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La educación inclusiva es un aspecto que en las instituciones de aprendizaje se pretende adquirir, con las herramientas tecnológicas en evolución contante es necesario aplicar distintos modelos de enseñanza, para este contexto se necesita una adaptación curricular frente a la demande del problema de discapacidad intelectual, en la actualidad aún no se logra emplear estrategias y técnicas que aporten de manera efectiva en dicho proceso dentro de las aulas de clases y que sobre todo abordar el tema de las NEE. En este sentido el objetivo de la investigación es analizar las adaptaciones curriculares para implementar el uso de las Tics en estudiantes con discapacidad intelectual. Este trabajo esta alineado a un estudio descriptivo con enfoque cualitativo, enmarcado en diversos métodos entre ellos; de revisión bibliográfica, análisis y síntesis. Los resultados obtenidos han permitido conocer la importancia de realizar adaptaciones curriculares basadas en las fortalezas y potencialidades (habilidades, destrezas) del alumno, para incrementar sus conocimientos.

Context Maternally inherited STX16 deletions that cause loss of methylation at GNAS exon A/B and thereby reduce Gsα expression are the most frequent cause of autosomal dominant pseudohypoparathyroidism type Ib (AD-PHP1B). Early identification of these disease-causing variants in the children of affected and unaffected female carriers would prompt treatment with calcium and calcitriol once parathyroid hormone (PTH) levels increase, thereby preventing hypocalcemia and associated complications. Objective This study aimed to determine when PTH and calcium abnormalities develop after birth if a STX16 deletion is inherited maternally. Methods Forty-four children of affected (n = 7) or unaffected (n = 7) females with a STX16 deletion were investigated for the presence of these variants. If a deletion was identified, measurement of PTH, calcium, phosphate, and thyrotropin (TSH) was advised. Results The STX16 deletion that causes AD-PHP1B was identified in 25 children. Pretreatment laboratory results were available for 19 of those cases. Elevated PTH levels were detected by 2 years of age, and these were progressively higher if laboratory testing was first performed after establishing the genetic defect later in life. Total serum calcium levels remained within normal limits until about 5 years of age. TSH levels showed no consistent rise over time. Conclusion Establishing whether a STX16 deletion is inherited from a female carrier of a disease-causing variant rapidly establishes the diagnosis of AD-PHP1B. Several years before overt hypocalcemia developed, PTH levels increased, thereby establishing the onset of PTH resistance. Our findings provide diagnostic guidance and when treatment with calcium and calcitriol should be considered in order to prevent hypocalcemia and associated sequelae.

Preferential transmission of a genetic mutation to the next generation, referred to as transmission ratio distortion (TRD), is well established for several dominant disorders, but underlying mechanisms remain undefined. Recently, TRD was reported for patients affected by pseudohypoparathyroidism type Ia or pseudopseudohypoparathyroidism. To determine whether TRD is observed also for autosomal dominant pseudohypoparathyroidism type Ib (AD-PHP1B), we analyzed kindreds with the frequent 3-kb STX16 deletion or other STX16/GNAS mutations. If inherited from a female, these genetic defects lead to loss-of-methylation at exon A/B alone or at all three differentially methylated regions (DMR), resulting in parathyroid hormone (PTH)-resistant hypocalcemia and hyperphosphatemia and possibly resistance to other hormones. In total, we investigated 212 children born to 80 females who are unaffected carriers of a STX16/GNAS mutation (n = 47) or affected by PHP1B (n = 33). Of these offspring, 134 (63.2%) had inherited the genetic defect (p = .00012). TRD was indistinguishable for mothers with a STX16/GNAS mutation on their paternal (unaffected carriers) or maternal allele (affected). The mechanisms favoring transmission of the mutant allele remain undefined but are likely to include abnormalities in oocyte maturation. Search for mutations in available descendants of males revealed marginally significant evidence for TRD (p = .038), but these analyses are less reliable because many more offspring of males than females with a STX16/GNAS mutation were lost to follow-up (31 of 98 versus 6 of 218). This difference in follow-up is probably related to the fact that inheritance of a mutation from a male does not have clinical implications, whereas inheritance from an affected or unaffected female results in PHP1B. Lastly, affected PHP1B females had fewer descendants than unaffected carriers, but it remains unclear whether abnormal oocyte development or impaired actions of reproductive hormones are responsible. Our findings highlight previously not recognized aspects of AD-PHP1B that are likely to have implications for genetic testing and counseling. © 2020 American Society for Bone and Mineral Research (ASBMR).

Pseudohypoparathyroidism type Ib (PHP1B) is characterized by resistance to parathyroid hormone (PTH) leading to hypocalcemia and hyperphosphatemia, and in some cases resistance toward additional hormones. Patients affected by this disorder all share a loss-of-methylation (LOM) at the differentially methylated GNAS exon A/B, which reduces expression of the stimulatory G protein α-subunit (Gsα) from the maternal allele. This leads in the proximal renal tubules, where the paternal GNAS allele does not contribute much to expression of this signaling protein, to little or no Gsα expression thereby causing PTH resistance. We now describe a PHP1B patient with a de novo genomic GNAS duplication of approximately 88 kb, which is associated with LOM restricted to exon A/B alone. Multiplex ligation-dependent probe amplification (MLPA), comparative genomic hybridization (CGH), and whole-genome sequencing (WGS) established that the duplicated DNA fragment extends from GNAS exon AS1 (telomeric breakpoint) to a small region between two imperfect repeats just upstream of LOC105372695 (centromeric breakpoint). Our novel duplication is considerably shorter than previously described duplications/triplications in that portion of chromosome 20q13 and it does not affect methylation at exons AS and XL. Based on these and previous findings, it appears plausible that the identified genomic abnormality disrupts in cis the actions of a transcript that is required for establishing or maintaining exon A/B methylation. Our findings extend the molecular causes of PHP1B and provide additional insights into structural GNAS features that are required for maintaining maternal Gsα expression and for preventing PTH-resistance. © 2020 American Society for Bone and Mineral Research (ASBMR).

