Body composition in patients with hepatic glycogen storage diseases.

OBJECTIVES: The present study aimed to evaluate the body composition of hepatic glycogen storage disorders (GSDs) through dual energy x-ray absorptiometry. METHODS: This was an exploratory, observational, cross-sectional study. Twenty-four patients with GSD (type Ia: n = 13, Ib: n = 5, III: n = 2, and IX-α/ß/γ: n = 4; female sex: n = 13; age <8 y: n = 3, 8-19 y: n = 14, and >19 y: n = 7) were included. Three-day dietary records were collected in the week preceding dual energy x-ray absorptiometry. Body composition findings were correlated with clinical parameters, uncooked cornstarch (UCCS) regimen, dietary intake, and markers of treatment adherence. RESULTS: An elevated fat mass (FM) index was found in 16 of 21 patients (age 8-19 y: n = 10 and >19 y: n = 6; GSD type Ia: n = 12, Ib: n = 2, III: n = 1, and IX-γ: n = 1). A lean mass (LM) index evaluation showed no LM deficits in relation to corresponding reference populations. Relative skeletal muscle index values were decreased in 2 of 7 adult patients (type Ib: n = 1 and IX-α: n = 1). UCCS (g/d) correlated positively with the FM index (rs = 0.7; P ≤ 0.01). In contrast, relative UCCS intake (g/kg body weight) was negatively associated with LM/kg (rs = -0.8; P ≤ 0.01). CONCLUSIONS: These findings suggest a high frequency of elevated FM in patients with hepatic GSDs. We also suggest that treatment with UCCS is associated with excess weight in these patients. Additionally, the treatment strategy can impair protein intake, and lead to a decrease in LM.

Two different presentations of de novo variants of CSNK2B: two case reports.

BACKGROUND: Poirier-Bienvenu neurodevelopmental syndrome is a neurologic disorder caused by mutations in the CSNK2B gene. It is mostly characterized by early-onset seizures, hypotonia, and mild dysmorphic features. Craniodigital syndrome is a recently described disorder also related to CSNK2B, with a single report in the literature. OBJECTIVE: To report two unrelated cases of children harboring CSNK2B variants (NM_001320.6) who presented with distinct diseases. CASE REPORT: Case 1 is a 7-month-old, Caucasian, female patient with chief complaints of severe hypotonia and drug-refractory myoclonic epilepsy, with a likely pathogenic de novo variant c.494A>G (p.His165Arg). Case 2 is a 5-year-old male, Latino patient with craniodigital intellectual disability syndrome subjacent to a de novo, likely pathogenic variant c.94G>T (p.Asp32Tyr). His dysmorphic features included facial dysmorphisms, supernumerary nipples, and left-hand postaxial polydactyly. CONCLUSION: This report suggest that the CSNK2B gene may be involved in the physiopathology of neurodevelopmental disorders and variable dysmorphic features.

Oropharyngeal Dysphagia in Mucopolysaccharidoses: Evidence from Videofluoroscopic Swallowing Study

Abstract Mucopolysaccharidoses (MPS) are rare inborn errors of metabolism, leading to the accumulation of glycosaminoglycans (GAG) in distinct tissues. We investigated oropharyngeal dysphagia using the videofluoroscopic swallowing study (VFSS) in patients with different MPS types. Since there is a lack of studies systematically evaluating this disorder in this population, the use of a standard technique should contribute to better evaluate individuals with MPS. A cross-sectional and observational study enrolling patients followed by an outpatient service for lysosomal diseases at the Genetics Service of the Hospital de Clínicas de Porto Alegre (SGM/HCPA) was conducted. Patients underwent semi-structured interviews, clinical evaluation and VFSS. Nineteen patients were evaluated, including patients with MPS types I (16%), II (42%), IIIb (10%) and IVa (32%). Nearly all patients (95%) presented with oropharyngeal dysphagia in the VFSS. The most frequent findings were impaired chewing during oral phase (94%) and reduced laryngeal elevation in the pharyngeal phase (72%). Oropharyngeal dysphagia constituted a prevalent symptom in the studied cohort regardless of MPS type. Our data reinforces the notion that this disorder should be objectively assessed since it can significantly compromise the nutrition and the hydration of these patients as well as lead to tracheobronchial aspiration, thus resulting in aspiration pneumonia and even death eventually.

