A re-investigation of saliva collection procedures that highlights the risk of potential positive interference in cortisol immunoassay

Background

The use of saliva for measurement of cortisol permits non-invasive study of adrenal function, but collection can be technically difficult, particularly in small infants. Saliva collection can be assisted by citric acid to increase saliva flow, or by the use of cotton or polyester swabs in the mouth.

Aim

To determine whether different methods of saliva collection affect cortisol radioimmunoassay (RIA) performance.

Experimental

Cortisol was measured in saliva collected from 16 adults using intra-oral cotton swabs or polyester swabs, compared with saliva dribbled directly into a pot either alone (plain saliva) or after citric acid had been placed on the tongue. An in-house RIA, without prior extraction, was used to measure cortisol with an encapsulated sheep antibody.

Results

Mean (median) salivary cortisol was 10.9 (10.5) nmol L⁻¹ in plain saliva, 10.4 (8.4) nmol L⁻¹ in citric acid stimulated saliva; 25.3 (25.1) nmol L⁻¹ in saliva collected on cotton swabs, and 27.9 (27.3) nmol L⁻¹ collected on polyester swabs. Cortisol in saliva collected using citric acid was not significantly different from plain saliva (p = 0.997), but cortisol in saliva collected using cotton and polyester swabs was significantly higher than that of plain saliva (p < 0.01).

Conclusion

The use of cotton or polyester swabs for collection of saliva can result in spuriously high levels of cortisol when measured by RIA.     

Cortisol release and heart rate variability in horses during road transport

Based on plasma cortisol concentrations it is widely accepted that transport is stressful to horses. So far, cortisol release during transport has not been evaluated in depth by non-invasive techniques such as analysis of salivary cortisol and faecal cortisol metabolites. Transport also causes changes in heart rate and heart rate variability (HRV). In this study, salivary cortisol, faecal cortisol metabolites, heart rate and HRV in horses transported by road for short (one and 3.5 h) and medium duration (8 h) were determined. With the onset of transport, salivary cortisol increased immediately but highest concentrations were measured towards the end of transport (4.1 ± 1.6, 4.5 ± 2.6, 6.5 ± 1.8 ng/ml in horses transported for one, 3.5 and 8 h, respectively). Faecal cortisol metabolite concentrations did not change during transport, but 1 day after transport for 3.5 and 8 h had increased significantly (p < 0.01), reflecting intestinal passage time. Compared to salivary cortisol, changes in faecal cortisol metabolites were less pronounced. Heart rate increased and beat-to-beat (RR) interval decreased (p < 0.05) with the onset of transport. Standard deviation of heart rate increased while root mean square of successive RR differences (RMSSD) decreased in horses transported for 3.5 (from 74 ± 5 to 45 ± 6 ms) and 8 h (from 89.7 ± 7 to 59 ± 7 ms), indicating a reduction in vagal tone. In conclusion, transport of horses over short and medium distances leads to increased cortisol release and changes in heart rate and HRV indicative of stress. The degree of these changes is related to the duration of transport. Salivary cortisol is a sensitive parameter to detect transient changes in cortisol release.     

Urinary free cortisone, but not cortisol, is associated with urine volume in severe obesity

Background

High urine volume enhances urinary free cortisol (UFF) and cortisone (UFE) excretion rates in normal-weight adults and children. Renal excretion rates of glucocorticoids (GC) and their metabolites are frequently altered in obesity. The aim of the present study was to investigate whether UFF and UFE excretion is also affected by urine volume in severely obese subjects.

Experimental

In 24-h urine samples of 59 extremely obese subjects (mean BMI 45.3 ± 8.9 kg/m²) and 20 healthy lean subjects (BMI 22.1 ± 1.8 kg/m²), UFF and UFE, tetrahydrocortisol (THF), 5α-tetrahydrocortisol (5α-THF), and tetrahydrocortisone (THE) were quantified by RIA. The sum of THF, 5α-THF, and THE (GC3), the three major GC metabolites, reflects daily cortisol secretion. 11β-Hydroxysteroid dehydrogenase type 2 (11β-HSD2) activity was assessed by the ratio UFE/UFF. Daily GC excretion rates were corrected for urine creatinine and adjusted for gender and body weight.

