We present the case of a young adult who had lethargy and significant weight loss for the three weeks before his death. The history of the present illness suggested a prodrome of several weeks, with progressive weakness indicating an advancing metabolic decompensation. To our knowledge, this is the first study performed on human brain tissue with type 1 diabetes (T1D) and likely diabetic ketoacidosis (DKA) before treatment. We studied neuroinflammatory markers in an insulin-deficient state without treatment compared with those found in a treated patient with T1D/DKA of similar age and race who died shortly after treatment. The frontal cortex and hippocampus were stained for tight junction proteins, RAGE, NLRP3, and HMGB1. Other markers that can disrupt the blood-brain barrier, such as IL-17, IL-6, IL-1β, GFAP, and IL-10 were also tested. This case study reveals that neuroinflammatory markers are expressed in the DKA brain at a lower level before treatment than those found to be expressed in the brain after treatment. These findings suggest that in DKA, dehydration minimizes inflammation which could be exacerbated with fluids promoting neuroinflammation and cognitive deficits. These findings require further studies and could identify therapeutic targets to reduce the progression of neuroinflammation and brain edema.

Question What is the analytical and clinical validity of a mass spectrometric method evaluating very long-chain fatty acyl-lysophosphatidylcholine species for the detection of X-linked adrenoleukodystrophy among newborns in North Carolina? Findings In this newborn diagnostic screening study of 52& x202f;301 dried blood spot specimens, 3 male infants were confirmed to have X-linked adrenoleukodystrophy, 3 female infants were identified as heterozygous for X-linked adrenoleukodystrophy (carriers), 1 female infant had a peroxisome biogenesis disorder, 1 female infant had Aicardi-Goutieres syndrome, 1 male infant received an indeterminate diagnosis, and 3 female infants had false-positive results. Meaning The newborn screening results suggested that detecting increased concentrations of very long-chain fatty acyl-lysophosphatidylcholine species in dried blood spots is an effective method for identifying infants with X-linked adrenoleukodystrophy. This diagnostic study evaluates the performance of a single-tier screening assay for X-linked adrenoleukodystrophy among newborns in North Carolina. Importance X-linked adrenoleukodystrophy (X-ALD) is a peroxisomal genetic disorder in which an accumulation of very long-chain fatty acids leads to inflammatory demyelination in the central nervous system and to adrenal cortex atrophy. In 2016, X-ALD was added to the US Recommended Uniform Screening Panel. Objective To evaluate the performance of a single-tier newborn screening assay for X-ALD in North Carolina. Design, Setting, and Participants This diagnostic screening study was of all newborn dried blood spot specimens received in the North Carolina State Laboratory of Public Health between January 2 and June 1, 2018, excluding specimens of insufficient quantity or quality. A total of 52& x202f;301 specimens were screened for X-ALD using negative ionization high-performance liquid chromatography tandem mass spectrometry to measure C24:0- and C26:0-lysophosphatidylcholine concentrations. Sanger sequencing of the adenosine triphosphate-binding cassette subfamily D member 1 (ABCD1) gene was performed on screen-positive specimens. Exposures A medical and family history, newborn physical examination, sequencing of ABCD1 on dried blood spot samples, and plasma analysis of very long-chain fatty acids were obtained for all infants with screen-positive results. Main Outcomes and Measures The prevalence of X-ALD in North Carolina and the positive predictive value and false-positive rate for the first-tier assay were determined. Results Of 52& x202f;301 infants tested (47.8% female, 50.6% male, and 1.7% other or unknown sex), 12 received screen-positive results. Of these 12 infants, 8 were confirmed with a genetic disorder: 3 male infants with X-ALD, 3 X-ALD-heterozygous female infants, 1 female infant with a peroxisome biogenesis disorder, and 1 female infant with Aicardi-Goutieres syndrome. Four infants were initially classified as having false-positives results, including 3 female infants who were deemed unaffected and 1 male infant with indeterminate results on confirmatory testing. The positive predictive value for X-ALD or other genetic disorders for the first-tier assay was 67%, with a false-positive rate of 0.0057%. Conclusions and Relevance This newborn screening pilot study reported results on 2 lysophosphatidylcholine analytes, identifying 3 male infants with X-ALD, 3 X-ALD-heterozygous female infants, and 3 infants with other disorders associated with increased very long-chain fatty acids. These results showed successful implementation in a public health program with minimal risk to the population. The findings will support other state laboratories planning to implement newborn screening for X-ALD and related disorders.