Magnetic resonance imaging findings of the posterior fossa in 47 patients with mucopolysaccharidoses: A cross-sectional analysis.

BACKGROUND: Mucopolysaccharidoses (MPS) is a group of hereditary multisystemic lysosomal disorders. Most neuroimaging studies in MPS have focused on the supratentorial compartment and craniocervical junction abnormalities, and data regarding posterior fossa findings are scarce in the literature. Thus, our purpose is to describe posterior fossa findings on magnetic resonance imaging (MRI) of MPS patients. METHODS: We reviewed routine MRI scans of MPS patients being followed up at our institution (types I, II, III, IV, and VI), focusing on posterior fossa structures. RESULTS: Forty-seven MPS patients were included. MRI-visible perivascular spaces were commonly found in the midbrain and adjacent to the dentate nuclei (85% and 55% of patients, respectively). White-matter lesion was not identified in most cases. Its most frequent localizations were in the pons and cerebellum (34% and 30% of patients, respectively). Enlargement of cerebrospinal fluid (CSF) spaces in the posterior fossa was present in 55% of individuals and was more frequent in neuronopathic patients (73% vs 40%; P = .02). Cerebellar volume was classified as normal, apparent macrocerebellum, atrophic, and hypoplastic in 38%, 38%, 21%, and 3% of patients, respectively. A depression of the posterior fossa floor in the midline sagittal plane was found in 22 patients (47%), which was statistical significantly associated with enlargement of CSF spaces (P = .02) and with apparent macrocerebellum (P = .03). CONCLUSION: The present study compiled the main posterior fossa findings in MPS patients. Classically described in the supratentorial compartment, MRI-visible perivascular spaces, white matter lesions, and enlarged perivascular spaces were also found in the posterior fossa. However, atrophy, which commonly affects cerebral hemispheres, was not the most frequent cerebellar morphology found in our study. Moreover, potential findings for future research were described.

Revealing the clinical phenotype of atypical neuronal ceroid lipofuscinosis type 2 disease: Insights from the largest cohort in the world.

AIM: Neuronal ceroid lipofuscinosis type 2 (CLN2) disease is an autosomal recessive inherited neurodegenerative lysosomal storage disorder caused by deficient tripeptidyl peptidase 1 (TPP1) enzyme, leading to progressive deterioration of neurological functions commonly occurring in children aged 2-4 years and culminating in early death. Atypical cases associated with earlier or later symptom onset, or even protracted course, have already been reported. Such variable manifestations may constitute an additional challenge to early diagnosis and initiation of appropriate treatment. The present work aimed to analyse clinical data from a cohort of Latin American CLN2 patients with atypical phenotypes. METHODS: Experts in inborn errors of metabolism from Latin America selected patients from their centres who were deemed by the clinicians to have atypical forms of CLN2, according to the current literature on this topic and their practical experience. Clinical and genetic data from the medical records were retrospectively revised. All cases were presented and analysed by these experts at an Advisory Board Meeting in São Paulo, Brazil, in October 2018. RESULTS: Seizures, language abnormalities and behavioural disorders were found as the first manifestations, appearing at the median age of 6 years, an older age than classically described for the late infantile form. Three novel mutations were also identified. CONCLUSION: Our findings reinforce the inclusion of CLN2 in the differential diagnosis of children presenting with seizures, behavioural disorders and language abnormalities. Early diagnosis will allow early initiation of specific therapy.

Oral, dental, and craniofacial features in chronic acid sphingomyelinase deficiency.

The aim of this study was to evaluate the oral, dental, and craniofacial features of individuals affected by the chronic forms of acid sphingomyelinase deficiency (ASMD). This study comprised a sample of adult and pediatric patients (n = 8) with chronic ASMD. The individuals underwent oral examinations to evaluate the occurrence of caries, as well as full-mouth periodontal examinations, to assess the occurrence and severity of periodontal diseases. Panoramic and profile radiographs were obtained to analyze dental conditions and craniofacial parameters. Participants also answered questionnaires to identify systemic impairment, parafunctional habits, and bruxism. Dental anomalies of size, shape, and number were found, with agenesis and microdontia being the predominant findings. The average of caries experience was 11.75 (±8.1). Only one patient had periodontal health and all adult individuals had periodontitis at different stages and degrees. Bruxism was found in 87.5% of the sample. The convex profile and maxillary and mandibular retrusion were the most relevant findings in the cephalometric analysis. It is concluded that individuals with chronic ASMD, in addition to several systemic manifestations, present significant modifications in their oral health, from a greater occurrence of dental anomalies, caries, periodontal disease, in addition to skeletal changes.