The sodium-phosphate cotransporter NPT2a plays a key role in the reabsorption of filtered phosphate in proximal renal tubules, thereby critically contributing to phosphate homeostasis. Inadequate urinary phosphate excretion can lead to severe hyperphosphatemia as in tumoral calcinosis and chronic kidney disease (CKD). Pharmacological inhibition of NPT2a may therefore represent an attractive approach for treating hyperphosphatemic conditions. The NPT2a-selective small-molecule inhibitor PF-06869206 was previously shown to reduce phosphate uptake in human proximal tubular cells in vitro. Here, we investigated the acute and chronic effects of the inhibitor in rodents and report that administration of PF-06869206 was well tolerated and elicited a dose-dependent increase in fractional phosphate excretion. This phosphaturic effect lowered plasma phosphate levels in WT mice and in rats with CKD due to subtotal nephrectomy. PF-06869206 had no effect on Npt2a-null mice, but promoted phosphate excretion and reduced phosphate levels in normophophatemic mice lacking Npt2c and in hyperphosphatemic mice lacking Fgf23 or Galnt3. In CKD rats, once-daily administration of PF-06869206 for 8 weeks induced an unabated acute phosphaturic and hypophosphatemic effect, but had no statistically significant effect on FGF23 or PTH levels. Selective pharmacological inhibition of NPT2a thus holds promise as a therapeutic option for genetic and acquired hyperphosphatemic disorders.

Rodent models are commonly used to evaluate parathyroid hormone (PTH) and PTH-related protein (PTHrP) ligands and analogues for their pharmacologic activities and potential therapeutic utility toward diseases of bone and mineral ion metabolism. Divergence, however, in the amino acid sequences of rodent and human PTH receptors (rat and mouse PTH1Rs are 91% identical to the human PTH1R) can lead to differences in receptor-binding and signaling potencies for such ligands when assessed on rodent vs human PTH1Rs, as shown by cell-based assays in vitro. This introduces an element of uncertainty in the accuracy of rodent models for performing such preclinical evaluations. To overcome this potential uncertainty, we used a homologous recombination-based knockin (KI) approach to generate a mouse (in-host strain C57Bl/6N) in which complementary DNA encoding the human PTH1R replaces a segment (exon 4) of the murine PTH1R gene so that the human and not the mouse PTH1R protein is expressed. Expression is directed by the endogenous mouse promoter and hence occurs in all biologically relevant cells and tissues and at appropriate levels. The resulting homozygous hPTH1R-KI (humanized) mice were healthy over at least 10 generations and showed functional responses to injected PTH analog peptides that are consistent with a fully functional human PTH1R in target bone and kidney cells. The initial evaluation of these mice and their potential utility for predicting behavior of PTH analogues in humans is reported here.

Individuals affected by pseudohypoparathyroidism type 1A (PHP1A) display hyperphosphatemia and hypocalcemia despite elevated PTH levels, as well as features of Albright Hereditary Osteodystrophy (AHO). PHP1A is caused by variants involving the maternal GNAS exons 1-13 encoding the stimulatory G protein alpha-subunit (Gs alpha). MLPA and aCGH analysis led in a male PHP1A patient to identification of a de novo 1284-bp deletion involving GNAS exon 1. This novel variant overlaps with a previously identified 1438-bp deletion in another PHP1A patient (ref. Li et al. (2020) {[}13], patient 2) that extends from the exon 1 promoter into the up-stream intronic region. This latter deletion is associated with reduced methylation at GNAS exon A/B, i.e. the differentially methylated region (DMR) that is demethylated in most pseudohypoparathyroidism type 1B (PHP1B) patients. In contrast, genomic DNA from our patient revealed no evidence for an epigenetic GNAS defect as determined by MS-MLPA and pyrosequencing. These findings thus reduce the region, which, in addition to other nucleotide sequences telomeric of exon A/B, may undergo histone modifications or interacts with transcription factors and possibly as-yet unknown proteins that are required for establishing the maternal methylation imprints at this site. Taken together, nucleotide deletions or changes within an approximately 1300-bp region telomeric of exon A/B could be a cause of PHP1B variants with complete or incomplete loss-of-methylation at the exon A/B DMR. In addition, when investigating patients with suspected PHP1A, MLPA should be considered to search for structural abnormalities within this difficult to analyze genomic region comprising GNAS exon 1.

The PTH/PTHrP receptor (PTHR1) mediates the actions of parathyroid hormone (PTH) and PTH-related peptide (PTHrP) by coupling this G protein-coupled receptor (GPCR) to the alpha-subunit of the heterotrimeric stimulatory G protein (Gsα) and thereby to the formation of cAMP. In growth plates, PTHrP-dependent activation of the cAMP/PKA second messenger pathway prevents the premature differentiation of chondrocytes into hypertrophic cells resulting in delayed growth plate closure. Heterozygous mutations in GNAS, the gene encoding Gsα, lead to a reduction in cAMP levels in growth plate chondrocytes that is sufficient to cause shortening of metacarpals and/or -tarsals, i. e. typical skeletal aspects of Albright’s Hereditary Osteodystrophy (AHO). However, heterozygous mutations in other genes, including those encoding PTHrP, PRKAR1A, PDE4D, and PDE3A, can lead to similar or even more pronounced acceleration of skeletal maturation that is particularly obvious in hands and feet, and reduces final adult height. Genetic mutations other than those resulting in Gsα haploinsufficiency thus reduce intracellular cAMP levels in growth plate chondrocytes to a similar extent and thereby accelerate skeletal maturation.

