0.24; p  < 0.01) and ( B ) pO ₂ ( R  = +0.60; p  < 0.0001), as well as ( C ) lower pCO ₂ ( R  = − 0.64; p  < 0.0001) and ( D ) bicarbonates ( R  = − 0.47; p  < 0.001). Data were obtained on room air.

At 19 months, she was admitted to the PICU for an afebrile seizure, metabolic acidosis, mild hyperammonemia, loose stools, and dehydration. Bilateral serous otitis was also noted. A rhinovirus infection was later confirmed by molecular testing of nasopharyngeal secretions. Her subsequent PICU course is summarized in Table 2 . Enteral feeding was attempted following stabilization, 20 hours after PICU admission (H ₂₀ ), and was discontinued 46 hours later (H ₆₆ ) when the patient developed hemodynamic instability. Signs of abdominal compartment syndrome developed. Exploratory laparotomy revealed severe ischemia with diffuse pneumatosis from the duodenum to the terminal ileum. The ascending colon and part of the descending colon were necrotic and subtotal colectomy was performed. Marked visceral edema was present, precluding closure of the abdomen. Postoperatively, rapidly progressive multiple organ dysfunction syndrome developed. Given the poor prognosis, supportive care was withdrawn after consultation with the parents. Septic workup was negative. Autopsy revealed the pathognomonic signs of NEC, including intestinal pneumatosis and coagulation necrosis ( Fig. 4 ). ^{10 11} Neuropathological examination showed severe brain atrophy, white matter microvacuolization, and Alzheimer type II glial changes (swollen astrocytes) compatible with mitochondrial degeneration.

Table 2. Clinical course in the PICU.

First day

(H 0 ) Lactate, 15 mmol/L; serum ammonia, 349 μmol/L; capillary pH 7.1; pCO 2 13 mm Hg; bicarbonate 4 mmol/L; pO 2 254 mm Hg on 28% FiO 2 , oxygen stopped 2 h later; urea, 8 mmol/L; creatinine, 51 μmol/L; hemoglobin, 160 g/L with normal neutrophil and lymphocyte counts; alanine aminotransferase, 240 U/L without cholestasis; creatine kinase, 222 U/L

(H 0 ) Focal secondarily generalized seizure (no epileptic activity on EEG)

(H 1 ) Cerebral CT scan ( Fig. 2A, B )

(H 2 ) Treatment with sodium benzoate/phenylacetate, L-carnitine, carglumic acid, Coenzyme Q, thiamine, riboflavine, and biotin

(H 4 ) Increased femoral SvO 2 (80%)

(H 0 –H 24 ) Rehydration: 175 mL/kg/d; intravenous sodium bicarbonate (14 mEq/kg/d)

(H 20 ) Nasoduodenal feeding (Nutren Jr and Pro-Phree in equal amounts, 1 cal/mL, protein 1.5 g/100 mL), started at 20 mL/h; plus Intralipid 20% (1 g/kg; 90 cal/24 h); total, 74 cal/kg/d

Second day

(H 24 –H 48 ) Fluids at maintenance with intravenous sodium bicarbonate (10 mEq/kg/d)

(H 29 ) Enteral feeding increased to 30 mL/h plus Intralipid 20% (1 g/kg; 90 cal/24 h)

(H 39 ) Endotracheal intubation due to persistent agitation and lactic acidosis

Third day

(H 46 ) Decreasing femoral SvO 2 : 88%; (H 51 ) 68%; (H 58 ) 58%; (H 61 ) 55%

(H 58 ) Fever (38.7°C rectal)

(H 63 ) Volume loading (50 mL/kg); (H 64 ) norepinephrine 0.05 µg/kg/min; (H 65 ) SvO 2 59%

(H 66 ) Abdominal distension; enteral feeding stopped

(H 67 ) Norepinephrine 0.07 µg/kg/min and red blood cell transfusion; femoral SvO 2 89%

(H 68 ) First arterial blood gas: pH 7.48; PaCO 2 14 mm Hg; PaO 2 199 mm Hg on 30% FiO 2 ; HCO 3 − 10 mmol/L; lactate 10 mmol/L

(H 68 ) Ultrasound shows portal vein gas; intestinal pneumatosis on abdominal radiograph

(H 72 ) Oliguria (intravesical pressure: 33 cm H 2 O) with high mean airway pressure: 15 cm H 2 O (H 64 ), 24 cm H 2 O (H 72 )

(H 75 ) Exploratory laparotomy and subtotal colectomy

Arterial PaO 2 : (H 70 ) 228 mm Hg, on 60% FiO 2 ; (H 72 ) 77, 60%; (H 75 ) 66, 60%; (H 77 ) 116, 100%; (H 79 ) 56, 70%; (H 81 ) 111, 90%; (H 83 ) 482, 100%; (H 87 ) 107, 60%.

Abbreviations: EEG, electroencephalogram; FiO ₂ , fraction of inspired oxygen; PICU, pediatric intensive care unit; SvO ₂ , venous oxygen saturation.

Fig. 4.

Histology of the ileocolectomy specimen: ( A ) healthy resection margins, 25×; ( B ) diffuse mucosal (M) necrosis and submucosal (SM) pneumatosis (circled areas), but normal muscularis propria (MP), 100×; ( C ) enterocolitis with ghosts of residual mucosa, 25×. ( D ) Colon autopsy specimen: diffuse necrosis (M), transmural hemorrhagic infarction, and extensive pneumatosis intestinalis (SM, circle); 25×. Hematoxylin and eosin stains.