Progressive eye pathology in mucopolysaccharidosis type I mice and effects of enzyme replacement therapy.

BACKGROUND: Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder caused by α-L-iduronidase deficiency, resulting in accumulation of glycosaminoglycans (GAG). Ophthalmological manifestations are common in MPS I patients and often lead to visual impairment. Accumulation of GAG in corneal or retinal tissues reduces vision causing corneal opacity and neurosensory complications. One available treatment for MPS I patients is enzyme replacement therapy (ERT), but the results of such treatment on eye disease are still debatable. Therefore, we aimed to determine the progression of ocular manifestations as well as the effectiveness of intravenous ERT in MPS I. METHODS: Corneal and retinal analyses were perform in eyes from 2- to 8-month normal and MPS I mice. Some MPS I mice received ERT (1.2 mg/kg of laronidase) every 2 weeks from 6 to 8 months and histological findings were compared with controls. Additionally, cornea from two MPS I patients under ERT were evaluated. RESULTS: Mouse corneal tissues had GAG accumulation early in life. In the retina, we found a progressive loss of photoreceptor cells, starting at 6 months. ERT did not improve or stabilize the histological abnormalities. MPS I patients, despite being on ERT for over a decade, presented GAG accumulation in the cornea, corneal thickening, visual loss and needed corneal transplantation. CONCLUSION: We provide data on the time course of ocular alteration in MPS I mice. Our results also suggest that ERT is not effective in treating the progressive ocular manifestations in MPS I mice and fails to prevent corneal abnormalities in patients.

Perthes-Like Disease Masquerading Non-Classical MPS

Abstract Mucopolysaccharidoses (MPS) are inborn errors of metabolism caused by deficient lysosomal enzymes, leading to organomegaly, hip osteonecrosis, coarse facial features, bone deformities, joint stiffness, cardiac and pulmonary symptoms (MPS VI) or hypermobility (MPS IVA). Some patients may present with non-classical forms of the disease in which osteoarticular abnormalities are the initial symptoms of non-classical forms. As orthopedists and surgeons are the specialists most frequently consulted before the diagnosis, it is critical that MPS may be considered as a differential diagnosis for patients with bone dysplasia. Experts in Latin America reviewed medical records focusing on disease onset, first symptoms and the follow-up clinical and surgical outcomes of non-classical MPS VI and IVA patients. All patients displayed orthopedic issues, which worsened over time, followed by cardiac and ophthalmological abnormalities. Our findings enlighten the necessity of including non-classical MPS as possible diagnosis for patients who report osteoarticular abnormalities in absence of inflammation.

Sensitivity, advantages, limitations, and clinical utility of targeted next-generation sequencing panels for the diagnosis of selected lysosomal storage disorders.

Lysosomal storage disorders (LSDs) constitute a heterogeneous group of approximately 50 genetic disorders. LSDs diagnosis is challenging due to variability in phenotype penetrance, similar clinical manifestations, and a high allelic heterogeneity. A powerful tool for the diagnosis of the disease could reduce the "diagnostic odyssey" for affected families, leading to an appropriate genetic counseling and a better outcome for current therapies, since enzyme replacement therapies have been approved in Brazil for Gaucher, Fabry, and Pompe diseases, and are under development for Niemann-Pick Type B. However, application of next-generation sequencing (NGS) technology in the clinical diagnostic setting requires a previous validation phase. Here, we assessed the application of this technology as a fast, accurate, and cost-effective method to determine genetic diagnosis in selected LSDs. We have designed two panels for testing simultaneously 11 genes known to harbor casual mutations of LSDs. A cohort of 58 patients was used to validate those two panels, and the clinical utility of these gene panels was tested in four novel cases. We report the assessment of a NGS approach as a new tool in the diagnosis of LSDs in our service.

Inborn Errors of Metabolism with Hypoglycemia: Glycogen Storage Diseases and Inherited Disorders of Gluconeogenesis.

