Ethylmalonic encephalopathy masquerading as meningococcemia.

Ethylmalonic encephalopathy (MIM #602473) is a rare autosomal recessive metabolic condition caused by biallelic variants in ETHE1 (MIM #608451), characterized by global developmental delay, infantile hypotonia, seizures, and microvascular damage. The microvascular changes result in a pattern of relapsing spontaneous diffuse petechiae and purpura, positional acrocyanosis, and pedal edema, hemorrhagic suffusions of mucous membranes, and chronic diarrhea. Here, we describe an instructive case in which ethylmalonic encephalopathy masqueraded as meningococcal septicemia and shock. Ultrarapid whole-genome testing (time to result 60 h) and prompt biochemical analysis facilitated accurate diagnosis and counseling with rapid implementation of precision treatment for the metabolic crisis related to this condition. This case provides a timely reminder to consider rare genetic diagnoses when atypical features of more common conditions are present, with an early referral to ensure prompt biochemical and genomic diagnosis.

Mitochondrial Dysfunction and Redox Homeostasis Impairment as Pathomechanisms of Brain Damage in Ethylmalonic Encephalopathy: Insights from Animal and Human Studies.

Ethylmalonic encephalopathy (EE) is a severe intoxication disorder caused by mutations in the ETHE1 gene that encodes a mitochondrial sulfur dioxygenase involved in the catabolism of hydrogen sulfide. It is biochemically characterized by tissue accumulation of hydrogen sulfide and its by-product thiosulfate, as well as of ethylmalonic acid due to hydrogen sulfide-induced inhibition of short-chain acyl-CoA dehydrogenase. Patients usually present with early onset severe brain damage associated to encephalopathy, chronic hemorrhagic diarrhea and vascular lesions with petechial purpura and orthostatic acrocyanosis whose pathophysiology is poorly known. Current treatment aims to reduce hydrogen sulfide accumulation, but does not significantly prevent encephalopathy and most fatalities. In this review, we will summarize the present knowledge obtained from human and animal studies showing that disruption of mitochondrial and redox homeostasis may represent relevant pathomechanisms of tissue damage in EE. Mounting evidence show that hydrogen sulfide and ethylmalonic acid markedly disturb critical mitochondrial functions and induce oxidative stress. Novel therapeutic strategies using promising candidate drugs for this devastating disease are also discussed.

Lymphomatoid papulosis type E with a CD56+ immunophenotype presenting with purpura-like lesions.

Lymphomatoid papulosis (LyP) type E is a recently described variant characterized by the occurrence of large necrotic eschar-like lesions displaying microscopically angioinvasive and angiodestructive infiltrates of CD30+ lymphocytes, frequently coexpressing CD8. Rare cases of LyP type E with a CD56+ immunophenotype have been described. Herein, we describe a 36-year-old woman with LyP type E, characterized by purpura-like lesions on her left ankle. Initially, she presented with left ankle swelling, petechiae and ecchymosis, and rapidly developing necrotic papules, all of which resolved spontaneously over a period of a few months without intentional therapy. Biopsy revealed CD30 and CD56 positive atypical cell infiltrates with marked angiocentricity and angiodestruction. Awareness of this rare LyP variant and its correct recognition, even if the clinical presentation is unusual, is important to avoid aggressive treatment.

Bateman purpura (dermatoporosis): a localized scurvy treated by topical vitamin C - double-blind randomized placebo-controlled clinical trial.