In 1980, Farfel and colleagues (NEJM, 1980;303:237–42) provided first evidence for two distinct variants of pseudohypoparathyroidism (PHP) that present with hypocalcemia and impaired parathyroid hormone (PTH)‐stimulated urinary cAMP and phosphate excretion, either in the presence or absence of Albright's hereditary osteodystrophy (AHO). An “abnormal allele” and an “unexpressed allele” were considered as underlying defects, predictions that turned out to be correct for both forms of PHP. Patients affected by the first variant (now referred to as PHP1A) were later shown to be carriers of inactivating mutations involving the maternal GNAS exons encoding Gsα. Patients affected by the second variant (now referred to as PHP1B) were shown in the current study to carry a maternal 3‐kb STX16 deletion, the most frequent cause of autosomal dominant PHP1B, which is associated with loss of methylation at GNAS exon A/B that reduces or abolishes maternal Gsα expression. However, the distinct maternal mutations leading to either PHP1A or PHP1B are disease‐causing only because paternal Gsα expression in the proximal renal tubules is silenced, ie, “unexpressed.” Our findings resolve at the molecular level carefully conducted investigations reported some 41 years ago that had provided first clues for the existence of two distinct PHP variants. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Brachydactyly type E, which can be an isolated finding or part of a syndrome in combination with other clinical anomalies, involves metacarpals and metatarsals with or without short phalanges. Herein we report two unrelated Turkish females who presented with brachydactyly type E and vitamin D deficiency in the absence of marked alterations in serum calcium, phosphate, and parathyroid hormone. After excluding disease-causing variants in two candidate genes, PTHLH and PDE4D, we identified different pathogenic variants in TRPS1, the gene mutated in patients with tricho-rhino-phalangeal syndrome (TRPS). In one of the patients, who displayed severe brachydactyly and short stature, we identified a novel heterozygous missense pathogenic variant in exon 6 (c.2783A>G, p.Tyr928Cys), located within the GATA DNA-binding domain. The second patient, who had relatively milder brachydactyly and was of normal height, carried a heterozygous nonsense pathogenic variant in exon 4 (c. 1870C>T, p.Arg624Ter), which has been previously described. Both pathogenic variants segregated in affected family members. The patients additionally showed sparse hair and a bulbous nose, consistent with the clinical features of TRPS. Our findings, in addition to identifying the genetic cause of brachydactyly in two unrelated kindreds, emphasize the role of pathogenic TRPS1 variants in the development of brachydactyly type E and highlight the GATA DNA-binding region of TRPS1 protein with respect to phenotype-genotype correlation.

Context Pseudohypoparathyroidism type Ib (PHP1B) is characterized by hypocalcemia and hyperphosphatemia due to parathyroid hormone resistance in the proximal renal tubules. Maternal pathogenic STX16/GNAS variants leading to maternal epigenetic GNAS changes impair expression of the stimulatory G protein alpha-subunit (Gsα) thereby causing autosomal dominant PHP1B. In contrast, genetic defects responsible for sporadic PHP1B (sporPHP1B) remain mostly unknown. Objective Determine whether PHP1B encountered after in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) causes GNAS remethylation defects similar to those in sporPHP1B. Design Retrospective analysis. Results Nine among 36 sporPHP1B patients investigated since 2000, all with loss of methylation (LOM) at the 3 maternal GNAS differentially methylated regions (DMRs) and gain of methylation at the paternal NESP DMR, had been conceived through IVF or ICSI. Besides abnormal GNAS methylation, IVF/ICSI PHP1B cases revealed no additional imprinting defects. Three of these PHP1B patients have dizygotic twins, and 4 have IVF/ICSI-conceived siblings, all with normal GNAS methylation; 2 unaffected younger siblings were conceived naturally. Conclusion Sporadic and IVF/ICSI-conceived PHP1B patients revealed indistinguishable epigenetic changes at all 4 GNAS DMRs, thus suggesting a similar underlying disease mechanism. Given that remethylation at the 3 maternal DMRs occurs during oogenesis, male factors are unlikely to cause LOM postfertilization. Instead, at least some of the sporPHP1B variants could be caused by a defect or defects in an oocyte-expressed gene that is required for fertility and for re-establishing maternal GNAS methylation imprints. It remains uncertain, however, whether the lack of GNAS remethylation alone and the resulting reduction in Gsα expression is sufficient to impair oocyte maturation.

Many chemicals in commonly used household and industrial products are being released into the environment, yet their toxicity is poorly understood. The synthetic phenolic antioxidant, 4,4′-thiobis(6-t-butyl-m-cresol) (CAS 96-69-5; TBBC) is present in many common products made of rubber and plastic. Yet, this phenolic antioxidant has not been tested for potential toxicity and developmental disruption in amphibians, a sensitive and susceptible class. We investigated whether acute and chronic exposure to TBBC would interfere with thyroid hormone-dependent developmental processes in the frog Silurana tropicalis and thus affect its early life-stage development. We exposed S. tropicalis embryos at the Nieuwkoop-Faber (NF) 9–10 stage to TBBC at nominal concentrations (0, 25, 50, 75, 100, 200 and 400 μg/L) to determine the 96h lethal concentrations and sublethal effects. We conducted a chronic exposure starting at stage NF47-48 to three sublethal TBBC nominal concentrations (0, 0.002, 0.1 and 5 μg/L) for 48–52 days to evaluate effects on growth and metamorphosis. The 96h lethal and effective (malformations) TBBC concentrations (LC50 and EC50) were 70.5 and 76.5 μg/L, respectively. Acute exposure to all TBBC concentrations affected S. tropicalis growth and was lethal at 200 and 400 μg/L. Chronic exposure to sublethal TBBC concentrations reduced body size by 8% at 5 μg/L and body mass by 17% at 0.002 μg/L when metamorphosis was completed. This study demonstrates that TBBC is toxic, induces malformations and inhibits tadpole growth after acute and chronic exposures. These findings call for further investigations on the mode of actions of TBBC and related antioxidants for developmental disruption in amphibians.

