Degranulating Neutrophils Promote Leukotriene B4 Production by Infected Macrophages To Kill Leishmania amazonensis Parasites.

Neutrophils mediate early responses against pathogens, and they become activated during endothelial transmigration toward the inflammatory site. In the current study, human neutrophils were activated in vitro with immobilized extracellular matrix proteins, such as fibronectin (FN), collagen, and laminin. Neutrophil activation by FN, but not other extracellular matrix proteins, induces the release of the granules' contents, measured as matrix metalloproteinase 9 and neutrophil elastase activity in culture supernatant, as well as reactive oxygen species production. Upon contact with Leishmania amazonensis-infected macrophages, these FN-activated neutrophils reduce the parasite burden through a mechanism independent of cell contact. The release of granule proteases, such as myeloperoxidase, neutrophil elastase, and matrix metalloproteinase 9, activates macrophages through TLRs, leading to the production of inflammatory mediators, TNF-α and leukotriene B4 (LTB4), which are involved in parasite killing by infected macrophages. The pharmacological inhibition of degranulation reverted this effect, abolishing LTB4 and TNF production. Together, these results suggest that FN-driven degranulation of neutrophils induces the production of LTB4 and TNF by infected macrophages, leading to the control of Leishmania infection.

Vascular endothelial function is improved by oral glycine treatment in aged rats.

Glycine has been used to reduce oxidative stress and proinflammatory mediators in some metabolic disorders; however, its effect on the vasculature has been poorly studied. The aim of this work was to explore the effect of glycine on endothelial dysfunction in aged rats. Aortic rings with intact or denuded endothelium were obtained from untreated or glycine-treated male Sprague-Dawley rats at 5 and 15 months of age. Concentration-response curves to phenylephrine (PHE) were obtained from aortic rings incubated with N(G)-nitro-l-arginine methyl ester (l-NAME), superoxide dismutase (SOD), indomethacin, SC-560, and NS-398. Aortic mRNA expression of endothelial nitric oxide synthase (eNOS), NADPH oxidase 4 (NOX-4), cyclooxygenase 1 (COX-1), cyclooxygenase 2 (COX-2), tumour necrosis factor (TNF)-α, and interleukin-1 ß was measured by real time RT-PCR. The endothelial modulation of the contraction by PHE was decreased in aortic rings from aged rats. Glycine treatment improved this modulator effect and increased relaxation to acetylcholine. Glycine augmented the sensitivity for PHE in the presence of l-NAME and SOD. It also reduced the contraction by incubation with indomethacin, SC-560, and NS-398. Glycine increased the mRNA expression of eNOS and decreased the expression of COX-2 and TNF-α. Glycine improved the endothelium function in aged rats possibly by enhancing eNOS expression and reducing the role of superoxide anion and contractile prostanoids that increase the nitric oxide bioavailability.

Role of Perivascular Adipose Tissue in Aorta Reactivity from Obese and Hyperglycemic CD-1 Mice: New Insights into Perivascular Adipose Tissue.

Background: Perivascular adipose tissue (PVAT) plays an essential role in cardiovascular homeostasis. However, during obesity and diabetes, its role in vascular tone regulation is unclear. This study aimed to evaluate the function of the PVAT on aorta reactivity in the lean and cafeteria (CAF) diet-induced obese-hyperglycemic mice model. Methods: Aorta reactivity to phenylephrine, KCl, and acetylcholine was analyzed in lean (n = 6) and obese mice (n = 6). Also, nitric oxide (NO-) and cyclooxygenase participation, in the presence (n = 6) and absence (n = 6) of PVAT, were examined in the aortas. Results: After a CAF diet for 19 weeks, obese mice showed increased body weight, glucose intolerance, and hypercholesterolemia concerning lean mice. Vascular reactivity to phenylephrine was reduced significantly in the aorta of obese mice. In contrast, the contraction produced by KCl (80 mM) was increased in the aorta of obese mice independent of PVAT. Acetylcholine-induced vasorelaxation diminished in the aortas of obese mice in the presence of PVAT. Nonselective inhibition of cyclooxygenases likely shows that PVAT and endothelium release vasorelaxant prostanoids. Conclusions: The results suggest that PVAT modulates aorta reactivity by releasing NO-, decreasing the α1-adrenergic response to phenylephrine, and probably releasing vasorelaxant prostanoids. The data suggest that PVAT regulates the vascular smooth muscle and endothelial function in a CAF diet-induced obese-hyperglycemic mice model.