BACKGROUND: Bateman purpura is characterized by diffuse senile skin atrophy, senile purpura and spontaneous stellar pseudocicatrices. Cutaneous changes in the course of ageing have been related to lower levels of ascorbic acid into the dermis of elderly people. OBJECTIVE: In this study, we postulate that senile purpura could be linked to dermal vitamin C deficiency and could be corrected by topical administration of this vitamin. METHODS: A 12-weeks, hemi-member (forearm or leg), randomized double-blind comparative study was conducted in 18 patients with Bateman purpura aged over than 60 years. At each visit, clinical assessment and biometrological measurements were performed. Clinical examination and scoring by experts showed a significant improvement on the vitamin C-treated side compared with the control, with reduction of haemorrhage areas, increase of dermal thickness. RESULTS: Twice-daily application of 5% topical vitamin C led to a clinically apparent improvement of the skin symptoms and allows beneficial effects on skin elasticity and thickness. Bateman purpura, a classical sign of photoaging whose origin has not clearly been recognized could be improved by vitamin C applied on to the skin. CONCLUSION: These results confirm the hypothesis of the underlying role of vitamin C deficiency in the determinism of Bateman purpura.

Bioenergetics dysfunction, mitochondrial permeability transition pore opening and lipid peroxidation induced by hydrogen sulfide as relevant pathomechanisms underlying the neurological dysfunction characteristic of ethylmalonic encephalopathy.

Hydrogen sulfide (sulfide) accumulates at high levels in brain of patients with ethylmalonic encephalopathy (EE). In the present study, we evaluated whether sulfide could disturb energy and redox homeostasis, and induce mitochondrial permeability transition (mPT) pore opening in rat brain aiming to better clarify the neuropathophysiology of EE. Sulfide decreased the activities of citrate synthase and aconitase in rat cerebral cortex mitochondria, and of creatine kinase (CK) in rat cerebral cortex, striatum and hippocampus supernatants. Glutathione prevented sulfide-induced CK activity decrease in the cerebral cortex. Sulfide also diminished mitochondrial respiration in cerebral cortex homogenates, and dissipated mitochondrial membrane potential (ΔΨm) and induced swelling in the presence of calcium in brain mitochondria. Alterations in ΔΨm and swelling caused by sulfide were prevented by the combination of ADP and cyclosporine A, and by ruthenium red, indicating the involvement of mPT in these effects. Furthermore, sulfide increased the levels of malondialdehyde in cerebral cortex supernatants, which was prevented by resveratrol and attenuated by glutathione, and of thiol groups in a medium devoid of brain samples. Finally, we verified that sulfide did not alter cell viability and DCFH oxidation in cerebral cortex slices, primary cortical astrocyte cultures and SH-SY5Y cells. Our data provide evidence that bioenergetics disturbance and lipid peroxidation along with mPT pore opening are involved in the pathophysiology of brain damage observed in EE.

Successful treatment of a patient with ethylmalonic encephalopathy by intravenous N-acetylcysteine.

Ethylmalonic encephalopathy (EE) is an autosomal recessive devastating metabolic disorder affecting the brain, gastrointestinal tract, peripheral vessels and rarely the other vascular organs. We report a 10-month-old girl who presented as a meningococcemia clinic but later diagnosed ethylmalonic encephalopathy. Molecular analyses revealed a homozygous c.554 T > G; p. L185R mutation in ETHE1 gene. She was only partially benefited from riboflavine, coenzyme Q10, metronidazole, N-acetylcysteine and symptomatic treatment and discharged from hospital with the sequela of oxygene dependance and developmental delay. We observed N-acetylcysteine 100 mg/kg/day intravenous infusion theraphy may be the most important drug especially in comatous EE patients.

Untargeted Metabolomics Analysis Reveals a Link between ETHE1-Mediated Disruptive Redox State and Altered Metabolic Regulation.