Although hyperinsulinism is the predominant inherited cause of hypoglycemia in the newborn period, inborn errors of metabolism are the primary etiologies after 1 month of age. Disorders of carbohydrate metabolism often present with hypoglycemia when fasting occurs. The presentation, diagnosis, and management of the hepatic glycogen storage diseases and disorders of gluconeogenesis are reviewed.

Long-term outcomes of systemic therapies for Hurler syndrome: an international multicenter comparison.

PURPOSE: Early treatment is critical for mucopolysaccharidosis type I (MPS I), justifying its incorporation into newborn screening. Enzyme replacement therapy (ERT) treats MPS I, yet presumptions that ERT cannot penetrate the blood-brain barrier (BBB) support recommendations that hematopoietic cell transplantation (HCT) treat the severe, neurodegenerative form (Hurler syndrome). Ethics precludes randomized comparison of ERT with HCT, but insight into this comparison is presented with an international cohort of patients with Hurler syndrome who received long-term ERT from a young age. METHODS: Long-term survival and neurologic outcomes were compared among three groups of patients with Hurler syndrome: 18 treated with ERT monotherapy (ERT group), 54 who underwent HCT (HCT group), and 23 who received no therapy (Untreated). All were followed starting before age 5 years. A sensitivity analysis restricted age of treatment below 3 years. RESULTS: Survival was worse when comparing ERT versus HCT, and Untreated versus ERT. The cumulative incidences of hydrocephalus and cervical spinal cord compression were greater in ERT versus HCT. Findings persisted in the sensitivity analysis. CONCLUSION: As newborn screening widens treatment opportunity for Hurler syndrome, this examination of early treatment quantifies some ERT benefit, supports presumptions about BBB impenetrability, and aligns with current guidelines to treat with HCT.

Diagnosis and Management of Classica Homocystinuria in Brazil: A Summary of 72 Late-Diagnosed Patients

Abstract This study described a broad clinical characterization of classical homocystinuria (HCU) in Brazil. This was a cross-sectional, observational study including clinical and biochemical data from 72 patients (60 families) from Brazil (South, n = 13; Southeast, n = 37; Northeast, n = 8; North, n = 1; and Midwest, n = 1). Parental consanguinity was reported in 42% of families. Ocular manifestations were the earliest detected symptom (53% of cases), the main reason for diagnostic suspicion (63% of cases), and the most prevalent manifestation at diagnosis (67% of cases). Pyridoxine responsiveness was observed in 14% of patients. Only 22% of nonresponsive patients on treatment had total homocysteine levels <100 mmol/L. Most commonly used treatment strategies were pyridoxine (93% of patients), folic acid (90%), betaine (74%), vitamin B12 (27%), and low-methionine diet + metabolic formula (17%). Most patients diagnosed with HCU in Brazil are late diagnosed, express a severe phenotype, and poor metabolic control. Milder forms of HCU are likely underrepresented due to underdiagnosis.

Correction to: Correlation of CSF flow using phase-contrast MRI with ventriculomegaly and CSF opening pressure in mucopolysaccharidoses.

After publication of the article [1], it has been brought to our attention that the full funding acknowledgement is missing from the original article.

Abnormal polyamine metabolism is unique to the neuropathic forms of MPS: potential for biomarker development and insight into pathogenesis.

The mucopolysaccharidoses (MPS) are rare genetic disorders marked by severe somatic and neurological symptoms. Development of treatments for the neurological manifestations of MPS has been hindered by the lack of objective measures of central nervous system disease burden. Identification of biomarkers for central nervous system disease in MPS patients would facilitate the evaluation of new agents in clinical trials. High throughput metabolite screening of cerebrospinal fluid (CSF) samples from a canine model of MPS I revealed a marked elevation of the polyamine, spermine, in affected animals, and gene therapy studies demonstrated that reduction of CSF spermine reflects correction of brain lesions in these animals. In humans, CSF spermine was elevated in neuropathic subtypes of MPS (MPS I, II, IIIA, IIIB), but not in subtypes in which cognitive function is preserved (MPS IVA, VI). In MPS I patients, elevated CSF spermine was restricted to patients with genotypes associated with CNS disease and was reduced following hematopoietic stem cell transplantation, which is the only therapy currently capable of improving cognitive outcomes. Additional studies in cultured neurons from MPS I mice showed that elevated spermine was essential for the abnormal neurite overgrowth exhibited by MPS neurons. These findings offer new insights into the pathogenesis of CNS disease in MPS patients, and support the use of spermine as a new biomarker to facilitate the development of next generation therapeutics for MPS.

