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The lead optimization stage of a drug discovery program generally involves the design, synthesis, and assaying of hundreds to thousands of compounds. The design phase is usually carried out via traditional medicinal chemistry approaches and/or structure-based drug design (SBDD) when suitable structural information is available. Two of the major limitations of this approach are (1) difficulty in rapidly designing potent molecules that adhere to myriad project criteria, or the multiparameter optimization (MPO) problem, and (2) the relatively small number of molecules explored compared to the vast size of chemical space. To address these limitations, we have developed AutoDesigner, a

The lead optimization stage of a drug discovery program generally involves the design, synthesis and assaying of hundreds to thousands of compounds. The design phase is usually carried out via traditional medicinal chemistry approaches and/or structure based drug design (SBDD) when suitable structural information is available. Two of the major limitations of this approach are (1) difficulty in rapidly designing potent molecules that adhere to myriad project criteria, or the multiparameter optimization (MPO) problem, and (2) the relatively small number of molecules explored compared to the vast size of chemical space. To address these limitations we have developed AutoDesigner, a de novo design algorithm. AutoDesigner employs a cloud-native, multi-stage search algorithm to carry out successive rounds of chemical space exploration and filtering. Millions to billions of virtual molecules are explored and optimized while adhering to a customizable set of project criteria such as physicochemical properties and potency. Additionally, the algorithm only requires a single ligand with measurable affinity and a putative binding model as a starting point, making it amenable to the early stages of a SBDD project where limited data is available. To assess the effectiveness of AutoDesigner, we applied it to the design of novel inhibitors of D-amino acid oxidase (DAO), a target for the treatment of schizophrenia. AutoDesigner was able to generate and efficiently explore over 1 billion molecules to successfully address a variety of project goals. The compounds generated by AutoDesigner that were synthesized and assayed (1) simultaneously met not only physicochemical criteria, clearance and central nervous system (CNS) penetration (Kp,uu) cutoffs, but also potency thresholds; (2) fully utilize structural data to discover and explore novel interactions and a previously unexplored subpocket in the DAO active site. The reported data demonstrate that AutoDesigner can play a key role in accelerating the discovery of novel, potent chemical matter within the constraints of a given drug discovery lead optimization campaign.

ObjectiveTurner Syndrome is associated with several phenotypic conditions associated with a higher risk of subsequent comorbidity. We aimed to evaluate the prevalence of congenital malformations and the occurrence of age-related comorbid conditions and to determine whether the frequencies of congenital and acquired conditions depend on X chromosome gene dosage, as a function of karyotype subgroup.Design and methodsThis national retrospective observational cohort study includes 1501 patients. We evaluated the prevalence of congenital malformations and the cumulative incidence of subsequent specific comorbidities at five-year intervals, from the ages of 10 to 30 years, with stratification by karyotype subgroup: 45,X (n = 549), 45,X/46,isoXq (n = 280), 46,X,r(X)/46,XX (n = 106), 45,X/46,XX (n = 221), presence of Y (n = 87).ResultsMedian age was 9.4 (3.7–13.7) years at first evaluation and 16.8 (11.2–21.4) years at last evaluation. Congenital heart (18.9%) malformations were more frequent in 45,X patients, and congenital renal (17.2%) malformations were more frequent in 45,X, 45,X/46,isoXq and 46,X,r(X)/46,XX patients than in those with 45,X/46,XX mosaicism or a Y chromosome (P ConclusionThese data suggest that X gene chromosome dosage, particularly for Xp genes, contributes to the risk of developing comorbidities.