GNAS exons 1–13 encode the biallelically expressed alpha-subunit of the stimulatory G protein (Gαs). Additional transcripts derived from this locus use alternative first exons that undergo parent-specific methylation, thus allowing transcription only from the non-modified allele. Pseudohypoparathyroidism type Ia (PHP1A) is characterized by Albright's Hereditary Osteodystrophy (AHO) and resistance to multiple hormones; this disorder is caused by maternal inactivating mutations involving Gαs exons. In contrast, pseudohypoparathyroidism type Ib (PHP1B) is characterized mostly by resistance to PTH and often mild TSH resistance, usually without AHO features. The autosomal dominant variant of PHP1B (AD-PHP1B) is caused by maternal deletions in GNAS or STX16 that reduce Gαs expression through loss-of-methylation at GNAS exon A/B alone or at multiple differentially methylated regions (DMR). Several large maternal deletions involve not only GNAS exons 1–13, but also one or several GNAS DMRs, thus causing PHP1A combined with apparent GNAS epigenetic changes that are indistinguishable from those observed in PHP1B. Some of these deletions include a large CpG island extending from exon A/B to the intron between GNAS exons 1 and 2, but there is no evidence for parent-specific exon 1 methylation. We now describe a family in which the female proband and her daughter presented with hypocalcemia, elevated PTH levels, shortened metacarpals, and obesity, but without obvious neurocognitive abnormalities. A maternally inherited 2015-bp deletion that includes GNAS exon 1was identified thereby establishing the diagnosis of PHP1A. The centromeric deletion breakpoint is located 178 bp upstream of exon 1, yet no methylation changes were observed at exon A/B. This novel deletion therefore refines further the region between exon A/B and exon 1 that is critical for establishing or maintaining normal methylation at GNAS exon A/B. 1.

Pseudohypoparathyroidism type Ib (PHP1B) is characterized primarily by resistance to parathyroid hormone (PTH) and thus hypocalcemia and hyperphosphatemia, in most cases without evidence for Albright hereditary osteodystrophy (AHO). PHP1B is associated with epigenetic changes at one or several differentially-methylated regions (DMRs) within GNAS, which encodes the α-subunit of the stimulatory G protein (Gsα) and splice variants thereof. Heterozygous, maternally inherited STX16 or GNAS deletions leading to isolated loss-of-methylation (LOM) at exon A/B alone or at all maternal DMRs are the cause of autosomal dominant PHP1B (AD-PHP1B). In this study, we analyzed three affected individuals, the female proband and her two sons. All three revealed isolated LOM at GNAS exon A/B, whereas the proband's healthy maternal grandmother and uncle showed normal methylation at this locus. Haplotype analysis was consistent with linkage to the STX16/GNAS region, yet no deletion could be identified. Whole-genome sequencing of one of the patients revealed a large heterozygous inversion (1,882,433 bp). The centromeric breakpoint of the inversion is located 7,225 bp downstream of GNAS exon XL, but its DMR showed no methylation abnormality, raising the possibility that the inversion disrupts a regulatory element required only for establishing or maintaining exon A/B methylation. Because our three patients presented phenotypes consistent with PHP1B, and not with PHP1A, the Gsα promoter is probably unaffected by the inversion. Our findings expand the spectrum of genetic mutations that lead to LOM at exon A/B alone and thus biallelic expression of the transcript derived from this alternative first GNAS exon. © 2017 American Society for Bone and Mineral Research.

Jansen's metaphyseal chondrodysplasia (JMC) is a rare disease of bone and mineral ion physiology that is caused by activating mutations in PTHR1. Ligand-independent signaling by the mutant receptors in cells of bone and kidney results in abnormal skeletal growth, excessive bone turnover, and chronic hypercalcemia and hyperphosphaturia. Clinical features further include short stature, limb deformities, nephrocalcinosis, and progressive losses in kidney function. There is no effective treatment option available for JMC. In previous cell-based assays, we found that certain N-terminally truncated PTH and PTHrP antagonist peptides function as inverse agonists and thus can reduce the high rates of basal cAMP signaling exhibited by the mutant PTHR1s of JMC in vitro. Here we explored whether one such inverse agonist ligand, [Leu11 ,dTrp12 ,Trp23 ,Tyr36 ]-PTHrP(7-36)NH2 (IA), can be effective in vivo and thus ameliorate the skeletal abnormalities that occur in transgenic mice expressing the PTHR1-H223R allele of JMC in osteoblastic cells via the collagen-1α1 promoter (C1HR mice). We observed that after 2 weeks of twice-daily injection and relative to vehicle controls, the IA analog resulted in significant improvements in key skeletal parameters that characterize the C1HR mice, because it reduced the excess trabecular bone mass, bone marrow fibrosis, and levels of bone turnover markers in blood and urine. The overall findings provide proof-of-concept support for the notion that inverse agonist ligands targeted to the mutant PTHR1 variants of JMC can have efficacy in vivo. Further studies of such PTHR1 ligand analogs could help open paths toward the first treatment option for this debilitating skeletal disorder. © 2019 American Society for Bone and Mineral Research.