Leishmania infantum Defective in Lipophosphoglycan Biosynthesis Interferes With Activation of Human Neutrophils.

Visceral leishmaniasis (VL) is often associated with hematologic manifestations that may interfere with neutrophil response. Lipophosphoglycan (LPG) is a major molecule on the surface of Leishmania promastigotes, which has been associated with several aspects of the parasite-vector-host interplay. Here, we investigated how LPG from Leishmania (L.) infantum, the principal etiological agent of VL in the New World, influences the initial establishment of infection during interaction with human neutrophils in an experimental setting in vitro. Human neutrophils obtained from peripheral blood samples were infected with either the wild-type L. infantum (WT) strain or LPG-deficient mutant (∆lpg1). In this setting, ∆lpg1 parasites displayed reduced viability compared to WT L. infantum; such finding was reverted in the complemented ∆lpg1+LPG1 parasites at 3- and 6-h post-infection. Confocal microscopy experiments indicated that this decreased survival was related to enhanced lysosomal fusion. In fact, LPG-deficient L. infantum parasites more frequently died inside neutrophil acidic compartments, a phenomenon that was reverted when host cells were treated with Wortmannin. We also observed an increase in the secretion of the neutrophil collagenase matrix metalloproteinase-8 (MMP-8) by cells infected with ∆lpg1 L. infantum compared to those that were infected with WT parasites. Furthermore, collagen I matrix degradation was found to be significantly increased in ∆lpg1 parasite-infected cells but not in WT-infected controls. Flow cytometry analysis revealed a substantial boost in production of reactive oxygen species (ROS) during infection with either WT or ∆lpg1 L. infantum. In addition, killing of ∆lpg1 parasites was shown to be more dependent on the ROS production than that of WT L. infantum. Notably, inhibition of the oxidative stress with Apocynin potentially fueled ∆lpg1 L. infantum fitness as it increased the intracellular parasite viability. Thus, our observations demonstrate that LPG may be a critical molecule fostering parasite survival in human neutrophils through a mechanism that involves cellular activation and generation of free radicals.

DC-SIGN Mediates the Interaction Between Neutrophils and Leishmania amazonensis-Infected Dendritic Cells to Promote DC Maturation and Parasite Elimination.

Background: Leishmaniasis is a neglected arthropod-borne disease that affects millions of people worldwide. Successful Leishmania infections require the mitigation of immune cell functions leading to parasite survival and proliferation. A large body of evidence highlights the involvement of neutrophils (PMNs) and dendritic cells (DCs) in the establishment of immunological responses against these parasites. However, few studies, contemplate to what extent these cells interact synergistically to constrain Leishmania infection. Objective: We sought to investigate how PMNs and infected DCs interact in an in vitro model of Leishmania amazonensis infection. Material and Methods: Briefly, human PMNs and DCs were purified from the peripheral blood of healthy donors. Next, PMNs were activated with fibronectin and subsequently co-cultured with L. amazonensis-infected DCs. Results: We observed that L. amazonensis-infected DC exhibited lower rates of infection when co-cultivated with either resting or activated PMNs. Surprisingly, we found that the release of neutrophil enzymes was not involved in Leishmania killing. Next, we showed that the interaction between PMNs and infected-DCs was intermediated by DC-SIGN, further suggesting that parasite elimination occurs in a contact-dependent manner. Furthermore, we also observed that TNFα and ROS production was dependent on DC-SIGN-mediated contact, as well as parasite elimination is dependent on TNFα production in the co-culture. Finally, we observed that direct contact between PMNs and DCs are required to restore the expression of DC maturation molecules during L. amazonensis infection. Conclusion: Our findings suggest that the engagement of direct contact between PMNs and L. amazonensis-infected DC via DC-SIGN is required for the production of inflammatory mediators with subsequent parasite elimination and DC maturation.