Results

In extremely obese subjects, urine volume was significantly associated with creatinine-corrected UFE and 11β-HSD2 activity after adjustment for gender and BMI (r = 0.47, p = 0.0002 and r = 0.31, p = 0.02, respectively). However, urine volume was not associated with creatinine-corrected UFF and GC3 (p = 0.4 and p = 0.6, respectively). In lean controls, urine volume was significantly associated with creatinine-corrected UFE and UFF (r = 0.58, p = 0.01 and r = 0.55, p = 0.02, respectively), whereas urine volume was not associated with 11β-HSD2 activity after appropriate adjustment (p = 0.3).

Conclusions

In severe obesity, in contrast to normal weight, renal excretion of UFE, but not of UFF is affected by fluid intake. This discrepancy may be due to the increased renal 11β-HSD2 activity in obesity.     

Dynamics of endogenous glucocorticoid secretion and its metabolism in Kawasaki disease

Objective

Kawasaki disease (KD) is a severe inflammatory disease that occurs in childhood. Recently, the initial corticosteroid therapy for KD has been reconsidered because its efficacy is controversial. The aim of this study was to evaluate the dynamic change in endogenous glucocorticoid levels and their relation with 11beta-hydroxysteroid dehydrogenase (11β-HSD) activity in the acute phase of KD.

Study design

Sixteen KD patients were investigated. Cortisol and cortisone levels, the cortisol/cortisone ratio and C-reactive protein (CRP) levels were measured on admission, before the first intravenous immunoglobulin (IVIG) therapy and convalescence.

Results

The 16 patients were divided into two groups. Group A included patients who received the first IVIG on admission and blood samples were collected before the first IVIG and convalescence. Group B included patients whose blood samples were collected at three different time points (on admission, before the first IVIG, and convalescence). CRP and cortisol levels and the cortisol/cortisol ratio were markedly higher before the first IVIG than those of convalescence in all patients except in one patient. In Group B patients, both serum cortisol levels and the cortisol/cortisone ratio on admission were significantly increased compared with those before the first IVIG (cortisol: p < 0.005, cortisol/cortisone: p < 0.001).

Conclusions

Decreases in cortisol levels and the cortisol/cortisone ratio before the first IVIG may be explained by a reduction in adrenal secretion and/or local glucocorticoid action through 11β-HSD activity. These findings suggest that exogenous glucocorticoid treatment in combination with the first IVIG at the acute stage may play a synergetic role in KD.     

Neurohormonal and cytokine fluctuations following transcatheter closure for an atrial septal defect

Introduction

Inflammation and neurohormonal activation are considered to be involved in the development of earlier and/or later complications in congenital heart disease patients, even after a successful repair of the lesion. It is not yet clarified what is the role of the therapeutic interventions in the occurrence of such a response and how it could be associated with possible postoperative complications.

Aim

We sought to assess the inflammatory and neurohormonal response to transcatheter closure of secundum type atrial septal defects (ASD) over a six-month follow-up period. We also evaluated the association between the respective markers and catheterization data as well as echocardiographic measurements.

Methods

Plasma concentrations of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), N-terminal-proatrial natriuretic peptide (NT-proANP) and N-terminal-probrain natriuretic peptide (NT-proBNP) were assessed and echocardiographic measurements were performed in twenty-eight patients with atrial septal defect prior to, and at the first, second and sixth months post transcatheter closure. Thirty-three age-matched healthy volunteers were also enrolled.