Objective Autosomal dominant hypocalcemia (ADH) is characterized by hypocalcemia and inappropriately low PTH concentrations. ADH type 1 is caused by activating mutations in the calcium-sensing receptor (CASR), a G-protein-coupled receptor signaling through α11 (Gα11) and αq (Gαq) subunits. Heterozygous activating mutations in GNA11, the gene encoding Gα11, underlie ADH type 2. This study describes disease characteristics in a family with ADH caused by a gain-of-function mutation in GNA11. Design A three-generation family with seven members (3 adults, 4 children) presenting with ADH. Methods Biochemical parameters of calcium metabolism, clinical, genetic and brain imaging findings were analyzed. Results Sanger sequencing revealed a heterozygous GNA11 missense mutation (c.1018G>A, p.V340M) in all seven hypocalcemic subjects, but not in the healthy family members (n=4). The adult patients showed clinical symptoms of hypocalcemia, while the children were asymptomatic. Plasma ionized calcium ranged from 0.95 to 1.14mmol/L, yet plasma PTH was inappropriately low for the degree of hypocalcemia. Serum 25OHD was normal. Despite hypocalcemia 1,25(OH)2D and urinary calcium excretion were inappropriately in the reference range. None of the patients had nephrocalcinosis. Two adults and one child (of the two MRI scanned children) had distinct intracranial calcifications. All affected subjects had short stature (height s.d. scores ranging from −3.4 to −2.3 vs −0.5 in the unaffected children). Conclusions The identified GNA11 mutation results in biochemical abnormalities typical for ADH. Additional features, including short stature and early intracranial calcifications, cosegregated with the mutation. These findings may indicate a wider role for Gα11 signaling besides calcium regulation.

Proximal tubular resistance to parathyroid hormone (PTH) resulting in hypocalcemia and hyperphosphatemia are preeminent abnormalities in pseudohypoparathyroidism type Ib (PHP1B), but resistance toward other hormones as well as variable features of Albright's Hereditary Osteodystrophy (AHO) can occur also. Genomic DNA from PHP1B patients shows epigenetic changes at one or multiple differentially methylated regions (DMRs) within GNAS, the gene encoding Gαs and splice variants thereof. In the autosomal dominant disease variant, these methylation abnormalities are caused by deletions in STX16 or GNAS on the maternal allele. The molecular defect(s) leading to sporadic PHP1B (sporPHP1B) remains in most cases unknown and we therefore analyzed 60 sporPHP1B patients and available family members by microsatellite markers, single nucleotide polymorphisms (SNPs), multiplex ligation-dependent probe amplification (MLPA), and methylation-specific MLPA (MS-MLPA). All investigated cases revealed broad GNAS methylation changes, but no evidence for inheritance of two paternal chromosome 20q alleles. Some patients with partial epigenetic modifications in DNA from peripheral blood cells showed more complete GNAS methylation changes when testing their immortalized lymphoblastoid cells. Analysis of siblings and children of sporPHP1B patients provided no evidence for an abnormal mineral ion regulation and no changes in GNAS methylation. Only one patient revealed, based on MLPA and microsatellite analyses, evidence for an allelic loss, which resulted in the discovery of two adjacent, maternally inherited deletions (37,597 and 1427 bp, respectively) that remove the area between GNAS antisense exons 3 and 5, including exon NESP. Our findings thus emphasize that the region comprising antisense exons 3 and 4 is required for establishing all maternal GNAS methylation imprints. The genetic defect(s) leading in sporPHP1B to epigenetic GNAS changes and thus PTH-resistance remains unknown, but it seems unlikely that this disease variant is caused by heterozygous inherited or de novo mutations involving GNAS.

Context The α subunit of the stimulatory G protein (Gαs) links numerous receptors to adenylyl cyclase. Gαs, encoded by GNAS, is expressed predominantly from the maternal allele in certain tissues. Thus, maternal heterozygous loss-of-function mutations cause hormonal resistance, as in pseudohypoparathyroidism type Ia, whereas somatic gain-of-function mutations cause hormone-independent endocrine stimulation, as in McCune-Albright syndrome. Objective We report two unrelated boys presenting with a new combination of clinical findings that suggest both gain and loss of Gαs function. Design and Setting Clinical features were studied and sequencing of GNAS was performed. Signaling capacities of wild-type and mutant Gαs were determined in the presence of different G protein–coupled receptors (GPCRs) under basal and agonist-stimulated conditions. Results Both unrelated patients presented with unexplained hyponatremia in infancy, followed by severe early onset gonadotrophin-independent precocious puberty and skeletal abnormalities. An identical heterozygous de novo variant (c.1136T>G; p.F376V) was found on the maternal GNAS allele in both patients; this resulted in a clinical phenotype that differed from known Gαs-related diseases and suggested gain of function at the vasopressin 2 receptor (V2R) and lutropin/choriogonadotropin receptor (LHCGR), yet increased serum PTH concentrations indicative of impaired proximal tubular PTH1 receptor (PTH1R) function. In vitro studies demonstrated that Gαs-F376V enhanced ligand-independent signaling at the PTH1R, LHCGR, and V2R and, at the same time, blunted ligand-dependent responses. Structural homology modeling suggested mutation-induced modifications at the C-terminal α5 helix of Gαs that are relevant for interaction with GPCRs and signal transduction. Conclusions The Gαs p.F376V mutation causes a previously unrecognized multisystem disorder.

Context Early-onset obesity, characteristic for disorders affecting the leptin–melanocortin pathway, is also observed in pseudohypoparathyroidism type 1A (PHP1A), a disorder caused by maternal GNAS mutations that disrupt expression or function of the stimulatory G protein α-subunit (Gsα). Mutations and/or epigenetic abnormalities at the same genetic locus are also the cause of pseudohypoparathyroidism type 1B (PHP1B). However, although equivalent biochemical and radiographic findings can be encountered in these related disorders caused by GNAS abnormalities, they are considered distinct clinical entities. Objectives To further emphasize the overlapping features between both disorders, we report the cases of several children, initially brought to medical attention because of unexplained early-onset obesity, in whom PHP1B or PHP1A was eventually diagnosed. Patients and Methods Search for GNAS methylation changes or mutations in cohorts of patients with early-onset obesity. Results Severe obesity had been noted in five infants, with a later diagnosis of PHP1B due to STX16 deletions and/or abnormal GNAS methylation. These findings prompted analysis of 24 unselected obese patients, leading to the discovery of inherited STX16 deletions in 2 individuals. Similarly, impressive early weight gains were noted in five patients, who initially lacked additional Albright hereditary osteodystrophy features but in whom PHP1A due to GNAS mutations involving exons encoding Gsα was diagnosed. Conclusions Obesity during the first year of life can be the first clinical evidence for PHP1B, expanding the spectrum of phenotypic overlap between PHP1A and PHP1B. Importantly, GNAS methylation abnormalities escape detection by targeted or genome-wide sequencing strategies, raising the question of whether epigenetic GNAS analyses should be considered for unexplained obesity.

