Mandarin essential oil as an antimicrobial in ethanolic fermentation: Effects on Limosilactobacillus fermentum and Saccharomyces cerevisiae.

The antibacterial activity of citrus essential oils (EOs) in the context of combating Limosilactobacillus fermentum, one of the most important bacterial contaminants in the bioethanol production industry, has never been explored previously. Industrial processes usually utilize sulfuric acid for cell treatment to decrease bacterial contamination. However, due to the hazardous nature of sulfuric acid, an alternative to it is highly desirable. Therefore, in the present study, the efficacy of Fremont IAC 543 mandarin EO against a strain of L. fermentum (ATCC® 9338™) was evaluated under proliferative/nonproliferative conditions, in both pure culture and co-culture with an industrial strain of Saccharomyces cerevisiae. The mandarin EO exhibited higher effectiveness against L. fermentum compared to that against S. cerevisiae under nonproliferative conditions (added to water rather than to culture medium). At the concentration of 0·05%, the EO was as effective as the acid solution with pH 2·0 in reducing the count of L. fermentum almost 5 log CFU ml-1 cycles, while the concentration of 0·1% led to the complete loss of bacterial culturability. When L. fermentum was co-cultured with S. cerevisiae, the efficacy of the EO against the bacterial strain was reduced. However, despite this reduced efficacy in co-culture, mandarin EO may be considered effective in combating L. fermentum and could be applied in processes where this bacterium proves to be unfavourable and does not interact with S. cerevisiae.

GnRH analog is ineffective in increasing adult height in girls with puberty onset after 7 years of age: a systematic review and meta-analysis.

Objective We assessed the effectiveness of puberty blockade with a gonadotropin-releasing hormone (GnRH) analog in increasing adult height (AH) in girls with puberty onset between 7 and 10 years of age. Methods We performed a systematic review and included controlled studies in which girls with early puberty (EP) were assigned to the GnRH analog or no treatment groups. The primary outcome analyzed was AH. Search strategies were applied to the MEDLINE, EMBASE, LILACS and CENTRAL databases. Results We identified 1514 references, and six studies fulfilled our eligibility criteria. Two studies were randomized and four were not randomized. At the baseline of each trial, height, chronological age, bone age, predicted AH (PAH) and target height (TH) were equal between the groups. All studies used intramuscular triptorelin every 28 days in the intervention groups. The mean duration of the therapy was 2 years. Meta-analysis of AH among the six studies (comprising 332 girls) showed no significant difference between the groups (mean difference = 0.50 cm, 95% confidence interval = -0.72 to 1.73 cm, I 2 = 0%). In a sub-group analysis based on PAH (<155 cm and 155 cm and equal to TH), there was no difference in average AH between the groups. The quality of evidence according to the Grading of Recommendations Assessment, Development and Evaluation approach was low. Conclusion We found no evidence from controlled experimental and observational studies that compared with no treatment, the use of GnRH analogs improved AH in girls with EP.

Ethanol addition enhances acid treatment to eliminate Lactobacillus fermentum from the fermentation process for fuel ethanol production.

Fermentation is one of the most critical steps of the fuel ethanol production and it is directly influenced by the fermentation system, selected yeast, and bacterial contamination, especially from the genus Lactobacillus. To control the contamination, the industry applies antibiotics and biocides; however, these substances can result in an increased cost and environmental problems. The use of the acid treatment of cells (water-diluted sulphuric acid, adjusted to pH 2·0-2·5) between the fermentation cycles is not always effective to combat the bacterial contamination. In this context, this study aimed to evaluate the effect of ethanol addition to the acid treatment to control the bacterial growth in a fed-batch system with cell recycling, using the industrial yeast strain Saccharomyces cerevisiae PE-2. When only the acid treatment was used, the population of Lactobacillus fermentum had a 3-log reduction at the end of the sixth fermentation cycle; however, when 5% of ethanol was added to the acid solution, the viability of the bacterium was completely lost even after the first round of cell treatment. The acid treatment +5% ethanol was able to kill L. fermentum cells without affecting the ethanol yield and with a low residual sugar concentration in the fermented must. SIGNIFICANCE AND IMPACT OF THE STUDY: In Brazilian ethanol-producing industry, water-diluted sulphuric acid is used to treat the cell mass at low pH (2·0) between the fermentative cycles. This procedure reduces the number of Lactobacillus fermentum from 107 to 104  CFU per ml. However, the addition of 5% ethanol to the acid treatment causes the complete loss of bacterial cell viability in fed-batch fermentation with six cell recycles. The ethanol yield and yeast cell viability are not affected. These data indicate the feasibility of adding ethanol to the acid solution replacing the antibiotic use, offering a low cost and a low amount of residue in the biomass.