Confirmation of neonatal screening: reference intervals and evaluation of methodological changes in TSH measurement.

Neonatal reference values for serum thyrotropin are scarce and comprise only small numbers of patients. During 2006, changes were made in IMMULITE kits for TSH measurement. To validate methodological changes, 80 serum samples from patients were evaluated and to establish reference intervals, 334 neonates and infants were analyzed (divided into 4 groups). Group 1 (G1) (48-72 h of life) (n=153), group 2A (G2A) (7-10 days of life) (n=65), group 2B (G2B) (11-14 days of life) (n=35), group 3 (G3) (28-40 days of life) (n=81). Current kits overestimate TSH results by 26 to 37%; TSH (mIU/L) reference intervals (percentile 2.5-97.5) were G1 (1.1-12.7), G2A (1.8-9.8), G2B (1.1-7.1) (p < 0.03 vs. G2A), G3 (1.2-6.9). We suggest that during the second week of life, reference values should be divided into an early stage and a late stage, at least, for there to be an adequate interpretation of borderline measurements in newborn thyroid screening.

Efecto ecotóxico y preventivo del uso de aceites esenciales en la mastitis bovina / Ecotoxic and preventive effect of the use of essential oils in bovine mastitis

The essential oils of Lippia citriodora (Ort.) and Lippia origanoides (Kunth) have shown antimicrobial activity associated with mastitis. The objective of this study was to evaluate its ecotoxic effect with the Artemia salina bioassay and the prevention of mastitis through an in vivo test in cattle (n=20) with a product based on these oils using a conventional product as a control. Contact hypersensitivity, the effect on somatic cells, and residuality in mil k samples were evaluated. The results of the Artemia salina bioassay were 10.05 and 19.36 (µg/mL) respectively. No negative effects or contact hypersensitivity were observed, and no residual metabolites were found in post - test milk. The somatic cell count showed 75% effectiveness in the prevention of mastitis with essential oils compared to 62.5% with the conventional product. The evaluated formulation could be used in the prevention of bovine mastitis safely, further investigation is required.

Los aceites esenciales de Lippia citriodora (Ort.) y Lippia origanoides (Kunth), han mostrado acti vidad antimicrobiana asociada a la mastitis. El objetivo de este estudio fue evaluar su efecto ecotóxico con el bioensayo Artemia salina y la prevención de mastitis mediante un ensayo in vivo en bovinos (n=20) con un producto a base de estos aceites utiliz ando como control un producto convencional. Se evaluó la hipersensibilidad de contacto, efecto en células somáticas y residualidad en muestra de leche. Los resultados del bioensayo de Artemia salina fueron 10,05 y 19,36 (µg/mL) respectivamente. No se obser varon efectos negativos, ni hipersensibilidad de contacto, y no se encontraron metabolitos residuales en leche posterior al ensayo. El conteo de células somáticas mostró efectividad en la prevención de mastitis del 75% con aceites esenciales frente al 62.5 % del producto convencional. La formulación evaluada podría ser utilizada en la prevención de la mastitis bovina de forma segura, se requiere profundizar en la investigación.

Screening of essential oils against oxacillin - resistant Staphylococcus aureus strains isolated from bovine mastitis / Screening de aceites esenciales frente a Staphylococcus aureus oxacilino - resistentes extraidos de aislados de mastitis bovina

Bovine mastitis is a disease wi th far - reaching consequences for the dairy industry. Staphylococcus aureus is a pathogen that is especially resistant to antibiotics. The objective of this study was to evaluate the antimicrobial activity of the essential oils Lippia citriodora (Lam.), Thy mus vulgaris (L), and a mixture of the essential oils Lippia citriodora and Thymus vulgaris (50/50 v/v), against isolates of oxacillin - resistant Staphylococcus aureus (n=15) of positive cases of bovine mastitis. For the statistical analysis, the IBM SPSS s tatistical package was used. The mixture of essential oils ( Lippia citriodora and Thymus vulgaris (50/50 v/v)) obtained the most significant antimicrobial activity in relation to pure essential oils. It is therefore concluded that the mixture of these oils boosts their antimicrobial activity ( p <0.05). The minimum inhibitory and bactericidal concentration of this mixture for the total isolations was 12 µL/L and 25 µL/mL, respectively.