Short metacarpals and/or metatarsals are typically observed in pseudohypoparathyroidism (PHP) type Ia (PHP1A) or pseudo-PHP (PPHP), disorders caused by inactivating GNAS mutations involving exons encoding the alpha-subunit of the stimulatory G protein (Gsα). Skeletal abnormalities similar to those in PHP1A/PPHP were present in several members of an extended Belgian family without evidence for abnormal calcium and phosphate regulation. Direct nucleotide sequencing of genomic DNA from an affected individual (190/III-1) excluded GNAS mutations. Instead, whole exome analysis revealed a novel heterozygous A>G change at nucleotide -3 upstream of PTHLH exon 3 that encodes the last two amino acids of the prosequence and the mature PTHrP. The same nucleotide change was also found in her affected mother and maternal aunt (190/II-2, 190/II-1), and her affected twin sons (190/IV-1, 190/IV-2), but not in her unaffected daughter (190/IV-3) and sister (190/III-2). Complementary DNA derived from immortalized lymphoblastoid cells from 190/IV-2 (affected) and 190/IV-3 (unaffected) was PCR-amplified using forward primers located either in PTHLH exon 1 (noncoding) or exon 2 (presequence and most of the prosequence), and reverse primers located in the 3'-noncoding regions of exons 3 or 4. Nucleotide sequence analysis of these amplicons revealed for the affected son 190/IV-2, but not for the unaffected daughter 190/IV-3, a heterozygous insertion of genomic nucleotides -2 and -1 causing a frameshift after residue 34 of the pre/prosequence and thus 29 novel residues without homology to PTHrP or any other protein. Our findings extend previous reports indicating that PTHrP haploinsufficiency causes skeletal abnormalities similar to those observed with heterozygous GNAS mutations. © 2018 American Society for Bone and Mineral Research.

Pseudohypoparathyroidism 1A (PHP1A) is a rare, genetic disorder. Most patients with PHP1A have cognitive impairment but this has not been systematically studied. We hypothesized that children with PHP1A would have lower intelligent quotient (IQ) scores than controls. To evaluate cognition and behavior, we prospectively enrolled children with PHP1A, one unaffected sibling (when available) and controls matched on BMI/age/gender/race. Evaluations included cognitive and executive function testing. Parents completed questionnaires on behavior and executive function. We enrolled 16 patients with PHP1A, 8 unaffected siblings, and 15 controls. Results are presented as mean (SD). The PHP1A group had a composite IQ of 85.9 (17.2); 25% had a composite IQ

Background RT-PCR (reverse transcriptase polymerase chain reaction) is often considered the “gold standard” for diagnosis of Zika Virus (ZIKV) infection; however, it has been shown to have low sensitivity. A possible remedy is to study ZIKV-specific IgG (ZsIgG) and IgM (ZsIgM) antibodies. However, the in vitro cross-reactivities of Dengue virus (DENV) and ZIKV-specific antibodies are well known, leading to diagnostic difficulties in an area with co-circulation of the two viruses. Our goal was to use Zika and Dengue serologic assays to build a classification model that improves upon the PPV of commercial kits while maintaining sensitivity. Methods We conducted a prospective longitudinal study in Southern Mexico where DENV and ZIKV co-circulation occurs (NCT02831699). Patients were included in two cohorts: a cohort of subjects presenting with a febrile rash meeting WHO/PAHO Zika case definition and a household cohort. After signed consent, all subjects enrolled were evaluated on study-visit Days 0, 3 and 7 (for fever rash cohort) and 28. We considered a subject “true positive” for ZIKV or DENV if RT-PCR positive at any time point. The healthy household cohort (with no positive RT-PCR) was considered “true negatives.” We fit a statistical decision tree taking as inputs serial serology measurements and outputting a predicted disease category. Funded in part by the NCI Contract No. HHSN261200800001E. Funded in part by the Mexican Ministry of Health. Results As of March 2018, we have 32 subjects in the Zika PCR+ group, 32 in the Dengue PCR+ group, and 68 in the household group. Our decision tree (Figure 1) achieved PPV of at least 90% on all three disease categories, while maintaining sensitivity above 50%. The highest PPV achieved by the kit manufacturer recommended cutoffs while maintaining a sensitivity of at least 10% on Zika PCR+ subjects is 30/114 (26%), and for Dengue PCR+ subjects is 21/30 (70%). Conclusion Using serology data in a statistical decision tree improves the PPV exhibited by the kit manufacturer recommendations while still maintaining respectable sensitivity. Physicians in regions with co-circulating flaviviruses should be aware of the pitfalls of using only RT-PCR or using pre-established commercial cutoffs in the serology kits for diagnosis. Disclosures All authors: No reported disclosures.