Sonic Hedgehog pathway is upregulated in adamantinomatous craniopharyngiomas.

OBJECTIVES: Pituitary stem cells play a role in the oncogenesis of human adamantinomatous craniopharyngiomas (aCPs). We hypothesized that crosstalk between the Wnt/ß-catenin and Sonic Hedgehog (SHH) pathways, both of which are important in normal pituitary development, would contribute to the pathogenesis of aCPs. DESIGN: To explore the mRNA and protein expression of components of the SHH signaling pathway in aCPs and their relationship with the identification of CTNNB1/ß-catenin mutations and patients outcomes. PATIENTS AND METHODS: In 18 aCP samples, CTNNB1 was sequenced, and the mRNA expression levels of SHH pathway members (SHH, PTCH1, SMO, GLI1, GLI2, GLI3, and SUFU) and SMO, GLI1, GLI3, SUFU, ß-catenin, and Ki67 proteins were evaluated by quantitative real-time PCR and immunohistochemistry respectively. Anterior normal pituitaries were used as controls. Associations between molecular findings and clinical data were analyzed. RESULTS: The aCPs presented higher mRNA expression of SHH (+400-fold change (FC); P<0.01), GLI1 (+102-FC; P<0.001), and GLI3 (+5.1-FC; P<0.01) than normal anterior pituitaries. Longer disease-free survival was associated with low SMO and SUFU mRNA expression (P<0.01 and P=0.02 respectively). CTNNB1/ß-catenin mutations were found in 47% of the samples. aCPs with identified mutations presented with higher mRNA expression of SMO and GLI1 (+4.3-FC; P=0.02 and +10.2-FC; P=0.03 respectively). SMO, GLI1, GLI3, and SUFU staining was found in 85, 67, 93, and 64% of the samples respectively. Strong GLI1 and GLI3 staining was detected in palisade cells, which also labeled Ki67, a marker of cell proliferation. CONCLUSIONS: The upregulation of SHH signaling occurs in aCPs. Thus, activation of Wnt/ß-catenin and SHH pathways, both of which are important in pituitary embryogenesis, appears to contribute to the pathogenesis of aCP.

Maternal mood and concordant maternal and infant salivary cortisol during heel lance while in kangaroo care.

BACKGROUND: Maternal kangaroo care (MKC) is a naturalistic intervention that alleviates neonatal pain, and mothers are assumed to play a stress regulatory role in MKC. Yet, no MKC infant pain study has examined relationship between maternal and infant stress reactivity concurrently, or whether post-partum depression and/or anxiety (PPDA) alters maternal and neonatal stress response and the regulatory effects of MKC. OBJECTIVES: To examine the concordance of salivary cortisol reactivity between 42 mothers and their stable preterm infants during routine infant heel lance (HL) while in MKC and to compare salivary cortisol between groups of mothers with and without PPDA and their infants. METHODS: Maternal and infant salivary cortisol samples were collected pre-HL and 20 min post-HL with two additional maternal samples at night and in the morning. Mothers and infants were allocated to with PPDA versus without PPDA study groups on the basis of maternal post-natal mental health assessment scores. RESULTS: Higher mothers' cortisol pre-HL was weakly associated with higher infants' salivary cortisol in response to the HL procedure. Maternal depression and/or anxiety were not associated with infants' cortisol. During HL, both groups of mothers and infants showed no change in salivary cortisol. CONCLUSIONS: Concordance between mother and infant salivary cortisol supports the maternal stress regulatory role in MKC. MKC may have stress regulatory benefits for mothers and their preterm infants during HL independent of PPDA. Future MKC studies that target mothers with altered mood will help to build on these findings.

Potassium metabisulphite as a potential biocide against Dekkera bruxellensis in fuel ethanol fermentations.