Highlights • In myocardial ischemia, the integrity of the cardiac sarcolemma is severely stressed in the critical earliest moments upon reperfusion. Bolstering sarcolemma integrity improves myocyte survival. • This review focuses on cardiac sarcolemma stability and its role as a therapeutic target in ischemia-reperfusion injury. • Synthetic block copolymers have been shown to interface with the muscle membrane to confer membrane stabilization during stress. • Integrated multidisciplinary research teams, spanning cardiology, physiology, chemistry, and chemical engineering are essential to guide future mechanistic and translational studies of novel chemical-based membrane stabilizers for preserving viable heart muscle during ischemia-reperfusion injury in human patients., Summary The phospholipid bilayer membrane that surrounds each cell in the body represents the first and last line of defense for preserving overall cell viability. In several forms of cardiac and skeletal muscle disease, deficits in the integrity of the muscle membrane play a central role in disease pathogenesis. In Duchenne muscular dystrophy, an inherited and uniformly fatal disease of progressive muscle deterioration, muscle membrane instability is the primary cause of disease, including significant heart disease, for which there is no cure or highly effective treatment. Further, in multiple clinical forms of myocardial ischemia-reperfusion injury, the cardiac sarcolemma is damaged and this plays a key role in disease etiology. In this review, cardiac muscle membrane stability is addressed, with a focus on synthetic block copolymers as a unique chemical-based approach to stabilize damaged muscle membranes. Recent advances using clinically relevant small and large animal models of heart disease are discussed. In addition, mechanistic insights into the copolymer-muscle membrane interface, featuring atomistic, molecular, and physiological structure-function approaches are highlighted. Collectively, muscle membrane instability contributes significantly to morbidity and mortality in prominent acquired and inherited heart diseases. In this context, chemical-based muscle membrane stabilizers provide a novel therapeutic approach for a myriad of heart diseases wherein the integrity of the cardiac muscle membrane is at risk., Central Illustration

An all-atom phospholipid bilayer and triblock copolymer model was developed for molecular dynamics (MD) studies. These were performed to investigate the mechanism of interaction between membrane-stabilizing triblock copolymer P188 and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) lipid bilayers under applied lateral surface tension (γ) to model membrane mechanical stress. Results showed that P188 insertion is driven by the hydrophobic poly(propylene oxide) (PPO) core and dependent on bilayer area per lipid. Moreover, insertion of P188 increased the bilayer's resistance to mechanical rupture, as observed by a significant increase in the absolute lateral pressure required to disrupt the bilayer. To further investigate the specific chemical features of P188 underlying membrane stabilizer function, a series of MD simulations with triblock copolymers of the same class as P188 but of varying chemical composition and sizes were performed. Results showed that triblock copolymer insertion into the lipid bilayer is dependent on overall copolymer hydrophobicity, with higher copolymer hydrophobicity requiring a reduced bilayer area per lipid ratio for insertion. Further analysis revealed that the effect of copolymer insertion on membrane mechanical integrity was also dependent on hydrophobicity. Here, P188 insertion significantly increased the absolute apparent lateral pressure required to rupture the POPC bilayer, thereby protecting the membrane against mechanical stress. In marked contrast, highly hydrophobic copolymers decreased the lateral pressure necessary for membrane rupture and thus rendering the membrane significantly more susceptible to mechanical stress. These new in silico findings align with recent experimental findings using synthetic lipid bilayers and in muscle cells in vitro and mouse models in vivo. Collectively, these data underscore the importance of PEO-PPO-PEO copolymer chemical composition in copolymer-based muscle membrane stabilization in vitro and in vivo. All-atom modeling with MD simulations holds promise for investigating novel copolymers with enhanced membrane interacting properties.

Poloxamer 188, a triblock copolymer of poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO), protects cellular membranes from various stresses. Though numerous block copolymer variants exist, evaluation of alternative architecture, composition, and size has been minimal. Herein, cultured murine myoblasts are exposed to the stresses of hypotonic shock and isotonic recovery, and membrane integrity was evaluated by quantifying release of lactate dehydrogenase. Comparative evaluation of a systematic set of PEO-PPO diblock and PEO-PPO-PEO triblock copolymers demonstrates that the diblock architecture can be protective in vitro. Short PPO blocks hinder protection with >9 PPO units needed for protection at 150 µM and >16 units needed at 14 µM. Addition of a tert-butyl end group enhances protection at reduced concentration. When the end group and PPO length are fixed, increasing the PEO length improves protection. This systematic evaluation establishes a new in vitro screening tool for evaluating membrane-sealing amphiphiles and provides mechanistic insight to guide future copolymer design for membrane stabilization in vivo.