Correlation of CSF flow using phase-contrast MRI with ventriculomegaly and CSF opening pressure in mucopolysaccharidoses.

BACKGROUND: Very little is known about the incidence and prevalence of hydrocephalus in patients with mucopolysaccharidoses (MPS). The biggest challenge is to distinguish communicating hydrocephalus from ventricular dilatation secondary to brain atrophy, because both conditions share common clinical and neuroradiological features. The main purpose of this study is to assess the relationship between ventriculomegaly, brain and cerebrospinal fluid (CSF) volumes, aqueductal and cervical CSF flows, and CSF opening pressure in MPS patients, and to provide potential biomarkers for abnormal CSF circulation. METHODS: Forty-three MPS patients (12 MPS I, 15 MPS II, 5 MPS III, 9 MPS IV A and 2 MPS VI) performed clinical and developmental tests, and T1, T2, FLAIR and phase-contrast magnetic resonance imaging (MRI) followed by a lumbar puncture with the CSF opening pressure assessment. For the analysis of MRI variables, we measured the brain and CSF volumes, white matter (WM) lesion load, Evans' index, third ventricle width, callosal angle, dilated perivascular spaces (PVS), craniocervical junction stenosis, aqueductal and cervical CSF stroke volumes, and CSF glycosaminoglycans concentration. RESULTS: All the scores used to assess the supratentorial ventricles enlargement and the ventricular CSF volume presented a moderate correlation with the aqueductal CSF stroke volume (ACSV). The CSF opening pressure did not correlate either with the three measures of ventriculomegaly, or the ventricular CSF volume, or with the ACSV. Dilated PVS showed a significant association with the ventriculomegaly, ventricular CSF volume and elevated ACSV. CONCLUSIONS: In MPS patients ventriculomegaly is associated with a severe phenotype, increased cognitive decline, WM lesion severity and enlarged PVS. The authors have shown that there are associations between CSF flow measurements and measurements related to CSF volumetrics. There was also an association of volumetric measurements with the degree of dilated PVS.

Investigation of newborns with abnormal results in a newborn screening program for four lysosomal storage diseases in Brazil.

Lysosomal storage diseases (LSDs) are genetic disorders, clinically heterogeneous, mainly caused by defects in genes encoding lysosomal enzymes that degrade macromolecules. Several LSDs already have specific therapies that may improve clinical outcomes, especially if introduced early in life. With this aim, screening methods have been established and newborn screening (NBS) for some LSDs has been developed. Such programs should include additional procedures for the confirmation (or not) of the cases that had an abnormal result in the initial screening. We present here the methods and results of the additional investigation performed in four babies with positive initial screening results in a program of NBS for LSDs performed by a private laboratory in over 10,000 newborns in Brazil. The suspicion in these cases was of Mucopolysaccharidosis I - MPS I (in two babies), Pompe disease and Gaucher disease (one baby each). One case of pseudodeficiency for MPS I, 1 carrier for MPS I, 1 case of pseudodeficiency for Pompe disease and 1 carrier for Gaucher disease were identified. This report illustrates the challenges that may be encountered by NBS programs for LSDs, and the need of a comprehensive protocol for the rapid and precise investigation of the babies who have an abnormal screening result.

Hydrocephalus and mucopolysaccharidoses: what do we know and what do we not know?

INTRODUCTION: The precise incidence of hydrocephalus in patients with mucopolysaccharidoses (MPS) is hard to determine, because the condition lacks a formal, consensus-based definition. The diagnosis of hydrocephalus depends on symptom profile, presence of neuroimaging features, and the outcome of diagnostic tests. Although numerous techniques are used to identify MPS patients who are most likely to have hydrocephalus and respond to treatment, no definitive method exists to prove diagnosis. PURPOSE: The authors propose an algorithm to aid in the diagnosis and management of hydrocephalus in MPS patients. CONCLUSIONS: The theory of venous hypertension associated with the morphological changes in the skull base and craniocervical junction indicate the need for future neuroimaging studies including cerebrospinal fluid (CSF) and venous flow measurements to monitor hydrocephalus progression and select therapeutic interventions in MPS patients. Preoperative planning should also be based on the increased risk of intraoperative and postoperative hemorrhagic complications.