UNLABELLED: Dekkera bruxellensis is an important contaminant yeast of fuel ethanol fermentations in Brazil, whose system applies cell repitching between the fermentative cycles. This work evaluated the addition of potassium metabisulphite (PMB) on yeast growth and fermentative yields in pure and co-cultures of Saccharomyces cerevisiae and D. bruxellensis in two situations: addition to the acidic solution in which the cells are treated between the fermentative cycles or to the fermentation medium. In the range of 200-400 mg l(-1) , PMB was effective to control the growth of D. bruxellensis depending on the culture medium and strain. When added to the acidic solution (250 mg l(-1) ), a significant effect was observed in mixed cultures, because the inactivation of SO2 by S. cerevisiae most likely protected D. bruxellensis from being damaged by PMB. The physiological response of S. cerevisiae to the presence of PMB may explain the significant decrease in alcohol production. When added to the fermentation medium, PMB resulted in the control but not the death of D. bruxellensis, with less intensive effect on the fermentative efficiency. In co-culture with the addition of PMB, the fermentative efficiency was significantly lower than in the absence of PMB. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first to evaluate the action of potassium metabisulphite to control the growth of Dekkera bruxellensis in the fermentation process for fuel alcohol production. As near as possible of industrial conditions, the study simulates the addition of that substance in different points in the fermentation process, verifying in which situation the effects over the starter yeast and alcohol yield are minimal and over D. bruxellensis are maximal. Co-culture fermentations were carried out in cell-recycled batch system. The feasibility of using this substance for this specific fermentation is discussed in light of the possible biological and chemical interactions.

Polymorphisms of the adiponectin gene in gestational hypertension and pre-eclampsia.

Adiponectin is a hormone involved in energy homeostasis by regulating glucose and lipid metabolism. In addition, the adiponectin gene (ADIPOQ) has polymorphisms that can modulate the circulating concentration of adiponectin. Abnormal adiponectin levels have been associated with pre-eclampsia (PE); however, the influence of genetic polymorphisms on the development of hypertensive disorders of pregnancy is unclear. The aim of this study was to examine whether ADIPOQ polymorphisms are associated with gestational hypertension (GH) and/or PE. We studied 401 pregnant women: 161 healthy pregnant (HP), 113 pregnant with GH and 127 pregnant with PE. ADIPOQ polymorphisms -11391G>A (rs17300539), -11377C>G (rs266729), 45T>G (rs2241766) and 276G>T (rs1501299) were genotyped by allelic discrimination assays using real-time PCR. Haplotypes were inferred using the PHASE 2.1 program. We observed that the genotypic frequencies of the -11377C>G polymorphism were different in PE compared with HP (P<0.0125), with the CT genotype being more commonly found in PE patients than in HP women (P<0.0125). However, allelic frequencies of this single-nucleotide polymorphism were similar between PE and HP (P>0.0125). No difference was observed when GH and HP groups were compared (both P>0.0125). In addition, we found no difference in genotype or allele distributions for the -11391G>A, 45T>G and 276G>T polymorphisms when we compared GH or PE with HP (all P>0.0125). In conclusion, we found a modest association between the CG genotype of the -11377C>G polymorphism and PE.

Growth hormone pharmacogenetics: the interactive effect of a microsatellite in the IGF1 promoter region with the GHR-exon 3 and -202 A/C IGFBP3 variants on treatment outcomes of children with severe GH deficiency.