Defects in the gene encoding the persulfide dioxygenase ETHE1 are known to cause the severe inherited metabolic disorder ethylmalonic encephalopathy (EE). In spite of known clinical characteristics, the molecular mechanisms underlying the ETHE1 deficiency are still obscure. Herein, to further analyze the molecular phenotype of the disease, we applied an untargeted metabolomics approach on cultivated fibroblasts of EE patients for pinpointing alterations in metabolite levels. Metabolites, as direct signatures of biochemical functions, can decipher biochemical pathways involved in the cellular phenotype of patient cells. Using liquid chromatography-mass spectrometry-based untargeted metabolomics, we identified 18 metabolites that have altered levels in fibroblasts from EE patients. Our data demonstrate disrupted redox state in EE patient cells, which is reflected by significantly decreased level of reduced glutathione. Furthermore, the down-regulation of several intermediate metabolites such as the redox cofactors NAD(+) and NADH as well as Krebs cycle intermediates revealed clear alteration in metabolic regulation. Pantothenic acid and several amino acids exhibited decreased levels, whereas the ß-citrylglutamate with a putative role in brain development had an increased level in the EE patient cells. These observations indicate the severe impact of ETHE1 deficiency on cellular physiology and redox state, meanwhile suggesting targets for experimental studies on novel treatment options for the devastating metabolic disorder.

Quantitative proteomics suggests metabolic reprogramming during ETHE1 deficiency.

Deficiency of mitochondrial sulfur dioxygenase (ETHE1) causes the severe metabolic disorder ethylmalonic encephalopathy, which is characterized by early-onset encephalopathy and defective cytochrome C oxidase because of hydrogen sulfide accumulation. Although the severe systemic consequences of the disorder are becoming clear, the molecular effects are not well defined. Therefore, for further elucidating the effects of ETHE1-deficiency, we performed a large scale quantitative proteomics study on liver tissue from ETHE1-deficient mice. Our results demonstrated a clear link between ETHE1-deficiency and redox active proteins, as reflected by downregulation of several proteins related to oxidation-reduction, such as different dehydrogenases and cytochrome P450 (CYP450) members. Furthermore, the protein data indicated impact of the ETHE1-deficiency on metabolic reprogramming through upregulation of glycolytic enzymes and by altering several heterogeneous ribonucleoproteins, indicating novel link between ETHE1 and gene expression regulation. We also found increase in total protein acetylation level, pointing out the link between ETHE1 and acetylation, which is likely controlled by both redox state and cellular metabolites. These findings are relevant for understanding the complexity of the disease and may shed light on important functions influenced by ETHE1 deficiency and by the concomitant increase in the gaseous mediator hydrogen sulfide. All MS data have been deposited in the ProteomeXchange with the dataset identifiers PXD002741 (http://proteomecentral.proteomexchange.org/dataset/PXD002741) and PXD002742 (http://proteomecentral.proteomexchange.org/dataset/PXD002741).

Crystal structure of human persulfide dioxygenase: structural basis of ethylmalonic encephalopathy.

The ethylmalonic encephalopathy protein 1 (ETHE1) catalyses the oxygen-dependent oxidation of glutathione persulfide (GSSH) to give persulfite and glutathione. Mutations to the hETHE1 gene compromise sulfide metabolism leading to the genetic disease ethylmalonic encephalopathy. hETHE1 is a mono-iron binding member of the metallo-ß-lactamase (MBL) fold superfamily. We report crystallographic analysis of hETHE1 in complex with iron to 2.6 Å resolution. hETHE1 contains an αßßα MBL-fold, which supports metal-binding by the side chains of an aspartate and two histidine residues; three water molecules complete octahedral coordination of the iron. The iron binding hETHE1 enzyme is related to the 'classical' di-zinc binding MBL hydrolases involved in antibiotic resistance, but has distinctive features. The histidine and aspartate residues involved in iron-binding in ETHE1, occupy similar positions to those observed across both the zinc 1 and zinc 2 binding sites in classical MBLs. The active site of hETHE1 is very similar to an ETHE1-like enzyme from Arabidopsis thaliana (60% sequence identity). A channel leading to the active site is sufficiently large to accommodate a GSSH substrate. Some of the observed hETHE1 clinical mutations cluster in the active site region. The structure will serve as a basis for detailed functional and mechanistic studies on ETHE1 and will be useful in the development of selective MBL inhibitors.

Gastrointestinal and hepatic manifestations of mitochondrial disorders.