Correction: Biotinidase deficiency: Genotype-biochemical phenotype association in Brazilian patients.

[This corrects the article DOI: 10.1371/journal.pone.0177503.].

Nutritional Status and Body Composition in Patients With Hepatic Glycogen Storage Diseases Treated With Uncooked Cornstarch—A Controlled Study

Abstract Hepatic glycogen storage diseases (GSDs) are genetic diseases associated with fasting hypoglycemia. Periodic intake of uncooked cornstarch is one of the treatment strategies available for those disorders. For reasons that are still not clear, patients with hepatic GSDs may be overweight. Aims: To assess nutritional status and body composition in patients with hepatic GSDs receiving uncooked cornstarch. Methods: The sample included 25 patients with hepatic GSD (type Ia = 14; Ib = 6; III = 3; IX? = 1; IX? = 1), with a median age of 11.0 years (interquartile range [IQR] = 9.0-17.5), matched by age and gender with 25 healthy controls (median age = 12.0 years, IQR = 10.0-17.5). Clinical, biochemical, and treatment-related variables were obtained from medical records. Nutritional status and body composition were prospectively evaluated by bioelectrical impedance. Results: Patients and controls did not differ with regard to age and gender. Height was significantly reduced in patients (median = 1.43 m, IQR = 1.25-1.54) in comparison to controls (median = 1.54 m, IQR = 1.42-1.61; P = .04). Body mass index for age z-score and fat mass percentage were higher in patients (median = 1.84, IQR = 0.55-3.06; and 27.5%, IQR = 22.6-32.0, respectively) than in controls (median = 0.86, IQR = ?0.55 to 1.82; P = .04 and 21.1%, IQR = 13.0-28.3; P = .01, respectively). When patients were stratified by type, those with GSD Ia had significantly higher adiposity (median fat mass = 28.7%, IQR = 25.3-32.9) than those with GSD III and GSD IX?/? (median fat mass = 20.9%, IQR = 14.9-22.6; P = .02). Conclusions: Our findings suggest that patients with hepatic GSD on treatment with cornstarch, especially those with GSD Ia, exhibit abnormalities in nutritional status and body composition, such as short stature and a trend toward overweight and obesity.

Oxidative and nitrative stress and pro-inflammatory cytokines in Mucopolysaccharidosis type II patients: effect of long-term enzyme replacement therapy and relation with glycosaminoglycan accumulation.

Mucopolysaccharidosis type II (MPS II) is a lysosomal storage disease caused by a deficient activity of iduronate-2-sulfatase, leading to abnormal accumulation of glycosaminoglycans (GAG). The main treatment for MPS II is enzyme replacement therapy (ERT). Previous studies described potential benefits of six months of ERT against oxidative stress in patients. Thus, the aim of this study was to investigate oxidative, nitrative and inflammatory biomarkers in MPS II patients submitted to long term ERT. It were analyzed urine and blood samples from patients on ERT (mean time: 5.2years) and healthy controls. Patients presented increased levels of lipid peroxidation, assessed by urinary 15-F2t-isoprostane and plasmatic thiobarbituric acid-reactive substances. Concerning to protein damage, urinary di-tyrosine (di-Tyr) was increased in patients; however, sulfhydryl and carbonyl groups in plasma were not altered. It were also verified increased levels of urinary nitrate+nitrite and plasmatic nitric oxide (NO) in MPS II patients. Pro-inflammatory cytokines IL-1ß and TNF-α were increased in treated patients. GAG levels were correlated to di-Tyr and nitrate+nitrite. Furthermore, IL-1ß was positively correlated with TNF-α and NO. Contrastingly, we did not observed alterations in erythrocyte superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase activities, in reduced glutathione content and in the plasmatic antioxidant capacity. Although some parameters were still altered in MPS II patients, these results may suggest a protective role of long-term ERT against oxidative stress, especially upon oxidative damage to protein and enzymatic and non-enzymatic defenses. Moreover, the redox imbalance observed in treated patients seems to be GAG- and pro-inflammatory cytokine-related.