Insulin-like growth factor type 1 (IGF1) is a mediator of growth hormone (GH) action, and therefore, IGF1 is a candidate gene for recombinant human GH (rhGH) pharmacogenetics. Lower serum IGF1 levels were found in adults homozygous for 19 cytosine-adenosine (CA) repeats in the IGF1 promoter. The aim of this study was to evaluate the influence of (CA)n IGF1 polymorphism, alone or in combination with GH receptor (GHR)-exon 3 and -202 A/C insulin-like growth factor binding protein-3 (IGFBP3) polymorphisms, on the growth response to rhGH therapy in GH-deficient (GHD) patients. Eighty-four severe GHD patients were genotyped for (CA)n IGF1, -202 A/C IGFBP3 and GHR-exon 3 polymorphisms. Multiple linear regressions were performed to estimate the effect of each genotype, after adjustment for other influential factors. We assessed the influence of genotypes on the first year growth velocity (1st y GV) (n=84) and adult height standard deviation score (SDS) adjusted for target-height SDS (AH-TH SDS) after rhGH therapy (n=37). Homozygosity for the IGF1 19CA repeat allele was negatively correlated with 1st y GV (P=0.03) and AH-TH SDS (P=0.002) in multiple linear regression analysis. In conjunction with clinical factors, IGF1 and IGFBP3 genotypes explain 29% of the 1st y GV variability, whereas IGF1 and GHR polymorphisms explain 59% of final height-target-height SDS variability. We conclude that homozygosity for IGF1 (CA)19 allele is associated with less favorable short- and long-term growth outcomes after rhGH treatment in patients with severe GHD. Furthermore, this polymorphism exhibits a non-additive interaction with -202 A/C IGFBP3 genotype on the 1st y GV and with GHR-exon 3 genotype on adult height.

PP002. Study of polymorphisms of the adiponectin gene in women with gestational hypertension and preeclampsia.

INTRODUCTION: Adiponectin is involved in energy homeostasis by regulating glucose and lipid metabolism. Additionally, it presents anti-inflammatory and anti-atherosclerotic functions. Polymorphisms in adiponectin gene (ADIPOQ) can modulate the concentrations of adiponectin. The influence of these polymorphisms on the development of gestational hypertension (GH) and preeclampsia (PE) is unknown. OBJECTIVES: The aim of this work was to examine the influence of polymorphisms in the gene ADIPOQ on the development of gestational hypertension and preeclampsia. PATIENTS AND METHODS: We studied 401 pregnant women: 161 healthy pregnant (HP), 113 pregnant with gestational hypertension (GH) and 127 pregnant with preeclampsia (PE). Polymorphisms ADIPOQ -11391G>A (rs17300539), -11377C>G (rs266729), 45T>G (rs2241766) and 276G>T (rs1501299) were genotyped by allelic discrimination by PCR in real time. Haplotypes were inferred using the PHASE 2.1 program. RESULTS: There were no statistically significant differences in allele and genotype frequencies of the polymorphisms studied. In the analysis of haplotypes, we observed small differences in haplotype frequencies between groups, however, none of these differences was statistically significant (P>0.05). CONCLUSION: We found no association between the genotypic and allelic variants of the ADIPOQ gene polymorphisms with the development of gestational hypertension and preeclampsia.

Mineralocorticoid receptor p.I180V polymorphism: association with body mass index and LDL-cholesterol levels.

BACKGROUND/AIMS: Aldosterone and the mineralocorticoid receptor (MR) play a major role in sodium balance and blood pressure control. They are also involved in adipocyte metabolism. The aim of this study was to analyze the association between the MR p.I180V polymorphism with hypertension and markers of cardiovascular risk. DESIGN AND METHODS: Case-control study nested within a cohort of 2063 subjects followed since birth to date. All subjects (age 23-25 yr old) from the entire cohort with systolic and diastolic hypertension (no.=126) were paired with 398 normotensive controls. MR p.I180V genotype association with anthropometric and biochemical markers of cardiometabolic risk was tested. RESULTS: There was a significant association of the MR p.I180V genotype with body mass index (BMI) and LDL-cholesterol level (p<0.01). Hypertensive subjects carrying the polymorphic G allele (AG or GG genotypes) presented significantly higher BMI (30.0+/-6.0 vs 28.7+/-5.6 kg/m(2); p<0.01) and higher LDL-cholesterol (139.9+/-60.3 vs 109.9+/-35.5 mg/dl; p<0.01). The frequency of the polymorphism MR p.I180V was similar between hypertensive subjects and controls (p=0.15). CONCLUSIONS: The MR p.I180V polymorphism seems to be associated with cardiovascular risk factors including BMI and LDL-cholesterol levels. This original in vivo finding reinforces the role of MR in adipocyte biology and in cardiovascular disease.

Analysis of the putative regulatory region of the gastric inhibitory polypeptide receptor gene in food-dependent Cushing's syndrome.