The scientific premise, design, and structure-function analysis of chemical-based muscle membrane stabilizing block copolymers are reviewed here for applications in striated muscle membrane injury. Synthetic block copolymers have a rich history and wide array of applications from industry to biology. Potential for discovery is enabled by a large chemical space for block copolymers, including modifications in block copolymer mass, composition, and molecular architecture. Collectively, this presents an impressive chemical landscape to leverage distinct structure-function outcomes. Of particular relevance to biology and medicine, stabilization of damaged phospholipid membranes using amphiphilic block copolymers, classified as poloxamers or pluronics, has been the subject of increasing scientific inquiry. This review focuses on implementing block copolymers to protect fragile muscle membranes against mechanical stress. The review highlights interventions in Duchenne muscular dystrophy, a fatal disease of progressive muscle deterioration owing to marked instability of the striated muscle membrane. Biophysical and chemical engineering advances are presented that delineate and expand upon current understanding of copolymer-lipid membrane interactions and the mechanism of stabilization. The studies presented here serve to underscore the utility of copolymer discovery leading toward the therapeutic application of block copolymers in Duchenne muscular dystrophy and potentially other biomedical applications in which membrane integrity is compromised.

The heterotrimeric cardiac troponin complex is a key regulator of contraction and plays an essential role in conferring Ca(2+) sensitivity to the sarcomere. During ischemic injury, rapidly accumulating protons acidify the myoplasm, resulting in markedly reduced Ca(2+) sensitivity of the sarcomere. Unlike the adult heart, sarcomeric Ca(2+) sensitivity in fetal cardiac tissue is comparatively pH insensitive. Replacement of the adult cardiac troponin I (cTnI) isoform with the fetal troponin I (ssTnI) isoform renders adult cardiac contractile machinery relatively insensitive to acidification. Alignment and functional studies have determined histidine 132 of ssTnI to be the predominant source of this pH insensitivity. Substitution of histidine at the cognate position 164 in cTnI confers the same pH insensitivity to adult cardiac myocytes. An alanine at position 164 of cTnI is conserved in all mammals, with the exception of the platypus, which expresses a proline. Prolines are biophysically unique because of their innate conformational rigidity and helix-disrupting function. To provide deeper structure-function insight into the role of the TnC-TnI interface in determining contractility, we employed a live-cell approach alongside molecular dynamics simulations to ascertain the chemo-mechanical implications of the disrupted helix 4 of cTnI where position 164 exists. This important motif belongs to the critical switch region of cTnI. Substitution of a proline at position 164 of cTnI in adult rat cardiac myocytes causes increased contractility independent of alterations in the Ca(2+) transient. Free-energy perturbation calculations of cTnC-Ca(2+) binding indicate no difference in cTnC-Ca(2+) affinity. Rather, we propose the enhanced contractility is derived from new salt bridge interactions between cTnI helix 4 and cTnC helix A, which are critical in determining pH sensitivity and contractility. Molecular dynamics simulations demonstrate that cTnI A164P structurally phenocopies ssTnI under baseline but not acidotic conditions. These findings highlight the evolutionarily directed role of the TnI-cTnC interface in determining cardiac contractility.

Block copolymers can be synthesized in an array of architectures and compositions to yield diverse chemical properties. The triblock copolymer Poloxamer 188 (P188), the family archetype, consisting of a hydrophobic poly(propylene oxide) core flanked by hydrophilic poly(ethylene oxide) chains, can stabilize cellular membranes during stress. However, little is known regarding the molecular basis of membrane interaction by copolymers in living organisms. By leveraging diblock architectural design, discrete end-group chemistry modifications can be tested. Here we show evidence of an anchor and chain mechanism of interaction wherein titrating poly(propylene oxide) block end group hydrophobicity directly dictates membrane interaction and stabilization. These findings, obtained in cells and animals in vivo, together with molecular dynamics simulations, provide new insights into copolymer-membrane interactions and establish the diblock copolymer molecular architecture as a valuable platform to inform copolymer-biological membrane interactions. These results have implications for membrane stabilizers in muscular dystrophy and for other biological applications involving damaged cell membranes.