La mastitis bovina es una enfermedad de gran impacto para la industria lechera. El Staphylococcus aureus es uno de los principales patógenos, especialmente aquellos resistentes a los antibióticos. El objetivo de este estudio fue evaluar la actividad antimicrobiana de los aceites esenciales de Lippia citriodora (Lam.), Thymus vulgaris (L), y una mezcla de aceites esenciales de Lippia citriodora y Thymus vulgaris (50/50 v/v), frente a aislamientos clínicos de Staph ylococcus aureus oxacilino - resistentes (n=15) de mastitis bovina. Se utilizó p rograma estadístico IBM SPSS y se concluyó la diferencia significativa a un p <0.05. La mezcla de aceites esenciales ( Lippia citriodora y Thymus vulgaris (50/50 v/v)), obtuvo la m ayor actividad antimicrobiana en relación a los aceites esenciales puros, se concluye que la mezcla de estos aceites potencia su actividad antimicrobiana ( p <0.019). La concentración mínima inhibitoria y bactericida de esta mezcla fue del 12 µL/mL y 25 µL/m L, respectivamente, y puede ser una alternativa terapéutica.

Corrigendum: Immunization of Experimental Dogs With Salivary Proteins From Lutzomyia longipalpis, Using DNA and Recombinant Canarypox Virus Induces Immune Responses Consistent With Protection Against Leishmania infantum.

[This corrects the article DOI: 10.3389/fimmu.2018.02558.].

A novel ABCA12 pathologic variant identified in an Ecuadorian harlequin ichthyosis patient: A step forward in genotype-phenotype correlations.

BACKGROUND: Autosomal recessive congenital ichthyoses (ARCI) have been associated with different phenotypes including: harlequin ichthyosis (HI), congenital ichthyosiform erythroderma (CIE), and lamellar ichthyosis (LI). While pathogenic variants in all ARCI genes are associated with LI and CIE phenotypes, the unique gene associated with HI is ABCA12. In HI, the most severe ARCI form, pathogenic variants in both ABCA12 gene alleles usually have a severe impact on protein function. The presence of at least one non-truncating variant frequently causes a less severe congenital ichthyosis phenotype (LI and CIE). METHODS: We report the case of a 4-year-old Ecuadorian boy with a severe skin disease. Genetic diagnosis was performed by NGS. In silico predictions were performed using Alamut software v2.11. A review of the literature was carried out to identify all patients carrying ABCA12 splice-site and missense variants, and to explore their genotype-phenotype correlations. RESULTS: Genetic testing revealed a nonsense substitution, p.(Arg2204*), and a new missense variant, p.(Val1927Leu), in the ABCA12 gene. After performing in silico analysis and a comprehensive review of the literature, we conclude that p.(Val1927Leu) affects a well conserved residue which could either disturb the protein function or alter the splicing process, both alternatives could explain the severe phenotype of our patient. CONCLUSION: This case expands the spectrum of ABCA12 reported disease-causing variants which is important to unravel genotype-phenotype correlations and highlights the importance of missense variants in the development of HI.

Immunization with LJM11 salivary protein protects against infection with Leishmania braziliensis in the presence of Lutzomyia longipalpis saliva.