Gastric inhibitory polypeptide (GIP)-dependent Cushing's syndrome (CS) results from the ectopic expression of non-mutated GIP receptor (hGIPR) in the adrenal cortex. We evaluated whether mutations or polymorphisms in the regulatory region of the GIPR gene could lead to this aberrant expression. We studied 9.0kb upstream and 1.3kb downstream of the GIPR gene putative promoter (pProm) by sequencing leukocyte DNA from controls and from adrenal tissues of GIP- and non-GIP-dependent CS patients. The putative proximal promoter region (800 bp) and the first exon and intron of the hGIPR gene were sequenced on adrenal DNA from nine GIP-dependent CS, as well as on leukocyte DNA of nine normal controls. Three variations found in this region were found in all patients and controls; at position -4/-5, an insertion of a T was seen in four out of nine patients and in five out of nine controls. Transient transfection studies conducted in rat GC and mouse Y1 cells showed that the TT allele confers loss of 40% in the promoter activity. The analysis of the 8-kb distal pProm region revealed eight distal single nucleotide polymorphisms (SNPs) without probable association with the disease, since frequencies in patients and controls were very similar. In conclusion, mutations or SNPs in the regulatory region of the GIPR gene are unlikely to underlie GIP-dependent CS.

Cerebral midline developmental anomalies: endocrine, neuroradiographic and ophthalmological features.

BACKGROUND: Hypopituitarism may occur in patients with midline cerebral defects (MCD), including septo-optic dysplasia (SOD). HESX1 gene mutations have been associated with SOD. OBJECTIVE: To evaluate the endocrine, ophthalmological and neuroradiographic abnormalities in 18 patients with MCD and SOD without mutations at the HESX1 locus. STUDY DESIGN: The diagnosis of hypothalamic and pituitary abnormalities was confirmed by clinical findings and basal hormone values or functional tests. All patients underwent ophthalmological examination and neuroradiologic studies by MRI. RESULTS: The diagnosis of hypothalamic and pituitary abnormalities varied from 3 days to 13 years. Endocrine abnormalities were found in 88% of the patients: GH deficiency (72%), hypothyroidism (66%), hypogonadism (45%), diabetes insipidus (27%), adrenal insufficiency (10%), and precocious puberty (5%). Psychomotor retardation was observed in 55% and seizures in 22%. Visual status varied from normal to blindness. MRI confirmed heterogeneous intracranial malformations. CONCLUSIONS: Our data support the need for systematic and periodic endocrine evaluation of patients with MCD using a multidisciplinary approach.

Glucocorticoid receptor gene polymorphisms in ACTH-secreting pituitary tumours.

OBJECTIVE: The inhibitory action of glucocorticoids on the hypothalamic-pituitary axis is disrupted in ACTH-secreting pituitary tumours. The molecular events leading to the development of these tumours and their relative resistance to glucocorticoids are unknown. We investigated the presence of mutations and polymorphisms of the glucocorticoid receptor (GR) gene in corticotropinoma and their possible relationship with the tissue-specific resistance to glucocorticoids. DESIGN AND METHODS: DNA or RNA was extracted from 18 corticotropinomas and the GR gene was amplified by the polymerase chain reaction (PCR) or reverse transcriptase-PCR followed by automated direct sequencing. RESULTS: We did not identify any mutation in the coding region and the exon-intron boundary regions of the GR gene. The polymorphism AAT > AGT at codon 363 (N363S) was found in 17% and the polymorphism AAT > AAC at codon 766 (N766N) in 11% of tumours, both in heterozygous state. The polymorphisms at codons 22 and 23, at introns 3 and 4, and at codon 618, previously described in normal population, were not observed. CONCLUSIONS: Our results show that GR gene mutations are rare and unlikely to contribute to the glucocorticoid resistance observed in corticotropinomas. Polymorphisms in the GR gene might confer a selective advantage to tumorigenesis in corticotropinoma. However, there was no relationship between GR gene polymorphism and clinical presentation, tumour size or surgery outcome, suggesting that tumour growth may not be directly related to alterations of the GR gene structure.