Troponin I (TnI) is the molecular switch of the sarcomere. Cardiac myocytes express two isoforms of TnI during development. The fetal heart expresses the slow skeletal TnI (ssTnI) isoform and shortly after birth ssTnI is completely and irreversibly replaced by the adult cardiac TnI (cTnI) isoform. These two isoforms have important functional differences; broadly, ssTnI is a positive inotrope, especially under acidic/hypoxic conditions, whereas cTnI facilitates faster relaxation performance. Evolutionary directed changes in cTnI sequence suggest cTnI evolved to favor relaxation performance in the mammalian heart. To investigate the mechanism, we focused on several notable TnI isoform and trans-species-specific residues located in TnI’s helix 4 using structure/function and molecular dynamics analyses. Gene transduction of adult cardiac myocytes by cTnIs with specific helix 4 ssTnI substitutions, Q157R/A164H/E166V/H173N (QAEH), and A164H/H173N (AH), were investigated. cTnI QAEH is similar in these four residues to ssTnI and nonmammalian chordate cTnIs, whereas cTnI AH is similar to fish cTnI in these four residues. In comparison to mammalian cTnI, cTnI QAEH and cTnI AH showed increased contractility and slowed relaxation, which functionally mimicked ssTnI expressing myocytes. cTnI QAEH molecular dynamics simulations demonstrated altered intermolecular interactions between TnI helix 4 and cTnC helix A, specifically revealing a new, to our knowledge, electrostatic interaction between R171of cTnI and E15 of cTnC, which structurally phenocopied the ssTnI conformation. Free energy perturbation calculation of cTnC Ca2+ binding for these conformations showed relative increased calcium binding for cTnI QAEH compared to cTnI. Taken together, to our knowledge, these new findings provide evidence that the evolutionary-directed coordinated acquisition of residues Q157, A164, E166, H173 facilitate enhanced relaxation performance in mammalian adult cardiac myocytes.

Cardiac troponin I (cTnI) functions as the molecular switch of the thin filament. Studies have shown that a histidine button engineered into cTnI (cTnI A164H) specifically enhances inotropic function in the context of numerous pathophysiological challenges. To gain mechanistic insight into the basis of this finding, we analyzed histidine ionization states in vitro by studying the myofilament biophysics of amino acid substitutions that act as constitutive chemical mimetics of altered histidine ionization. We also assessed the role of histidine-modified cTnI in silico by means of molecular dynamics simulations. A functional in vitro analysis of myocytes at baseline (pH 7.4) indicated similar cellular contractile function and myofilament calcium sensitivity between myocytes expressing wild-type (WT) cTnI and cTnI A164H, whereas the A164R variant showed increased myofilament calcium sensitivity. Under acidic conditions, compared with WT myocytes, the myocytes expressing cTnI A164H maintained a contractile performance similar to that observed for the constitutively protonated cTnI A164R variant. Molecular dynamics simulations showed similar intermolecular atomic contacts between the WT and the deprotonated cTnI A164H variant. In contrast, simulations of protonated cTnI A164H showed various potential structural configurations, one of which included a salt bridge between His-164 of cTnI and Glu-19 of cTnC. This salt bridge was recapitulated in simulations of the cTnI A164R variant. These data suggest that differential histidine ionization may be necessary for cTnI A164H to act as a molecular sensor capable of modulating sarcomere performance in response to changes in the cytosolic milieu.

Congenital isolated ACTH deficiency (IAD) is a rare disease characterized by low plasma ACTH and cortisol levels and preservation of all other pituitary hormones. This condition was poorly defined before we identified TPIT, a T-box transcription factor with a specific role in differentiation of the corticotroph lineage in mice and humans, as its principal molecular cause.We have enlarged our series of IAD patients to better characterize the phenotype and the genotype of this rare disease.Each exon of the TPIT gene was amplified and sequenced in IAD patients without any identified cause. A functional analysis of each new TPIT mutation was performed.We described the largest series of 91 IAD patients and identified three distinct groups: neonatal onset complete or partial IAD or late onset IAD. We did not identify any TPIT mutation in patients with partial or late-onset IAD. However, we found a TPIT mutation in 65% of patients with neonatal-onset complete IAD. These patients are homozygous or compound heterozygous for TPIT mutations, and their parents are healthy heterozygous carriers. We identified nine new mutations: four missense, one one-nucleotide deletion, three splice-site mutations, and one large deletion. TPIT mutations lead to loss of function by different mechanisms, such as non-sense-mediated mRNA decay, abnormal mRNA splicing, loss of TPIT DNA binding or protein-protein interaction defects.TPIT mutations are responsible for two thirds of neonatal-onset complete IAD but can not be detected in partial or late-onset IAD.