Leishmania is transmitted in the presence of sand fly saliva. Protective immunity generated by saliva has encouraged identification of a vector salivary-based vaccine. Previous studies have shown that immunization with LJM11, a salivary protein from Lutzomyia longipalpis, is able to induce a Th1 immune response and protect mice against bites of Leishmania major-infected Lutzomyia longipalpis. Here, we further investigate if immunization with LJM11 recombinant protein is able to confer cross-protection against infection with Leishmania braziliensis associated with salivary gland sonicate (SGS) from Lutzomyia intermedia or Lu. longipalpis. Mice immunized with LJM11 protein exhibited an increased production of anti-LJM11 IgG, IgG1 and IgG2a and a DTH response characterized by an inflammatory infiltrate with the presence of CD4+ IFN-γ+ T cells. LJM11-immunized mice were intradermally infected in the ear with L. braziliensis in the presence of Lu. longipalpis or Lu. intermedia SGS. A significant reduction of parasite numbers in the ear and lymph node in the group challenged with L. braziliensis plus Lu. longipalpis SGS was observed, but not when the challenge was performed with L. braziliensis plus Lu. intermedia SGS. A higher specific production of IFN-γ and absence of IL-10 by lymph node cells were only observed in LJM11 immunized mice after infection. After two weeks, a similar frequency of CD4+ IFN-γ+ T cells was detected in LJM11 and BSA groups challenged with L. braziliensis plus Lu. longipalpis SGS, suggesting that early events possibly triggered by immunization are essential for protection against Leishmania infection. Our findings support the specificity of saliva-mediated immune responses and reinforce the importance of identifying cross-protective salivary antigens.

Immunization of Experimental Dogs With Salivary Proteins From Lutzomyia longipalpis, Using DNA and Recombinant Canarypox Virus Induces Immune Responses Consistent With Protection Against Leishmania infantum.

Metacyclic Leishmania promastigotes are transmitted by sand flies that inject parasites and saliva into the host's skin. Previous studies have demonstrated that DNA plasmids encoding Lutzomyia longipalpis salivary proteins LJM17 and LJL143, when used to immunize dogs, resulted in a systemic and local Th1 cell-mediated immunity that interfered in parasite survival in vitro. Here we evaluated the ability of these same salivary antigens to induce anti-Leishmania immunity and to confer protection by immunizing dogs using a novel vaccination strategy more suitable for use in the field. The strategy consisted of a single dose of plasmid followed by two doses of recombinant Canarypoxvirus (rCanarypoxvirus) expressing L. longipalpis salivary proteins (LJM17 or LJL143). Thirty days after the final immunization, dogs were intradermally challenged with 107Leishmania infantum promastigotes in the presence of L. longipalpis saliva. We followed the experimentally infected dogs for 10 months to characterize clinical, parasitological, and immunological parameters. Upon vaccination, all immunized dogs presented strong and specific humoral responses with increased serum concentrations of IFN-γ, TNF, IL-7, and IL-15. The serum of dogs immunized with LJM17 also exhibited high levels of IL-2, IL-6, and IL-18. L. infantum infection was established in all experimental groups as evidenced by the presence of anti-Leishmania IgG, and by parasite detection in the spleen and skin. Dogs immunized with LJM17-based vaccines presented higher circulating levels of IFN-γ, IL-2, IL-6, IL-7, IL-15, IL-18, TNF, CXCL10, and GM-CSF post-infection when compared with controls. Results demonstrated that relevant Leishmania-specific immune responses were induced following vaccination of dogs with L. longipalpis salivary antigen LJM17 administered in a single priming dose of plasmid DNA, followed by two booster doses of recombinant Canarypox vector. Importantly, a significant increase in pro-inflammatory cytokines and chemokines known to be relevant for protection against leishmaniasis was evidenced after challenging LJM17-vaccinated dogs as compared to controls. Although similar results were observed following immunization with LJL143, the pro-inflammatory response observed after immunization was attenuated following infection. Collectively, these data suggest that the LJM17-based vaccine induced an immune profile consistent with the expected protective immunity against canine leishmaniosis. These results clearly support the need for further evaluation of the LJM17 antigen, using a heterologous prime-boost vaccination strategy against canine visceral leishmaniosis (CVL).

Male-female differences in 6-sulfatoxymelatonin excretion in hypopituitary patients.