Whole exome sequencing and confirmatory studies in the parents later identified a homozygous frameshift mutation in exon 8 of FBXL4 , predicted to cause premature termination (c.1641_1642delTG; p.Cys547Ter), consistent with encephalomyopathic MTDPS13. ²

Discussion

Mitochondrial DNA depletion syndromes are a heterogeneous group of hereditary, mainly autosomal recessive disorders. ¹² The causal genes are involved in mitochondrial nucleotide metabolism, mtDNA replication, or mitochondrial fusion. ^{3 4} FBXL4 is an F-box family protein that is targeted to the mitochondrial intermembrane space, where it may be a signal for mitochondrial degradation by mitophagy. ^{4 5} F-box family proteins govern phosphorylation-dependent ubiquitination, tagging proteins for degradation in proteasomes. ^{4 13} The absence of FBXL4 might cause abnormal accumulation of defective mitochondrial proteins, but the pathophysiology of the disease is not yet known in detail. ⁴ Clinically, patients with FBXL4 deficiency can present with lactic acidosis, intrauterine or postnatal growth failure, severe feeding difficulties, global developmental delay, hypotonia, seizures, stroke-like episodes, ^{14 15} cardiomyopathy, heart malformation, bone marrow depression with neutropenia or lymphopenia, elevated levels of plasma aminotransferases, and proximal renal tubular acidosis. ^{2 5} The median age of death was 2 years, with death as young as 2 days of age; the oldest survivor reported so far was 36 years old. ¹⁶

NEC classically affects premature newborns. It has been reported among full-term infants with congenital heart disease and other conditions that restrict mesenteric blood flow, and in diseases with intestinal dysmotility, such as Hirschsprung's disease or gastroschisis. ^{10 17 18} To our knowledge, this is the first report of NEC in a patient with a mitochondrial DNA depletion syndrome and the second report of NEC in mitochondrial disease. The first occurred at 10 days of age in a patient born at 35 weeks' gestation with trifunctional protein deficiency, a disease of mitochondrial fatty acid oxidation. ¹⁹

In addition to mitochondrial DNA depletion, at least three interrelated factors may have contributed to the development of NEC in this patient: intestinal dysmotility, hypocarbia, and hyperoxia. Intestinal villi are particularly susceptible to circulatory insufficiency and hypoxia. ²⁰ Intestinal dysmotility is a cardinal feature of the commonest form of mitochondrial DNA depletion, mitochondrial neurogastrointestinal encephalopathy (mitochondrial DNA depletion syndrome type 1) ⁶ and is also reported in FBXL4 encephalomyopathy. ⁴ Dilatation and uncoordinated contraction of the intestinal musculature cause increased pressure in the intestinal wall. This, in turn, can decrease gut perfusion and may have contributed to the development of NEC in this patient. ¹⁰ Hyperventilation during a metabolic crisis results in hypocapnia, as observed in this patient. Systemic hypocapnia can decrease splanchnic perfusion, ^{20 21} which may have contributed to the development of NEC.

There are no published guidelines for the enteral nutrition of critically ill mitochondrial patients. The primary aim of nutrition in mitochondrial patients is to reverse catabolism by providing sufficient energy yet avoiding the complications of providing excess nutrients; protein, glucose, and lipid intakes are adjusted to avoid hyperammonemia, hyperglycemia, and hyperlipidemia, respectively. Empirically, in mitochondrial patients, we currently start with a low enteral infusion rate (e.g., 0.25 mL/kg/h in an infant), increasing at about half the pace of other critically ill patients, particularly if intestinal dysmotility or a mitochondrial DNA depletion syndrome is suspected. Parenteral nutrition is started, including dextrose, amino acids, and intralipid, and adjusted according to the earlier parameters. Adjustments of the feeding regimen must be individualized according to the intestinal and metabolic tolerance.

Current treatment of shock in patients without a mitochondrial disease aims to reinstate homeostasis by providing adequate oxygen delivery (cardiac output; SaO ₂ ; hemoglobin), thwarting a hypothetical oxygen debt presumed to have led to lactic acidosis. In contrast, patients with mitochondrial disorders may be at risk for oxidative damage if their poorly functioning mitochondria are exposed to high levels of oxygen. Mitochondria are the main source of free radicals in cells. ²² Elegant work in a mouse model of Leigh's disease showed that hyperoxia was rapidly fatal, whereas hypoxia was protective. ²³ Therefore, it is unlikely that current clinical paradigms used in critical care management apply to patients with mitochondrial diseases. We propose that hyperoxia should be avoided. During periods of metabolic decompensation in patients with mitochondrial disease, we aim to provide the necessary nutrients, oxygen, fluids, and rest, while avoiding excesses that may increase mitochondrial stress.

Conclusion

To our knowledge, this is the first report of NEC in a patient with a mitochondrial DNA depletion syndrome. Adequate nutrition is essential for the long-term treatment of mitochondrial disease patients. In retrospect, the use of trophic enteral feeding and parenteral nutrition to provide caloric needs might have been a better approach for this patient during the acute decompensation. The occurrence of NEC after the newborn period is exceptional and raises questions about the intestine in mitochondrial diseases in general, about the occurrence of intestinal dysmotility in mitochondrial DNA depletion syndromes in particular, and the roles of enteral and parenteral nutrition in the treatment of mitochondrial DNA depletion syndrome during decompensations.

Funding Statement

Funding J.S.J. is supported by the Burroughs Wellcome Fund Career Award for Medical Scientists (1012321.01) and CIHR (141956, 390615).