Hypophosphatasia (HPP) is a rare, inherited metabolic bone disorder characterized by low serum alkaline phosphatase activity and impaired bone mineralization. Clinical manifestations and severity of symptoms vary widely in HPP, ranging from in utero death to isolated dental manifestations in adults. Treatment with enzyme replacement therapy has been reported to improve outcomes in perinatal, infantile, and childhood forms of HPP. Here, we present a case of a boy with poor linear growth, mild limb bowing, and radiographic rickets who was diagnosed with HPP before 6 months of age. Treatment with enzyme replacement therapy was initiated at 7 months of age, after which significant improvements in radiographic findings and linear growth were demonstrated. This case highlights several important challenges in the diagnosis, classification, and management of HPP.

Deficiency of the short stature homeobox-containing (SHOX) gene is a frequent cause of short stature in children (2–15%). Here, we report 7 siblings with SHOX deficiency due to a point mutation in the SHOX gene. Index case was a 3-year-old male who presented for evaluation of short stature. His past medical history and birth history were unremarkable. Family history was notable for multiple individuals with short stature. Physical exam revealed short stature, with height standard deviation score (SDS) of −2.98, as well as arm span 3 cm less than his height. His laboratory workup was noncontributory for common etiologies of short stature. Due to significant familial short stature and shortened arm span, SHOX gene analysis was performed and revealed patient is heterozygous for a novel SHOX gene mutation at nucleotide position c.582. This mutation is predicted to cause termination of the SHOX protein at codon 194, effectively causing haploinsufficiency. Six out of nine other siblings were later found to also be heterozygous for the same mutation. Growth hormone was initiated in all seven siblings upon diagnosis and they have demonstrated improved height SDS.

Embryonic sexual differentiation and formation of the genitalia is complex and an interruption in this process results in a variety of abnormalities that can lead to primary amenorrhea. Understanding this process is critical to arrive at the correct diagnosis. We present a case of primary amenorrhea due to the rare clinical syndrome of Testicular Regression Syndrome. Patient presented to us at age 16 with delayed puberty and primary amenorrhea. She was normal in stature. There were no dysmorphic features. She had minimal breast development and sparse pubic hair. No clitoromegaly or labioscrotal fusion was noted. She had elevated gonadotropins, (FSH 79.3mIU/mL, LH 24.9mIU/mL) and low estradiol level (2.3 pg/mL). The remaining endocrine work up was normal. She subsequently had a pelvic ultrasound and well as an Abdominal/Pelvic MRI which revealed no uterus, nor gonads. Two normally positioned kidneys were noted. Her karyotype revealed 46 XY without mosaicism. C-banding indicated the patient's Y chromosome was grossly normal and FISH studies confirmed the presence of the SRY gene in the distal short arm of the Y chromosome. Further evaluation of her Y-chromosome with molecular genetic sequencing revealed no abnormalities of the SRY segment. Laparoscopy with goandectomy was planned due to the possibility of dysgenetic gonadal material that was not able to be visualized on MRI. Findings at laparoscopy revealed only a thick fold of peritoneum across the pelvis without a uterus or cervix. However, fallopian tubes and rudimentary utero-ovarian ligaments versus streak gonads were present. The fallopian tubes and this adjacent tissue were excised and send for pathologic examination. Pathologic examination revealed fallopian tubes and mesonephric duct remnants, though no ovarian or testicular tissue. She was begun on a graduated estrogen replacement regimen to simulate puberty. This patient presents phenotypically female with primary amenorrhea and an absence of internal genitalia. Gonadal dysgenesis is possible but the absence of mullerian structures is perplexing as this implies anti-mullerian hormone from functioning embryonic testis was present at some point. The absence of masculinization, however, indicates that the testes became non-functional before differentiation of the external genitalia occurs. This interesting constellation of findings: partial mullerian development, a seemingly normal XY karyotype, feminized external genitalia, and no gonadal tissue suggests that testes were present for a short time during embryonic development and then disappeared. Understanding the complex embryonic sequence of events that is responsible for sex determination allows one to understand this rare clinical presentation.