BACKGROUND Bio-geographical differences in fungal infection distribution have been observed around the world, confirming that climatic conditions are decisive in colonization. This research is focused on the impact of ultraviolet radiation (UV) on Aspergillus species, based on the consideration that an increase in UV-B radiation may have large ecological effects. RESULTS Conidia of six mycotoxigenic Aspergillus species isolated from vineyards located in the northeast and south of Spain were incubated for 15 days under light/dark cycles and temperatures between 20 and 30 °C per day. Additionally, 6 h of exposure to UV-A or UV-B radiation per day were included in the light exposure. UV irradiance used were 1.7 ± 0.2 mW cm−2 of UV-A (peak 365 nm) and 0.10 ± 0.2 mW cm−2 of UV-B (peak 312 nm). The intrinsic decrease in viability of conidia over time was accentuated when they were UV irradiated. UV-B radiation was more harmful. CONCLUSION Conidial sensitivity to UV light was marked in Aspergillus section Circumdati. Conidia pigmentation could be related to UV sensitivity. Different resistance was observed within species belonging to sections Flavi and Nigri. An increase in UV radiation could lead to a reduction in the Aspergillus spp. inoculum present in the field (vineyards, nuts, cereal crops). In addition, it could unbalance the spore species present in the field, leading to a higher predominance of dark-pigmented conidia. © 2015 Society of Chemical Industry

Compound heterozygous and homozygous (comp/hom) mutations in solute carrier family 34, member 3 (SLC34A3), the gene encoding the sodium (Na(+))-dependent phosphate cotransporter 2c (NPT2c), cause hereditary hypophosphatemic rickets with hypercalciuria (HHRH), a disorder characterized by renal phosphate wasting resulting in hypophosphatemia, correspondingly elevated 1,25(OH)2 vitamin D levels, hypercalciuria, and rickets/osteomalacia. Similar, albeit less severe, biochemical changes are observed in heterozygous (het) carriers and indistinguishable from those changes encountered in idiopathic hypercalciuria (IH). Here, we report a review of clinical and laboratory records of 133 individuals from 27 kindreds, including 5 previously unreported HHRH kindreds and two cases with IH, in which known and novel SLC34A3 mutations (c.1357delTTC [p.F453del]; c.G1369A [p.G457S]; c.367delC) were identified. Individuals with mutations affecting both SLC34A3 alleles had a significantly increased risk of kidney stone formation or medullary nephrocalcinosis, namely 46% compared with 6% observed in healthy family members carrying only the wild-type SLC34A3 allele (P=0.005) or 5.64% in the general population (P

We describe conservation built on local expertise such that it constitutes a hybrid form of traditional and bureaucratic knowledge. Researchers regularly ask how local knowledge might be applied to programs linked to protected areas. By examining the production of conservation knowledge in southern Mexico, we assert local expertise is already central to conservation. However, bureaucratic norms and social identity differences between lay experts and conservation practitioners prevent the public valuing of traditional knowledge. We make this point by contrasting 2 examples. The first is a master's thesis survey of local experts regarding the biology of the King Vulture (Sarcoramphus papa) in which data collection took place in communities adjacent to the Calakmul Biosphere Reserve. The second is a workshop sponsored by the same reserve that instructed farmers on how to monitor endangered species, including the King Vulture. In both examples, conservation knowledge would not have existed without traditional knowledge. In both examples, this traditional knowledge is absent from scientific reporting. On the basis of these findings, we suggest conservation outcomes may be improved by recognizing the knowledge contributions local experts already make to conservation programming. Mejorando los Resultados de la Conservacion con la Percepcion de Expertos y Burocratas Locales Haenn et al. Resumen La conservacion se describe como construida sobre el conocimiento local, de tal forma que constituye una forma hibrida del conocimiento tradicional y el burocratico. Los investigadores preguntan regularmente como el conocimiento local puede aplicarse a programas relacionados con areas protegidas. Al examinar la produccion del conocimiento de la conservacion en el sur de Mexico, afirmamos que el conocimiento local ya es central para la conservacion. Sin embargo, las normas burocraticas y las diferencias en la identidad social entre los expertos consagrados y los practicantes de la conservacion previenen la valoracion publica del conocimiento tradicional. Este punto lo hacemos al contrastar dos ejemplos. El primero es la encuesta de una tesis de maestria a los expertos locales con respecto a la biologia del zopilote rey (Sarcoramphus papa) en la cual la colecta de datos tuvo lugar en las comunidades adyacentes a la Reserva de la Biosfera de Calakmul. El segundo es un taller patrocinado por la misma reserva que instruyo a los granjeros sobre como monitorear a las especies en peligro, incluido el zopilote rey. En ambos ejemplos el conocimiento de la conservacion no podria haber existido sin el conocimiento tradicional. En ambos ejemplos, este conocimiento tradicional esta ausente del reporte cientifico. Con base en estos hallazgos, sugerimos que los resultados de la conservacion pueden mejorarse al reconocer las contribuciones para el conocimiento que los expertos locales ya hacen a la programacion de la conservacion.

Pseudohypoparathyroidism type-Ia (PHP-Ia), characterized by renal proximal tubular resistance to parathyroid hormone (PTH), results from maternal mutations of GNAS that lead to loss of α-subunit of the stimulatory G protein (Gαs) activity. Gαs expression is paternally silenced in the renal proximal tubule, and this genomic event is critical for the development of PTH resistance, as patients display impaired hormone action only if the mutation is inherited maternally. The primary clinical finding of PHP-Ia is hypocalcemia, which can lead to various neuromuscular defects including seizures. PHP-Ia patients frequently do not present with hypocalcemia until after infancy, but it has remained uncertain whether PTH resistance occurs in a delayed fashion. Analyzing reported cases of PHP-Ia with documented GNAS mutations and mice heterozygous for disruption of Gnas, we herein determined that the manifestation of PTH resistance caused by the maternal loss of Gαs, ie, hypocalcemia and elevated serum PTH, occurs after early postnatal life. To investigate whether this delay could reflect gradual development of paternal Gαs silencing, we then analyzed renal proximal tubules isolated by laser capture microdissection from mice with either maternal or paternal disruption of Gnas. Our results revealed that, whereas expression of Gαs mRNA in this tissue is predominantly from the maternal Gnas allele at weaning (3 weeks postnatal) and in adulthood, the contributions of the maternal and paternal Gnas alleles to Gαs mRNA expression are equal at postnatal day 3. In contrast, we found that paternal Gαs expression is already markedly repressed in brown adipose tissue at birth. Thus, the mechanisms silencing the paternal Gαs allele in renal proximal tubules are not operational during early postnatal development, and this finding correlates well with the latency of PTH resistance in patients with PHP-Ia.