In cardiac muscle, the troponin (cTn) complex is a key regulator of myofilament calcium sensitivity because it serves as a molecular switch required for translating myocyte calcium fluxes into sarcomeric contraction and relaxation. Studies of several species suggest that ectotherm chordates have myofilaments with heightened calcium responsiveness. However, genetic polymorphisms in cTn that cause increased myofilament sensitivity to activating calcium in mammals result in cardiac disease including arrhythmias, diastolic dysfunction, and increased susceptibility to sudden cardiac death. We hypothesized that specific residue modifications in the regulatory arm of troponin I (TnI) were critical in mediating the observed decrease in myofilament calcium sensitivity within the mammalian taxa. We performed large-scale phylogenetic analysis, atomic resolution molecular dynamics simulations and modeling, and computational alanine scanning. This study provides evidence that a His to Ala substitution within mammalian cardiac TnI (cTnI) reduced the thermodynamic potential at the interface between cTnI and cardiac TnC (cTnC) in the calcium-saturated state by disrupting a strong intermolecular electrostatic interaction. This key residue modification reduced myofilament calcium sensitivity by making cTnI molecularly untethered from cTnC. To meet the requirements for refined mammalian adult cardiac performance, we propose that compensatory evolutionary pressures favored mutations that enhanced the relaxation properties of cTn by decreasing its sensitivity to activating calcium.

A case of endometrioma of the right inguinal canal region, diagnosed preoperatively, is presented. The diagnosis was made on the basis of cyclical symptoms relating to menstrual periods, in combination with demonstration of blood products within an enhancing focal lesion in the inguinal region with magnetic resonance imaging. The case presented here is unique, as it is the first case, to our knowledge, of an endometriotic lesion in the inguinal canal to demonstrate the characteristic 'shading sign' at magnetic resonance imaging.

Muscle membrane vulnerability is a hallmark of Duchenne Muscular Dystrophy (DMD), an X-linked disease that results in progressive skeletal muscle weakness and cardiomyopathy. Cardiac disease is an increasing cause of death in DMD and there is currently no cure for DMD. One unique therapeutic approach is the use of membrane sealants to protect the fragile dystrophic muscle membranes from mechanical stress. We propose a structure-function strategy in understanding the mechanism by which block copolymers may protect dystrophic cardiac and skeletal muscles. Poloxamer 188 (P188) is a membrane sealant that has been shown by us and others to protect the fragile dystrophic myocardium under physiological stress but appears to have reduced efficacy in affecting the skeletal myopathy. P188 belongs to the tri-block copolymer family, which comprises molecules made of a hydrophobic polypropylene oxide (PPO) core flanked by hydrophilic chains of polyethylene oxide (PEO) moieties. These copolymers exist at various molecular weights and PPO to PEO ratios and it is unknown what structural properties of P188 confer its membrane protecting functionality. Mechanistic knowledge is requisite for a deeper understanding of muscle membrane protection by copolymer sealants to enable therapeutic application. Interestingly, our data shows that P188 is efficacious in isolated dystrophic skeletal myofibers suggesting that poor delivery and low diffusion into the core of dystrophic whole muscle in vivo limits P188 effectiveness. Ultimately, fully effective therapeutic strategies must simultaneously target both cardiac and skeletal muscle tissues. We will present data on copolymer structure-function understanding and discuss how these new data will shed light into the structural requisite for more efficacious and potent membrane sealants for dystrophic skeletal muscle in vivo.

Sensorineural hearing defect and goiter are common features of Pendred's syndrome. The clinical diagnosis of Pendred's syndrome remains difficult because of the lack of sensitivity and specificity of the thyroid signs. The identification of PDS as the causative gene allowed molecular screening and enabled a re-evaluation of the syndrome to identify potential diagnostic characteristics. This report presents the clinical and genotypic findings of 30 French families, for whom a diagnosis of Pendred's syndrome had been made. Twenty-seven families had at least one mutated allele. Twenty-eight different mutations were identified, 11 of which had never been previously reported. The main clinical characteristics were: early hearing loss, fluctuation in terms of during deafness evolution, and the presence of an enlarged vestibular aqueduct.