Background: Central diabetes insipidus (CDI) results from a number of conditions affecting the hypothalamic-neurohypophyseal system to cause vasopressin deficiency. Diagnosis of CDI is challenging, and clinical data and guidelines for management are lacking. We aim to characterize clinical and radiological characteristics of a cohort of pediatric patients with CDI.Methods: A chart review of 35 patients with CDI followed at North Carolina Children's Hospital from 2000 to 2015 was undertaken. The frequencies of specific etiologies of CDI and characteristic magnetic resonance imaging (MRI) findings were determined. The presence of additional hormone deficiencies at diagnosis and later in the disease course was ascertained. Patient characteristics and management strategies were evaluated.Results: The cohort included 14 female and 21 male patients with a median age of 4.7 years (range, less than 1 month to 16 years) at diagnosis. Median duration of follow-up was 5 years (range, 2 months to 16 years). The cause of CDI was intracranial mass in 13 patients (37.2 %), septo-optic dysplasia in 9 patients (25.7 %), holoprosencephaly in 5 patients (14.2 %), Langerhans cell histiocytosis in 3 patients (8.6 %), isolated pituitary hypoplasia in 2 patients (5.7 %), and encephalocele in 1 patient (2.9 %). Patients were symptomatic for a mean of 6.3 months (range, less than 1 month to 36 months) prior to diagnosis of CDI. Growth hormone (GH), thyrotropin (TSH), adrenocorticotropic hormone (ACTH), and gonadotropin deficiencies were present at diagnosis in 34, 23, 23, and 6 % of patients, respectively. GH, TSH, ACTH, and gonadotropin deficiencies were diagnosed during follow-up in 23, 40, 37, and 14 % of patients, respectively. In patients with structural CNS abnormalities, development of additional hormone deficiencies occurred anywhere from 2 months to 13 years after the time of initial presentation.Conclusions: All patients in our cohort had an underlying organic etiology for CDI, with intracranial masses and CNS malformations being most common. Therefore, MRI of the brain is indicated in all pediatric patients with CDI. Other pituitary hormone deficiencies should be investigated at diagnosis as well as during follow-up.

Vitamin D is important in multiple aspects of health and its effects are mediated through the Vitamin D Receptor (VDR). We wanted to test the hypothesis that specific haplotypes of the VDR gene are associated with markers of disease severity, inflammation and bone health in Sickle Cell Disease (SCD). Genotyping was performed on DNA specimens from 1141 study participants in the NIH-funded Silent Infarct Transfusion (SIT) trial. We used the clinical and laboratory data to create separate endothelial dysfunction, vaso-occlusive severity scores and phenotype variables. Seventy-nine Single Nucleotide Polymorphisms (SNP) in the VDR gene and three associated genes—CYP27B1, VD binding protein, retinoid X receptor, were evaluated. The discovery cohort individuals had VDR haplotype information from a prior Genome-Wide Association Study (GWAS). The validation cohort was analyzed for SNPs that were significant in the discovery cohort. The pheno-type data were obtained from the demographic and clinical information of the participants, and were used to create the severity scores, vaso-occlusive score, endothelial dysfunction severity, and overall severity score. Potential gene-gene interactions were analyzed for prediction of disease severity within each severity score. Two SNPs were associated with the overall severity score, 3 SNPs with the endothelial dysfunction severity score and 4 SNPs with the vaso-occlusive severity score. After permutation testing to correct for multiple comparisons, only one of the associations remained significant. SNP rs7965281 was found to be associated with the endothelial dysfunction severity score and remained significant after correcting for multiple comparisons using permutation testing. In the validation cohort, that SNP was again tested for association with each of the severity scores. There was no association with the endothelial or the overall severity score but a trend towards association with the vaso-occlusive severity score (p = 0.02). None of the known functional polymorphisms in the VDR gene were found to have an association with severity in sickle cell disease. Further work analyzing for gene-gene interaction using the same significant SNPs remains to be done in association with inflammatory markers and measure of bone health. Those studies may provide insight on the contribution of VDR polymorphisms to sickle cell disease severity.