OBJECTIVE: To evaluate melatonin secretion in adult hypopituitary patients with Growth Hormone deficiency (AGHD) on and off replacement therapy. SUBJECTS AND METHODS: We studied 48 subjects: 12 (6 males) untreated AGHD (AGHDnt), 20 (10 males) treated AGHD (AGHDt) and 16 healthy subjects (8 males) as control group (CG). We measured urinary 6-sulfatoxymelatonin (6-SM) in total (24 h samples), nocturnal (6-SMn): 1800-0800 and diurnal samples (6-SMd): 0800-1800. RESULTS: Significant differences were observed among the 3 groups of male subjects, in total 6-SM (p < 0.05), nocturnal 6-SM (p < 0.02) and nighttime-daytime delta values (p < 0.003). CG had significantly higher values than the AGHDnt in total 6-SM (p < 0.01), nocturnal 6-SM (p < 0.05) and nighttime-daytime delta values (p < 0.01). AGHDt patients showed significantly higher levels in nighttime-daytime delta values than AGHDnt patients (p < 0.05). In females, no significant differences were found among the 3 groups studied in total, nocturnal, diurnal or nighttime-daytime delta values. In males, significant correlations were found among total 6-SM (r = 0.58; p = 0.029), nocturnal 6-SM (r = 0.70; p = 0.006) and nighttime-daytime delta values (r = 0.71; p = 0.004) vs. serum IGF-1 levels in subjects evaluated. In females, significant correlations were found among total 6-SM (r = 0.57; p = 0.02) vs. serum IGF-1 levels in subjects evaluated. A tendency towards a significant correlation was found in diurnal 6-SM (r = 0.48; p = 0.07). CONCLUSIONS: Our findings show a sexual dimorphism in 6-SM excretion in AGHD patients and provide an interesting approach to a further understanding of some chronobiological disorders involved in GH deficiency.

Burden of Growth Hormone Deficiency and Excess in Children.

Longitudinal growth results from multifactorial and complex processes that take place in the context of different genetic traits and environmental influences. Thus, in view of the difficulties in comprehension of the physiological mechanisms involved in the achievement of normal height, our ability to make a definitive diagnosis of GH impairment still remains limited. There is a myriad of controversial aspects in relation to GH deficiency, mainly related to diagnostic controversies and advances in molecular biology. This might explain the diversity in therapeutic responses and may also serve as a rationale for new "nonclassical" treatment indications for GH. It is necessary to acquire more effective tools to reach an adequate evaluation, particularly while considering the long-term implications of a correct diagnosis, the cost, and safety of treatments. On the other hand, overgrowth constitutes a heterogeneous group of different pathophysiological situations including excessive somatic and visceral growth. There are overlaps in clinical and molecular features among overgrowth syndromes, which constitute the real burden for an accurate diagnosis. In conclusion, both GH deficiency and overgrowth are a great dilemma, still not completely solved. In this chapter, we review the most burdensome aspects related to short stature, GH deficiency, and excess in children, avoiding any details about well-known issues that have been extensively discussed in the literature.

Case-control study of mosquito nets against malaria in the Amazon region of Colombia.

The degree of effectiveness of mosquito nets against malaria in the Americas has remained uncertain. We carried out a case-control study of net use and mild malaria in the Amazonas state of Colombia. Two hundred ninety cases were enrolled via the Health Department services, and 977 community-based controls matched for age, sex, and place of residence. We found that a large proportion of the population (96% of controls) slept under nets. Nevertheless, we found a benefit of impregnated nets compared with no net use: adjusted odds ratio (OR) for mild malaria 0.44, 95% confidence interval (CI) 0.20-0.98. Nonimpregnated nets had a benefit that was only slightly smaller but not statistically significant (OR for mild malaria 0.54, 95% CI 0.25-1.18). Travel in the previous month had an odds ratio of 6.2 (95% CI 3.1-8.8) and a population attributable fraction of 13% compared with 11% for failure to use an impregnated net. We conclude that, in the Amazon region, promotion of mosquito net use and impregnation is justified, and that there is a need for measures to protect travelers from malaria.

Decreased 6-Sulfatoxymelatonin Excretion in Male GH-Deficient Children and Adolescents.