Inherited defects of oxidative phosphorylation lead to heterogeneous, often multisystem, mitochondrial diseases. This review highlights those mitochondrial syndromes with prominent gastrointestinal and hepatic symptoms, categorised according to underlying disease mechanism. Mitochondrial encephalopathies with major gastrointestinal involvement include mitochondrial neurogastrointestinal encephalopathy and ethylmalonic encephalopathy, which are each associated with highly specific clinical and metabolic profiles. Mitochondrial hepatopathies are most frequently caused by defects of mitochondrial DNA maintenance and expression. Although mitochondrial disorders are notorious for extreme clinical, biochemical and genetic heterogeneity, there are some pathognomonic clinical and metabolic clues that suggest a specific diagnosis, and these are highlighted. An approach to diagnosis of these complex disorders is presented, together with a genetic classification, including mitochondrial DNA disorders and nuclear-encoded defects of mitochondrial DNA maintenance and translation, OXPHOS complex assembly and mitochondrial membrane lipids. Finally, supportive and experimental therapeutic options for these currently incurable diseases are reviewed, including liver transplantation, allogeneic haematopoietic stem cell transplantation and gene therapy.

Cocaine-associated retiform purpura: a C5b-9-mediated microangiopathy syndrome associated with enhanced apoptosis and high levels of intercellular adhesion molecule-1 expression.

Cocaine-associated retiform purpura is a recently described entity characterized by striking hemorrhagic necrosis involving areas of skin associated with administration of cocaine. Levamisole, an adulterant in cocaine, has been suggested as the main culprit pathogenetically. Four cases of cocaine-associated retiform purpura were encountered in the dermatopathology practice of C. M. Magro. The light microscopic findings were correlated with immunohistochemical and immunofluorescence studies. All 4 cases showed a very striking thrombotic diathesis associated with intravascular macrophage accumulation. Necrotizing vasculitis was noted in 1 case. Striking intercellular adhesion molecule-1 (ICAM-1)/CD54 expression in vessel wall along with endothelial expression of caspase 3 and extensive vascular C5b-9 deposition was observed in all biopsies examined. Cocaine-induced retiform purpura is a C5b-9-mediated microvascular injury associated with enhanced apoptosis and prominent vascular expression of ICAM-1, all of which have been shown in prior in vitro and in vivo murine models to be a direct effect of cocaine metabolic products. Antineutrophilic cytoplasmic antibody and antiphospholipid antibodies are likely the direct sequelae of the proapoptotic microenvironment. The inflammatory vasculitic lesion could reflect the downstream end point reflective of enhanced ICAM-1 expression and the development of antineutrophilic cytoplasmic antibody. Levamisole likely works synergistically with cocaine in the propagation of this syndromic complex.

Altered sulfide (H(2)S) metabolism in ethylmalonic encephalopathy.

Hydrogen sulfide (sulfide, H(2)S) is a colorless, water-soluble gas with a typical smell of rotten eggs. In the past, it has been investigated for its role as a potent toxic gas emanating from sewers and swamps or as a by-product of industrial processes. At high concentrations, H(2)S is a powerful inhibitor of cytochrome c oxidase; in trace amounts, it is an important signaling molecule, like nitric oxide (NO) and carbon monoxide (CO), together termed "gasotransmitters." This review will cover the physiological role and the pathogenic effects of H(2)S, focusing on ethylmalonic encephalopathy, a human mitochondrial disorder caused by genetic abnormalities of sulfide metabolism. We will also discuss the options that are now conceivable for preventing genetically driven chronic H(2)S toxicity, taking into account that a complete understanding of the physiopathology of H(2)S has still to be achieved.

Characterization of patient mutations in human persulfide dioxygenase (ETHE1) involved in H2S catabolism.