Objective: To evaluate the effects of diabetic ketoacidosis (DKA) on neurocognitive functions in children and adolescents presenting with newonset type 1 diabetes.Methods: Newly diagnosed patients were divided into two groups: those with DKA and those without DKA (non-DKA). Following metabolic stabilization, the patients took a mini-mental status exam prior to undergoing a baseline battery of cognitive tests that evaluated visual and verbal cognitive tasks. Follow-up testing was performed 8-12 weeks after diagnosis. Patients completed an IQ test at follow-up.Results: There was no statistical difference between the DKA and non-DKA groups neither in alertness at baseline testing nor in an IQ test at followup. The DKA group had significantly lower baseline scores than the non-DKA group for the visual cognitive tasks of design recognition, design memory and the composite visual memory index (VMI). At follow-up, Design Recognition remained statistically lower in the DKA group, but the design memory and the VMI tasks returned to statistical parity between the two groups. No significant differences were found in verbal cognitive tasks at baseline or follow-up between the two groups. Direct correlations were present for the admission CO2 and the visual cognitive tasks of VMI, design memory and design recognition. Direct correlations were also present for admission pH and VMI, design memory and picture memory.Conclusion: Pediatric patients presenting with newly diagnosed type 1 diabetes and severe but uncomplicated DKA showed a definite trend for lower cognitive functioning when compared to the age-matched patients without DKA.

BACKGROUND:Treatment with recombinant human insulin-like growth factor-I (IGF-I) stimulates linear growth in children with severe IGF-I deficiency (IGFD). AIMS:To evaluate the efficacy and safety of treatment with IGF-I in patients with severe IGFD treated until adult or near-adult height. METHODS:Twenty-one children with severe IGFD were treated until adult or near-adult height under a predominantly open-label design. All patients were naive to IGF-I. Recombinant human IGF-I was administered subcutaneously in doses between 60 and 120 µg/kg twice daily. Nine patients received additional therapy with gonadotropin- releasing hormone (GnRH) analog for a mean period of 2.9 ± 1.8 years. RESULTS:Mean duration of treatment was 10.0 years. Mean height velocity increased from 3.1 cm/year prior to treatment to 7.4 cm/year during the first year of treatment. Height velocities during the subsequent years were lower, but remained above baseline for up to 12 years. Cumulative mean Δ height SD score at (near) adult height was +2. The observed mean gain in height was 13.4 cm more than had been expected without treatment. The adult height achieved by the patients also treated with GnRH analog was not different from those who received IGF-I therapy alone. There were no new safety signals identified in these patients, a subset of those previously reported. CONCLUSION:Long-term therapy with IGF-I improves adult height of patients with severe IGFD. Most patients did not bring their heights into the normal adult range.

Vitamin D-dependent rickets type 1 is an autosomal recessive disorder caused by an inactivating mutation of the 25-hydroxyvitamin-D-1α-hydroxylase (CYP27B1) gene. Clinical presentation is characterized by early onset of severe rickets and can include severe hypotonia. Here, we report a 16-month-old white male who presented with severe muscle weakness, failure to thrive, renal tubular dysfunction, and skeletal deformities, including osteopenia and multiple fractures. At presentation, he had severe hypocalcemia, hypophosphatemia, hypomagnesemia, and elevated alkaline phosphatase and parathyroid hormone levels, although normal 25-hydroxyvitamin D levels. DNA sequencing of the CYP27B1 gene revealed a novel mutation in exon 2 (c286_300de115) and a previously reported mutation in exon 7 (c.1166G>A). Once calcitriol therapy was initiated, the patient showed significant improvement in muscle strength and linear growth. Serum calcium, phosphorous, and alkaline phosphatase returned to normal range. Organic aciduria resolved and aminoaciduria significantly improved 2 months after parathyroid hormone levels normalized.