Troponin I (TnI) has a central, isoform-dependent role in ischemic contractile failure. The fetal heart expresses the slow skeletal TnI (ssTnI) isoform which confers protection from ischemia-mediated contractile failure relative to the adult expressed cardiac TnI (cTnI) isoform. A single codon substitution in cTnI, A164H, reverts significantly to an ssTnI phenotype, in specifics, conferring protection from ischemia-mediated contractile failure. Importantly, unlike ssTnI, cTnI A164H does not alter contractile performance under baseline conditions. The molecular mechanisms for the marked enhancement in myocyte function under pathophysiological conditions with no detected effects under baseline conditions have yet to be determined. Isoform specific residues in helix 4 of TnI were investigated through structure/function analysis to gain insight into this mechanism. Molecular dynamics simulation for cTnI, Q157R/A164H/E166V/H173N cTnI (QAEH), A164H cTnI, and ssTnI in complex with cTnC, showed that substitution of cTnI with the ssTnI residues alters the intermolecular interactions between TnI and cTnC. These findings suggest that these residues are important for the conformation of helix 4 in regards to cTnC. To investigate if these substitutions alter function, adult cardiac myocytes were transduced with adenovirus expressing QAEH cTnI and sarcomere dynamics were analyzed. QAEH cTnI showed a similar phenotype to ssTnI, increasing sarcomere shortening and slowing relaxation at pH 7.4 while maintaining normal contractile function under acidic conditions. To elucidate this further, double and triple mutants were analyzed to determine the importance of each of the three residues to baseline contractility. The data show that H173 and Q157 are necessary for the reduced contractility at baseline while E166 exerts the opposite effect. Taken together, these new findings suggests that the conformation of helix 4 in TnI is an important determinate of contractility in ischemia.

Introduction La recuperation cerebrale etait consideree comme impossible apres 12 min d’ischemie globale jusqu’a recemment. Le Poloxamer 188 (P188) est un tensioactif synthetique connue pour minimiser les lesions de reperfusion. Il a ete montre que le postconditionnement ischemique (PCI) et le sevoflurane (SEV) reduisent les lesions de reperfusion secondaires a un arret cardiaque. Nous emettons l’hypothese qu’une therapie multiple (M-RCP) avec la combinaison de P188, PCI et SEV au debut de la reanimation cardiorespiratoire (RCP) ameliore l’hemodynamique durant la RCP, reduit la dysfonction ventriculaire post-reanimation, ameliore la survie sans sequelle neurologique a 24 h et reduit les dommages des organes vitaux apres un arret cardiaque prolonge dans un modele porcin. Materiel et methodes Apres une fibrillation ventriculaire (FV) de 15 min, une RCP standard a ete realisee chez les animaux temoins (n = 8) et comparee aux animaux du groupe M-RCP (n-13) qui ont subi une VF non traitee 17 min. La M-RCP etait constituee : a) P188 : bolus IV (250 mg/kg) au debut de la RCP suivie d’une perfusion de 250 mg/kg de P188 en 4 h apres le retour a une circulation spontanee (RACS), b) PCI : reanimation cardiopulmonaire avec compression decompression active, valve d’impedance inspiratoire pendant 20 s, puis 20 s de pause pendant 3 cycles et c) SEV : insufflation de SEV 2 % pendant les 3 premieres min. L’adrenaline (0,5 mg) a ete utilisee chez tous les animaux. La survie et le score de categorie de performance cerebrale (CPC) ont ete evalues a 24 h. La Troponine, CK-MB, creatinine et fonction hepatique ont ete evaluees a 4 h. Un test exact de Fischer et un test t non apparie ont ete utilises pour l’analyse statistique. Resultats L’hemodynamique est ameliore durant la M-RCP ( Fig. 1 ) durant la RCP. Tous les animaux atteints ont eu un RACS. La FEVG etait ameliore (63 ± 13 a 1 h, 61 ± 8 a 4 h) vs temoins (38 ± 12 a 1 h, 36 ± 18 a 4 h) (p Fig. 1 ). Discussion Une therapie multiple qui cible les lesions de reperfusion au niveau des membranes mitochondriale et cellulaire ameliore les parametres hemodynamiques au cours de RCP et facilite la reanimation cerebrale, cardiaque et des organes vitaux apres un arret cardiaque non traitee prolongee.