Hydrogen sulfide (H(2)S) is a recently described endogenously produced gaseous signaling molecule that influences various cellular processes in the central nervous system, cardiovascular system, and gastrointestinal tract. The biogenesis of H(2)S involves the cytoplasmic transsulfuration enzymes, cystathionine ß-synthase and γ-cystathionase, whereas its catabolism occurs in the mitochondrion and couples to the energy-yielding electron transfer chain. Low steady-state levels of H(2)S appear to be controlled primarily by efficient oxygen-dependent catabolism via sulfide quinone oxidoreductase, persulfide dioxygenase (ETHE1), rhodanese, and sulfite oxidase. Mutations in the persulfide dioxgenase, i.e. ETHE1, result in ethylmalonic encephalopathy, an inborn error of metabolism. In this study, we report the biochemical characterization and kinetic properties of human persulfide dioxygenase and describe the biochemical penalties associated with two patient mutations, T152I and D196N. Steady-state kinetic analysis reveals that the T152I mutation results in a 3-fold lower activity, which is correlated with a 3-fold lower iron content compared with the wild-type enzyme. The D196N mutation results in a 2-fold higher K(m) for the substrate, glutathione persulfide.

Ethylmalonic encephalopathy associated with crescentic glomerulonephritis.

Ethylmalonic encephalopathy (EE) is a rare autosomal recessive disorder caused by mutations in the ETHE1 gene and characterized by chronic diarrhea, encephalopathy, relapsing petechiae and acrocyanosis. Nephrotic syndrome has been described in an infant with EE but the renal histology findings were not described in previous reports. We report a Palestinian girl with EE who presented with chronic diarrhea, encephalopathy, petechial rash and acrocyanosis. Subsequently, she developed progressive deterioration of renal function caused by rapidly progressive glomerulonephritis resulting in death within few days. This is, to our knowledge, the first reported occurrence of rapidly progressive glomerulonephritis in a child with ethylmalonic encephalopathy. Its presence is a serious complication associated with poor prognosis and may be explained by the diffuse vascular damage.

Persistent pigmented purpuric dermatitis: granulomatous variant.

The persistent pigmented purpuric dermatitides (PPPD) are a spectrum of dermatologic disorders characterized by petechial and pigmented macules usually confined to the lower limbs. Their etiology is unknown and several clinical variants are recognized. At the microscopic level they are characterized by angiocentric lymphocytic inflammation, red blood cell extravasation and hemosiderin deposition. A granulomatous variant of the PPPD has recently been described and to date eleven cases have been reported in the literature. In contrast to the conventional type, this variant is characterized histopathologically by ill-defined, non-necrotizing granulomata admixed with the lymphocytic inflammatory background. Although initially the granulomatous variant of the PPPD was thought to occur only in Asian patients, this sole racial predilection has not been substantiated. A tenuous association with hyperlipidemia has been noted but this requires further study. The principal importance of recognizing this entity lies in the need to include it in the histopathological differential diagnosis of granulomatous dermal infiltrates. We report here an additional patient with the granulomatous variant of PPPD and elaborate on this entity in the context of existing information in the literature.

Granulomatous pigmented purpura: report of a case and review of the literature.

The pigmented purpuric dermatoses (PPD) are a group of diseases characterized by petechiae and bronze discoloration of the skin on the lower extremities. Histopathologically, extravasation of erythrocytes with hemosiderin deposition, a perivascular lymphocytic infiltrate centered on the superficial capillaries and endothelial cell swelling are seen. The granulomatous variant of PPD (GPPD) was described in 1996 and only 10 cases have been reported since in the literature, almost exclusively in patients of East Asian descent only involving the extremities. We present a case of GPPD in a Caucasian, North American Ashkenazi Jewish woman involving the thighs, back, forearms and wrists with concomitant non-granulomatous PPD of the shins. She presented with an asymptomatic, spreading, cayenne pepper-like rash. This rash intermittently involved the lower extremities and back for 15 years, but now involves the thighs with accompanying pink papules on the back, wrists and forearms. Histopathology of the thigh and back lesions revealed superficial lichenoid granulomatous dermatitis with palisading lymphocytes and focal interface changes. Extravasated erythrocytes were seen, but vasculitis was absent. No lymphocytic atypicality was noted and T-cell gene rearrangement studies were non-clonal. This is the second reported case of GPPD in a non-Asian patient and the first case involving sites other than the extremities.