Phenotypic and clinical features of individuals with ring chromosome 18 [r(18)] vary with the extent of deletion of the short (18p-) or long arm (18q-). Most patients with r(18), therefore, demonstrate a clinical spectrum of both 18p- and 18q- deletions. Short stature, microcephaly, mental and motor retardation, craniofacial dysmorphism and extremity abnormalities are the most commonly reported features in patients with r(18). Abnormalities of chromosome 18, especially 18p- syndrome, are often reported with autoimmune thyroid disease and growth hormone deficiency, but reports of endocrine abnormalities associated with r(18) are rare. Here, we report a case of an African-American female with hyperthyroidism, type 1 diabetes mellitus, vitiligo and IgA deficiency associated with a r(18) chromosome complement. This patient additionally had mild intellectual disability and dysmorphic features. Karyotype analysis showed a de novo ring chromosome 18 (deletion 18q23-18qter and deletion 18p11.3-18pter). Although this unique association of autoimmune polyglandular endocrinopathy with ring chromosome 18 could be coincidental, we speculate that a gene or genes on chromosome 18 might play a role in the autoimmune process.

Abstract Abstract 2122 Vitamin D is important in multiple aspects of health, including the cardiovascular, immune and skeletal systems and its effects are mediated through the vitamin D receptor (VDR). The systems affected by vitamin D are also perturbed by sickle cell disease (SCD). Vitamin D deficiency is common in SCD, but its contribution to disease manifestations is not yet known. In normal populations, vitamin D has been shown to be associated with hypertension and vascular pathology. That association may be particularly relevant to the inflammatory / endothelial damage seen in sickle cell disease. We have used clinical and laboratory data to create separate inflammatory and vaso-occlusive severity scores. Our hypothesis is that specific VDR polymorphisms are associated with disease severity in sickle cell disease. DNA specimens from 1141 study participants in the NIH-funded Silent Infarct Transfusion (SIT) trial were used. In this multi-center international trial, the participants were children ages 4 to 13 years of age with SCD who were screened for the presence of silent cerebral infarction and had demographic and clinical data collected, as well as samples for a biologic repository for a self-renewing source of DNA. An initial 570 samples served as the discovery cohort. The subsequently enrolled 530 individuals formed our validation cohort. We evaluated 79 single nucleotide polymorphisms (SNP) in the VDR gene and three associated genes, CYP27B1, VD binding protein, retinoid X receptor, and tagging SNPs from the African American population from Hapmap. The discovery cohort individuals had VDR haplotype information from a prior genome-wide association study (GWAS), and analysis for additional VDR-related SNPs was performed using a specifically designed Sequenom assay. The validation cohort was analyzed for SNPs that were significant in the discovery cohort. Phenotype data was obtained from the demographic and clinical information of the participants, and was used to create the severity scores. The vaso-occlusive score includes: number of hospitalizations for pain, number of hospitalizations for acute chest, and avascular necrosis. The inflammatory severity score includes: priapism, transient ischemic attacks (TIA), silent cerebral infarct, systolic and diastolic blood pressure, transcranial doppler velocity, white count and baseline hemoglobin. The overall severity score includes all of the inflammatory and vaso-occlusive variables. To derive the scores, the variables were transformed into quartiles. Each individual subject was assigned values of 1, 2, 3, or 4 for each variable with 1 representing lowest severity and 4, the highest. In addition, in concert with prior analyses of the SIT data, the variable for number of hospitalizations for pain was used alone as a severity measure. The severity scores were not normally distributed and were not totally continuous distributions, so the Kruskal-Wallis test was used in association analysis. To look for complex genetic models including potential gene-gene interactions for prediction of disease severity, the Multifactor Dimensionality Reduction (MDR) method was utilized, with repeat analyses performed for each severity score. By univariate analysis, no associations were found between any of the VDR associated SNPs and the 3 severity scores. Using MDR in the discovery cohort, one SNP, rs7965281, was found to be associated with the inflammatory severity score. It remained significant after correcting for multiple comparisons with permutation analysis. In the validation cohort, rs7965281 was tested for association with each of the severity scores. There was no association with the inflammatory or the vaso-occlusive severity score but a trend towards association with the overall severity score (p= 0.08). All the SNPs were tested for association with the variable, number of hospitalizations for pain using regression analysis. Two additional SNPs, rs7855881 and rs34312136 were found to be nominally significant (p=0.01 and p=0.04 respectively). Rs7965281 shows a trend as well with p=0.06. In the literature, rs7965281 is associated with reduced risk for cutaneous melanoma in a large population based study as well as with blood pressure in a British population. Further work in our validation cohort, including MDR analysis for gene-gene interaction using the 3 significant SNPs remains to be done and this may provide further direction in future research. Disclosures: No relevant conflicts of interest to declare.