Many factors contribute to the growth failure of chronic renal failure: water and electrolytes disturbances, hypertonicity, phosphate or calcium wasting, secondary hyperparathyroidism, anemia, hypertension, metabolic acidosis, and malnutrition. In addition, the pubertal growth spurt is usually stunted. Growth hormone (GH) resistance is observed with low GH binding protein (GHBP) level, and normal or low IGF I levels despite elevated GH level. Elevated IGFBP levels may contribute to a reduced IGF activity, especially in dialysed patients. Glucocorticoid therapy in transplanted patients further contribute to poor growth and inhibited IGF I activity. As conventional treatments have a limited effect to improve growth, adult height is often far below -2 SD. GH therapy has proved to be successful, especially in young children, overpassing the hormonal resistance so that an adult height within the normal range may be reached.

Muscle membrane vulnerability is a hallmark of Duchenne Muscular Dystrophy (DMD), an X-linked neuromuscular disease that results in progressive skeletal muscle weakness and significant cardiomyopathy. We propose the use of membrane-sealant copolymers as an innovative and unique potential therapeutic for DMD. The tri-block copolymer family comprises molecules made of a hydrophobic polypropylene oxide (PPO) core flanked bilaterally by linear chains of hydrophilic polyethylene oxide (PEO) chemical moieties. Block copolymers exist at various molecular weights and PPO/PEO ratios and it is currently not known what structural properties confer membrane sealing capacity to copolymers of this family. We and other groups have shown that one such sealant, poloxamer P188, protects dystrophic hearts in vitro and in vivo in both small and large animal models of DMD but its apparent efficacy is significantly reduced in skeletal muscle in vivo. This underscores the importance of discovering more potent membrane sealants to treat all striated muscles in DMD. We have initiated a collaborative structure-function approach by implementing and refining an in vitro membrane injuring osmotic and shear stress assay to analyze the membrane sealing functions of these tri-block copolymer family members on dystrophic skeletal muscle to systematically determine the effects of PPO/PEO ratio and molecular weight on membrane protection. We will present progress on copolymer structure-function understanding and discuss how these new data will shed light into the structural requisite for more efficacious and potent membrane sealants.

Syringomyelia is rare in children aged less than 10 years, and bladder dysfunction is an unlikely first manifestation. This report describes a case of repeated episodes of acute urinary retention in a young girl revealing syringomyelia and Arnold-Chiari malformation.A 2.5 year-old girl was admitted because she was suffering from acute urinary retention. Her poor appetite had been treated with cyproheptadine, a histamine type I blocking drug. Clinical investigation revealed no local cause for this bladder dysfunction except moderate spasticity of the legs. Cystography showed no vesicoureteral reflux. Because the episodes of urinary retention recurred each day, magnetic resonance imaging (MRI) was performed; this showed the typical features of syringomyelia extending from C5 to T11 plus Arnold-Chiari malformation. The cyproheptadine was discontinued and the urinary retention disappeared.Cyproheptadine may have revealed latent neurogenic bladder in this case, although urodynamic studies, performed 3 months later, detected no bladder dysfunction.

A case of cutaneous herpes relapse with meningitis is reported in a 1.5 month-old infant treated during the first three weeks of life with acyclovir (ACV) for a neonatal herpes infection. Such a relapse has previously been described in older children as well as in adults. In this case report, there was immunological response to herpes virus infection, 2.5 months after the onset of the infection. The relapse is discussed taking into account the mechanism of action of ACV, the age of the patient and the immunological response profile. Because of the high risk of neurological involvement, we suggest that the relapse should be treated with ACV for a period of time longer than actually recommended.

The aim of this study was to evaluate with magnetic resonance imaging (MRI) the hypothalamic and pituitary structures in patients with idiopathic isolated growth hormone deficiency (GHD) or multiple pituitary hormone deficiencies (MPHD).