Proteomics reveals that redox regulation is disrupted in patients with ethylmalonic encephalopathy.

Deficiency of the sulfide metabolizing protein ETHE1 is the cause of ethylmalonic encephalopathy (EE), an inherited and severe metabolic disorder. To study the molecular effects of EE, we performed a proteomics study on mitochondria from cultured patient fibroblast cells. Samples from six patients were analyzed and revealed seven differentially regulated proteins compared with healthy controls. Two proteins involved in pathways of detoxification and oxidative/reductive stress were underrepresented in EE patient samples: mitochondrial superoxide dismutase (SOD2) and aldehyde dehydrogenase X (ALDH1B). Sulfide:quinone oxidoreductase (SQRDL), which takes part in the same sulfide pathway as ETHE1, was also underrepresented in EE patients. The other differentially regulated proteins were apoptosis inducing factor (AIFM1), lactate dehydrogenase (LDHB), chloride intracellular channel (CLIC4) and dimethylarginine dimethylaminohydrolase 1 (DDAH1). These proteins have been reported to be involved in encephalopathy, energy metabolism, ion transport, and nitric oxide regulation, respectively. Interestingly, oxidoreductase activity was overrepresented among the regulated proteins indicating that redox perturbation plays an important role in the molecular mechanism of EE. This observation may explain the wide range of symptoms associated with the disease, and highlights the potency of the novel gaseous mediator sulfide.

Foxp3 expression in cutaneous T-cell lymphocytic infiltrates.

BACKGROUND: The primary function of regulatory T cells (Treg cells) is to regulate the function and proliferation of immunologically responsive T cells; the transcription factor Foxp3 is expressed by this cell populace and is held to be the standard marker for Treg cells. DESIGN: A variety of cutaneous T-cell lymphocytic infiltrates were evaluated for Foxp3 expression. RESULTS: Of the 95 cases, 33 (35%) were reactive, 40 (42%) were prelymphomatous cutaneous T-cell dyscrasia and 22 were (23%) T-cell lymphoma. The reactive category included dermatomyositis, lupus erythematosus, hypersensitivity reactions and graft-vs.-host disease. The prelymphomatous dyscrasia category was represented chiefly by pityriasis lichenoides chronica (PLC) and pigmented purpuric dermatosis (PPD). The Foxp3 reactivity was less than 10% for cases of dermatomyositis and lupus erythematosus, 23% for hypersensitivity cases, 0% for graft-vs.-host disease, 16% for the dyscrasias and 11% for the lymphomas. Intermediate grade and aggressive lymphomas had very few Foxp3+ cells (< 5%). There were fewer numbers of Foxp3+ T cells in the monoclonal variants of PLC and PPD. CONCLUSIONS: T-reg cells may play a role in controlling the extent of T-cell proliferations in the skin with a lack of T-regulatory function permissive to the development of various T-cell disorders.

Vitality and age of conjunctival petechiae: the expression of P-selectin.

Confocal laser scanning microscopy (CLSM) results in displaying multiple ruptures of capillaries and venules define conjunctival petechiae as a rhexis haemorrhage. In order to distinguish between petechiae which have arisen before death as opposed to post-mortem artefacts and the establishment of the age of the vessel's damage, we carried out an investigation on a collection of samples from medico-legal cases (positive petechiae: n=65; post-mortem artefacts: n=12; control: n=19) to establish whether the expression of the endothelial adhesion molecule P-selectin as demonstrated by immunohistochemical methods is influenced by trauma before death and possibly by time. Except for cases with massive congestion in those cases where the victim had survived for around minutes, CLSM examination revealed a strong endothelial reaction at the site where the vessel had ruptured. It was impossible to detect a strong P-selectin expression signal in both those vessels which had not suffered any mechanical insult and those which had experienced post-mortem rupture. Therefore, the elevated expression of P-selectin can be used as a criterion to distinguish between those lesions which have occurred in life and those which are post-mortem artefacts.