AIM: To evaluate melatonin secretion in a group of untreated and treated male growth hormone (GH)-deficient children and adolescents. METHODS: We studied 44 male subjects: 8 untreated GH-deficient patients (GHDnt), 16 treated GH-deficient patients (GHDt) and 20 healthy children and adolescents as control group (CG). We measured urinary 6-sulfatoxymelatonin (6-SM) in total (24-hour samples), nocturnal (18.00-8.00 h) and diurnal samples (8.00-18.00 h). Levels of 6-SM were expressed as micrograms excreted per time interval and x0394; values (difference between nighttime and daytime values). RESULTS: Significant differences were observed among the 3 groups of pediatric subjects studied for total 6-SM (p < 0.0001), nocturnal 6-SM (p < 0.0001) and x0394; values (p < 0.0001). Subsequent analysis showed significantly higher levels for total 6-SM, nocturnal 6-SM and nighttime-daytime x0394; in the CG versus the GHDnt (p < 0.01) and in the CG versus the GHDt group (p < 0.01). No significant correlations were found between 6-SM excretion and insulin-like growth factor-1 values in the children and adolescents studied. CONCLUSIONS: GH-deficient patients showed lower levels of 6-SM. Our findings provide a different insight to a further understanding of some chronobiological disorders involved in GH deficiency in children.

Evaluation of diagnostic accuracy of insulin-like growth factor (IGF)-I and IGF-binding protein-3 in growth hormone-deficient children and adults using ROC plot analysis.

We critically evaluated the diagnostic value of IGF-I and IGF-binding protein-3 (IGFBP-3) in GH deficiency (GHD) in children and adults using receiver operating characteristic (ROC) plot analysis. Sixty-six children (chronological age, 1.3-15 yr) were studied: 34 GHD and 32 idiopathic short stature (ISS). Ninety-two adults (chronological age, 18-70 yr) were also evaluated: 72 GHD, 34 of childhood onset (AGHD-CO), and 38 of adult onset (AGHD-AO); and 20 healthy volunteers. The SD score (SDS) for IGF-I was calculated from 596 normal subjects (212 children and 384 adults), and the SDS for IGFBP-3 was calculated from 350 normal subjects (212 children and 138 adults). The ROC plot showed that the best IGF-I SDS cut-off line was -1.65 for children [sensitivity (S), 68%; specificity (Sp), 97%, diagnostic efficiency (DEf), 81%], the cut-off line for AGHD was -1.65 for AGHD-CO (S, 91%; Sp, 100%; DEf, 94%), and the cut-off line for AGHD-AO was -1.80 (S, 81%; Sp, 100%; DEf, 88%). For IGFBP-3 SDS, the best cut-off line was -1.80 for children (S, 90%; Sp, 60%; DEf, 78%); it was -1.45 for AGHD-CO (S, 90%; Sp, 75%; DEf, 82%) and -0.90 for AGHD-AO (S, 90%; Sp, 68%; DEf, 77%). An accurate diagnosis was obtained using IGF-I SDS alone in GHD children 65%; ISS, 97%; AGHD-CO, 92%; AGHD-AO, 86%, with IGFBP-3 SDS alone in GHD children 60%; ISS, 90%; AGHD-CO, 75%; AGHD-AO, 68%. Considering both, an accurate diagnosis was obtained in GHD children 60%; ISS, 87%; AGHD-CO, 71%; AGHD-AO, 64%. In conclusion, our findings support the need to use cut-off lines expressed in SDS obtained using an appropriate statistical methodology for better characterization of the various clinical presentations. IGF-I proved to be more useful because of its good diagnostic efficiency and accuracy in both children and adults, whereas IGFBP-3 did not significantly contribute to the diagnosis of GHD.

Basal ultrasensitive LH assay: a useful tool in the early diagnosis of male pubertal delay?

The aim of this study was to evaluate the usefulness of basal measurements of gonadotropins in distinguishing between constitutionally delayed puberty (DP) and hypogonadotropic hypogonadism (HH), comparing its diagnostic efficiency with that of the dynamic GnRH infusion test (0.83 microg/min during 120 min). We studied 20 males, chronological age (CA) 14-18 years, with a final diagnosis of DP (n = 8), partial HH (n = 5) and complete HH (n = 7), confirmed by follow-up. We also evaluated basal samples of ultrasensitive LH and FSH in 117 healthy control males (CA 2-19 yr), classified according to Tanner stage. In the control group, ROC plot analysis showed a cutoff to differentiate prepuberty from puberty of 0.65 IU/l for LH (sensitivity: 91%, specificity: 98%). Differences were found (p < 0.05) in basal LH and in maximal responses to GnRH in complete HH in relation to DP and partial HH. The diagnostic efficiency of the GnRH infusion test was 85%. For basal LH, a cut-off limit of 0.65 IU/l showed a diagnostic efficiency of 85% for complete HH and 100% for partial HH and DP. We conclude that, in our experience, basal LH levels above 0.65 IU/l measured by ultrasensitive assay would rule out a complete deficiency. It was not possible to differentiate DP from partial HH, either in basal samples or with the infusion test.