Background Syphilis has increased among people living with HIV (PLWH), particularly in men who have sex with men (MSM). Syphilis screening test should be routinely offered as part of comprehensive care for PLWH. We assessed the annual frequency of syphilis testing and positive test results in patients actively receiving care for HIV in Mexico City and its associated factors. Methods We retrospectively analyzed cohort data of patients receiving care for HIV in our clinic (2001–2017). We estimated the annual proportion of PLWH receiving care that was tested for Syphilis and the annual prevalence of Syphilis infection among those tested. We performed annual, cross-sectional analysis in patients receiving care. We included patients with at least one registered visit to the clinic each year and defined those that were engaged in care, as those with at least two visits separated by at least 3 months in a given year. To identify factors associated with being tested and with being infected we used multivariate logistic regression models with random intercepts and GEE using sex, age, education, socioeconomic status (SES), year, time in care, use of ART, being naïve and having AIDS at enrollment, being engaged in care, and number of hospitalizations as independent variables. Results 2896 patients were included. The proportion of patients tested for syphilis increased from 21% in 2001 to 81% in 2017. The prevalence of syphilis increased from 7% to 21% during the same period with an early peak in 2003–2004 (Figure 1). Over time, the prevalence of re-infections increased substantially and accounted for the most positive test from 2013 onwards. Men, younger age, MSM, increasing education, time receiving care, number of hospitalizations and being engaged in care during the year were positively associated with being tested (Figure 2). Men, younger age, MSM, patients not receiving ARTs, those with AIDS at enrollment, and not being engaged in care were at higher risk of infection (Figure 3). Conclusion Annual VDRL testing increased over time, as did prevalence of syphilis, particularly after 2010 (9% vs. 21%). Re-infections were frequent. Efforts are needed to increase screening among women, those newly enrolled in care, and with low SES. Preventive strategies are needed for men, particularly MSM, early after enrollment. Disclosures All authors: No reported disclosures.

Pseudohypoparathyroidism type Ib (PHP1B) is characterized primarily by resistance to parathyroid hormone (PTH) and thus hypocalcemia and hyperphosphatemia, in most cases without evidence for Albright hereditary osteodystrophy (AHO). PHP1B is associated with epigenetic changes at one or several differentially-methylated regions (DMRs) within GNAS, which encodes the α-subunit of the stimulatory G protein (Gsα) and splice variants thereof. Heterozygous, maternally inherited STX16 or GNAS deletions leading to isolated loss-of-methylation (LOM) at exon A/B alone or at all maternal DMRs are the cause of autosomal dominant PHP1B (AD-PHP1B). In this study, we analyzed three affected individuals, the female proband and her two sons. All three revealed isolated LOM at GNAS exon A/B, whereas the proband's healthy maternal grandmother and uncle showed normal methylation at this locus. Haplotype analysis was consistent with linkage to the STX16/GNAS region, yet no deletion could be identified. Whole-genome sequencing of one of the patients revealed a large heterozygous inversion (1,882,433 bp). The centromeric breakpoint of the inversion is located 7,225 bp downstream of GNAS exon XL, but its DMR showed no methylation abnormality, raising the possibility that the inversion disrupts a regulatory element required only for establishing or maintaining exon A/B methylation. Because our three patients presented phenotypes consistent with PHP1B, and not with PHP1A, the Gsα promoter is probably unaffected by the inversion. Our findings expand the spectrum of genetic mutations that lead to LOM at exon A/B alone and thus biallelic expression of the transcript derived from this alternative first GNAS exon. © 2017 American Society for Bone and Mineral Research.

XLαs, a variant of the stimulatory G protein α-subunit (Gsα), can mediate receptor-activated cAMP generation and, thus, mimic the actions of Gsα in transfected cells. However, it remains unknown whether XLαs can act in a similar manner in vivo. We have now generated mice with ectopic transgenic expression of rat XLαs in the renal proximal tubule (rptXLαs mice), where Gsα mediates most actions of PTH. Western blots and quantitative RT-PCR showed that, while Gsα and type-1 PTH receptor levels were unaltered, protein kinase A activity and 25-hydroxyvitamin D 1-α-hydroxylase (Cyp27b1) mRNA levels were significantly higher in renal proximal tubules of rptXLαs mice than wild-type littermates. Immunohistochemical analysis of kidney sections showed that the sodium-phosphate cotransporter type 2a was modestly reduced in brush border membranes of male rptXLαs mice compared to gender-matched controls. Serum calcium, phosphorus, and 1,25 dihydroxyvitamin D were within the normal range, but serum PTH was ∼30% lower in rptXLαs mice than in controls (152 ± 16 vs. 222 ± 41 pg/ml; P < 0.05). After crossing the rptXLαs mice to mice with ablation of maternal Gnas exon 1 (E1m−/+), male offspring carrying both the XLαs transgene and maternal Gnas exon 1 ablation (rptXLαs/E1m−/+) were significantly less hypocalcemic than gender-matched E1m−/+ littermates. Both E1m−/+ and rptXLαs/E1m−/+ offspring had higher serum PTH than wild-type littermates, but the degree of secondary hyperparathyroidism tended to be lower in rptXLαs/E1m−/+ mice. Hence, transgenic XLαs expression in the proximal tubule enhanced Gsα-mediated responses, indicating that XLαs can mimic Gsα in vivo.