OBJECTIVE Type 1 diabetes is an autoimmune disease characterized by the destruction of insulin-producing β-cells. NOD mice provide a useful tool for understanding disease pathogenesis and progression. Although much has been learned from studies with NOD mice, increased understanding of human type 1 diabetes can be gained by evaluating the pathogenic potential of human diabetogenic effector cells in vivo. Therefore, our objective in this study was to develop a small-animal model using human effector cells to study type 1 diabetes. RESEARCH DESIGN AND METHODS We adoptively transferred HLA-A2–matched peripheral blood mononuclear cells (PBMCs) from type 1 diabetic patients and nondiabetic control subjects into transgenic NOD-scid/γcnull/HLA-A*0201 (NOD-scid/γcnull/A2) mice. At various times after adoptive transfer, we determined the ability of these mice to support the survival and proliferation of the human lymphoid cells. Human lymphocytes were isolated and assessed from the blood, spleen, pancreatic lymph node and islets of NOD-scid/γcnull/A2 mice after transfer. RESULTS Human T and B cells proliferate and survive for at least 6 weeks and were recovered from the blood, spleen, draining pancreatic lymph node, and most importantly, islets of NOD-scid/γcnull/A2 mice. Lymphocytes from type 1 diabetic patients preferentially infiltrate the islets of NOD-scid/γcnull/A2 mice. In contrast, PBMCs from nondiabetic HLA-A2–matched donors showed significantly less islet infiltration. Moreover, in mice that received PBMCs from type 1 diabetic patients, we identified epitope-specific CD8+ T cells among the islet infiltrates. CONCLUSIONS We show that insulitis is transferred to NOD-scid/γcnull/A2 mice that received HLA-A2–matched PBMCs from type 1 diabetic patients. In addition, many of the infiltrating CD8+ T cells are epitope-specific and produce interferon-γ after in vitro peptide stimulation. This indicates that NOD-scid/γcnull/A2 mice transferred with HLA-A2–matched PBMCs from type 1 diabetic patients may serve as a useful tool for studying epitope-specific T-cell–mediated responses in patients with type 1 diabetes.

Despite decades of different treatment algorithms, the management of congenital adrenal hyperplasia (CAH) remains clinically challenging. This is due to the inherent difficulty of suppressing adrenal androgen production using near physiological dosing of glucocorticoids (GC). As a result, alternating cycles of androgen versus GC excess can occur and may lead to short stature, obesity, virilization, and alterations in puberty. Novel therapeutic alternatives, including new and more physiological means of GC delivery, inhibitors at the level of CRH or ACTH secretion and/or action, as well as "rescue strategies", such as GnRH analogs, anti-androgens, aromatase inhibitors, and estrogen receptor blockers, are available; many of these agents, however, still require active investigation in CAH. Bilateral adrenalectomy is effective but it is also still an experimental approach. Gene therapy and stem cells, to provide functional adrenal cortical tissue, are at preclinical stage but provide exciting avenues for a potential cure for CAH.