Tumoral versus non-tumoral hyperprolactinemia in children and adolescents: possible usefulness of the domperidone test.

UNLABELLED: The aim of this study was to evaluate the usefulness of the domperidone test for the difficult diagnosis between functional and tumoral hyperprolactinemia. We evaluated 36 patients, aged 5-18 years, 14 (12 F, 2 M) with hyperprolactinemia (non-tumoral: 10; pituitary adenoma: 4) and 22 individuals as a control group (prepubertal: 5 F, 8 M; pubertal: 4 F, 5 M). Basal prolactin (PRL) (IRMA-DPC), T4 and TSH and PRL 30 min post-domperidone (0.2 mg/kg b. wt i.v.) were measured. Non-tumoral hyperprolactinemic females showed basal PRL: 45 (29-80) (median and range) ng/ml; post-domperidone: 208 (116-290) ng/ml; delta PRL (PRL 30' - PRL 0'): 167 (77-252) ng/ml; and PRL ratio (PRL 30'/PRL 0'): 3.9 (2.3-7.6). Females with pituitary adenoma showed basal PRL: 129 (125-660) ng/ml; post-domperidone: 202 (150-535) ng/ml; delta PRL: 73 (25-135) ng/ml; and ratio: 1.2 (0.8-1.6). Two males, one with a non-tumoral hyperprolactinemia and the other one with a pituitary adenoma, presented, respectively, PRL 0':45, 160; PRL 30':130, 173; delta: 85, 13; ratio: 2.9, 1.1. All non-tumoral patients showed a PRL ratio (30'/0') > 2.3, while no patient with pituitary adenoma had a ratio > 1.6. CONCLUSIONS: PRL response to domperidone allowed us to characterize hyperprolactinemias, although the presence of a blunted response should be confirmed in a larger number of patients with tumors with 'low' PRL levels (dependence on etiology or basal PRL level?).

Effect of the parental origin of the X-chromosome on the clinical features, associated complications, the two-year-response to growth hormone (rhGH) and the biochemical profile in patients with turner syndrome.

BACKGROUND: It is possible that genes on the X chromosome are expressed differently depending of its parental origin. The objective of this study was to determine the influence of the parental origin of the X-chromosome on phenotypic variability, response to rhGH and on the biochemical profile of TS patients. METHODS: This was a cross-sectional multicenter correlational study carried out over three years in six Latin-American university hospitals. Unrelated 45,X TS patients (n = 93; 18.3 ± 8.5 years )) were evaluated. A subgroup (n = 34) of the patients were prospectively treated with rhGH over two years. DNA profiles of patients and their mothers were compared to determine the parental origin of the retained X-chromosome through 10 polymorphic X-chromosome-STRs. The association with clinical features, biochemical profiles and anthropometric data at the beginning and after two years of rhGH treatment was determined. RESULTS: Seventy two percent of patients retained the maternal X chromosome (Xm). A trend towards significance between maternal height and patients final height (p ≤ 0.07) in 45,Xm subjects was observed. There was no correlation between paternal height and patient height. No differences were detected between both groups in regard to dysmorphic features, classical malformations or increase in the height-SDS after rhGH. There were higher levels of triglycerides, total and LDL cholesterol in patients >20 years who retained the Xm. CONCLUSIONS: The parental origin of the retained X chromosome may influence lipid metabolism in TS patients, but its effect on growth seems to be minimal. No parental-origin-effect on the phenotypic features, associated anomalies and on the growth response to rhGH was found in 45,X TS individuals.