Salivary cortisol as a tool for physiological studies and diagnostic strategies

Salivary cortisol is an index of plasma free cortisol and is obtained by a noninvasive procedure. We have been using salivary cortisol as a tool for physiological and diagnostic studies, among them the emergence of circadian rhythm in preterm and term infants. The salivary cortisol circadian rhythm in term and premature infants was established between 8 and 12 postnatal weeks. In the preterm infants the emergence of circadian rhythm was parallel to the onset of sleep rhythm. We also studied the use of salivary cortisol for screening for Cushing's syndrome (CS) in control and obese outpatients based on circadian rhythm and the overnight 1 mg dexamethasone (DEX) suppression test. Salivary cortisol was suppressed to less than 100 ng/dl after 1 mg DEX in control and obese patients. A single salivary cortisol measurement at 23:00 h and again after 1 mg DEX above the 90th percentile of the obese group values had sensitivity and specificity of 93 and 93 percent (23:00 h), and 91 and 94 percent (after DEX), respectively. The sensitivity improved to 100 percent when we combined both parameters. We also studied 11 CS children and 21 age-matched primary obese children for whom salivary cortisol sensitivity and specificity were 100/95 percent (23:00 h), and 100/95 percent (1 mg DEX), respectively. Similar to adults, sensitivity and specificity of 100 percent were obtained by combining 23:00 h and 1 mg DEX. The measurement of salivary cortisol is a useful tool for physiological studies and for the diagnosis of CS in children and adults on an outpatient basis.

Salivary cortisol as a tool for physiological studies and diagnostic strategies.

Salivary cortisol is an index of plasma free cortisol and is obtained by a noninvasive procedure. We have been using salivary cortisol as a tool for physiological and diagnostic studies, among them the emergence of circadian rhythm in preterm and term infants. The salivary cortisol circadian rhythm in term and premature infants was established between 8 and 12 postnatal weeks. In the preterm infants the emergence of circadian rhythm was parallel to the onset of sleep rhythm. We also studied the use of salivary cortisol for screening for Cushing's syndrome (CS) in control and obese outpatients based on circadian rhythm and the overnight 1 mg dexamethasone (DEX) suppression test. Salivary cortisol was suppressed to less than 100 ng/dl after 1 mg DEX in control and obese patients. A single salivary cortisol measurement at 23:00 h and again after 1 mg DEX above the 90th percentile of the obese group values had sensitivity and specificity of 93 and 93% (23:00 h), and 91 and 94% (after DEX), respectively. The sensitivity improved to 100% when we combined both parameters. We also studied 11 CS children and 21 age-matched primary obese children for whom salivary cortisol sensitivity and specificity were 100/95% (23:00 h), and 100/95% (1 mg DEX), respectively. Similar to adults, sensitivity and specificity of 100% were obtained by combining 23:00 h and 1 mg DEX. The measurement of salivary cortisol is a useful tool for physiological studies and for the diagnosis of CS in children and adults on an outpatient basis.

The emergence of salivary cortisol circadian rhythm and its relationship to sleep activity in preterm infants.

OBJECTIVE: The circadian rhythm of cortisol is established at between 8 and 12 postnatal weeks in term infants. However, there is limited information about the effect of prematurity on this rhythm. We evaluated the emergence of the salivary cortisol circadian rhythm in premature infants and its relationship to the onset of sleep daily rhythm. DESIGN AND PATIENTS: A longitudinal study of a group of nine premature infants (gestational age 31-34 weeks) was performed. Salivary samples were obtained in the morning and at night at 2, 4, 8, 12, 16, 20 and 24 postnatal weeks and the babies' sleeping periods were recorded by their mothers. MEASUREMENTS: Cortisol was determined by RIA in 25-microl salivary samples. Two techniques based on assay coefficients of variation were used to characterize the circadian pattern of cortisol. RESULTS: Five infants (55%) established and maintained their cortisol rhythm at 2 and 8 postnatal weeks. In the remaining four infants the age of appearance was 12 and 16 weeks. This rhythm emerged in the group as a whole between 8 and 12 postnatal weeks. The circadian rhythm of sleep was detected starting from the eighth postnatal week. CONCLUSIONS: Our data suggest that in this group of premature infants the circadian maturation of the hypothalamic-pituitary-adrenal axis occurred at the same postnatal age as reported for term infants and that there was a parallelism between the appearance of such rhythm and the onset of sleep rhythm.