Computerized axial tomographic scans (CAT scans) of thorax and abdomen were used as part of the initial staging procedure of 50 consecutive patients with small-cell bronchogenic carcinoma (SCBC). The aims were to define the extent of the primary tumor and metastatic spread, and to compare the information obtained from CAT scans with conventional staging procedures. With CAT scanning, spread of the primary tumor was found to be far more extensive than originally believed when defined by conventional assessment. Of 35 patients conventionally staged as localized T1 or T2 tumors, 27 (77%) had their stage increased to extensive T3 disease. similarly, subcarinal lymph nodes were found in one patient when conventional methods were used (2%), but 16 patients (32%) were found when CAT scan was used. Unsuspected adrenal metastases were found in eight patients (16%) and retrocrural nodes in four patients (8%). Thirty-seven patients (74%) were found to have Stage III tumors when conventional staging was used; 47 patients (94%) were found to be Stage III following CAT scanning, demonstrating that, within the thorax, localized tumors are very uncommon. CAT scanning demonstrates the degree of intrathoracic spread very clearly, and has a valuable role in the initial investigation of patients with SCBC if the planned treatment includes radiotherapy.

Computerized axial tomographic scans (CAT scans) of thorax and abdomen were used as part of the initial staging procedure of 50 consecutive patients with small-cell bronchogenic carcinoma (SCBC). The aims were to define the extent of the primary tumor and metastatic spread, and to compare the information obtained from CAT scans with conventional staging procedures. With CAT scanning, spread of the primary tumor was found to be far more extensive than originally believed when defined by conventional assessment. Of 35 patients conventionally staged as localized T1 or T2 tumors, 27 (77%) had their stage increased to extensive T3 disease. similarly, subcarinal lymph nodes were found in one patient when conventional methods were used (2%), but 16 patients (32%) were found when CAT scan was used. Unsuspected adrenal metastases were found in eight patients (16%) and retrocrural nodes in four patients (8%). Thirty-seven patients (74%) were found to have Stage III tumors when conventional staging was used; 47 patients (94%) were found to be Stage III following CAT scanning, demonstrating that, within the thorax, localized tumors are very uncommon. CAT scanning demonstrates the degree of intrathoracic spread very clearly, and has a valuable role in the initial investigation of patients with SCBC if the planned treatment includes radiotherapy.

The clinical outcomes of seven girls presenting with pseudosexual precocity caused by isolated autonomous ovarian follicular cysts are presented. Six of the seven girls, aged 11 months to 6.9 years, had a unilateral ovarian cyst detected by ultrasound at the first acute episode. Plasma oestradiol was raised in only five of the cases, but all had a low response to luteinising hormone releasing hormone stimulation. Follow up lasted for up to eight years with recurrent episodes of variable frequency and severity in all seven patients. Evidence of McCune-Albright syndrome appeared later in only three patients. It could not be predicted from the initial symptoms or the clinical course. Mutations of the G(s)alpha protein leading to activation were investigated in the lymphocytes and ovarian and bone tissues of four patients. Only one patient showed a mutation in bone tissue. Close follow up with repeated searches for skeletal lesions remains necessary since the distribution of somatic mutations cannot be assessed by molecular studies. Most patients with recurrent ovarian cysts require a conservative approach.

Cardiac troponin I (cTnI) functions as the molecular switch of the thin filament. Studies have shown that a A164H button engineered into cTnI enhances inotropic function under pathophysiological challenges. In vitro studies of myofilament calcium sensitivity and sarcomere shortening kinetics in intact and permeablized myocytes at baseline (pH 7.4) indicated similar cellular contractile function and myofilament calcium sensitivity between myocytes expressing wildtype cTnI and cTnI A164H while A164R showed a hypercontractile phenotype associated with increased myofilament calcium sensitivity. Under acidic conditions, compared to depressed function in myocytes with wildtype cTnI, myocytes expressing cTnI A164H maintained myofilament calcium sensitivity and contractile performance comparable to the calcium sensitizer cTnI A164R. The role of histidine modified cTnI was assessed by molecular dynamics (MD) simulations and pKa calculations of the wildtype and histidine or arginine-modified cTnI (148-173): cTnC (1-90) complex. The simulations showed similar conformations between the wildtype and the deprotonated cTnI A164H variant. In contrast, simulations of protonated cTnI A164H and cTnI A164R showed diverse conformation changes, both of which included the formation of a cTnI His 164 and cTnC Glu 19 salt bridge. pKa calculations showed no significant pKa shift for all ionizable residues except for cTnI His 164 and cTnC Glu 19 when the salt bridge is formed. The data shed light into the potential mechanism of pH activation of cTnI A164H and the importance of electrostatic interactions in governing the biophysical adjustments in troponin function necessary for nuanced modulation of myofilament function in response to changes in the cytosolic milieu.