Results

IL-6 plasma levels, although higher preoperatively, [physical logarithm (ln) IL-6: 3.37 ± 0.66 vs 2.92 ± 0.44 pg/ml, p = 0.015], reached control levels postoperatively, at the end of the third month, whereas TNF-α and IL-10 were not influenced by the procedure. NT-proANP levels were elevated preoperatively compared to the control group (ln NT-proANP 3.78 ± 0.572 vs 3.48 ± 0.30, p = 0.031), with a further significant increase during the 1st month (ln NT-proANP 3.78 ± 0.572 vs 4.2 ± 0.42, p = 0.006), following the pattern of the left atrial volume enlargement, and remained high even 6 months after the procedure .On the other hand, the initially normal concentrations of NT-proBNP, after a transient significant increase during the first month postoperatively (ln NT-proBNP 3.56 ± 0.94 vs 4.58 ± 0.91, p < 0.0001) returned to the controls’ levels at the end of the third month. Preoperative concentrations of NT-proANP positively correlated with NT-proBNP concentrations and pulmonary to systemic flow ratio (Qp/Qs).

Conclusions

Transcatheter closure could improve, on a mid- term basis, the inflammatory process but natriuretic peptides’ secretion continues in parallel with left atrial volume increase. Further follow up is required to determine the long-term progress of the inflammatory and neurohormonal response to the procedure.     

Clinical significance of plasma CD146 and P-selectin in patients with type 2 diabetic nephropathy

Objective

To investigate the levels of plasma CD146 and P-selectin in patients with type 2 diabetic nephropathy at different stages.

Methods

A total of 80 patients with type 2 diabetes mellitus were enrolled in the present study. According to 24 h urinary albumin excretion ratio and renal function, they were further divided into group of diabetes without microalbuminuria (DN0, n = 20), microalbuminuria group (DN1, n = 20), macroalbuminuria group (DN2, n = 20) and renal insufficiency group (DN3, n = 20). Another 20 healthy subjects were enrolled as control group (non-DM). Plasma CD146 and P-selectin were measured by ELISA.

Results

Plasma CD146 and P-selectin were significantly increased in patients with type 2 diabetes with microalbuminuria (DN1) compared with health control (CD146: 415.3 ± 29.0 vs. 243.5 ± 14.7 ng/ml, P < 0.05; P-selectin: 66.8 ± 3.4 vs. 45.3 ± 2.7 ng/ml, P < 0.001). With the development of diabetic nephropathy, both plasma CD146 and P-selectin level progressively rise, with the highest levels in patients with significant renal insufficiency (DN3: 515.9 ± 36.9 and 81.5 ± 5.1 ng/ml respectively, P < 0.001). Moreover, the increase in CD146 is positively co-related to the rise of P-selectin in patients with type 2 diabetes.

Conclusion

Expression of CD146 and P-selectin in patients with type 2 diabetes is elevated, and they are positively correlated with severity of diabetic nephropathy.     

WT1 CpG islands methylation in human lung cancer: A pilot study

Background

CpG island hypermethylation of gene promoters and regulatory regions is a well-known mechanism of epigenetic silencing of tumor suppressors and is directly linked to carcinogenesis. Wilm’s tumor gene (WT1) is a tumor suppressor protein involved in the regulation of human cell growth and differentiation and a modulator of oncogenic K Ras signaling in lung cancer. Changes in the pattern of methylation of the WT1 gene have not yet been studied in detail in human lung cancer. In this study we compared the methylation profile of WT1 gene in samples of neoplastic and non-neoplastic lung tissue taken from the same patients.

Methods

DNA was extracted from neoplastic and normal lung tissue obtained from 16 patients with non small cell lung cancer (NSCLC). The methylation status of 29 CpG islands in the 5′ region of WT1 was determined by pyrosequencing. Statistical analysis was carried out by T test and Mann Whitney test.

Results

The mean percentage of methylation, considering all CpG islands of WT1 in the neoplastic tissues of the 16 NSCLC patients, was 16.2 ± 3.4, whereas in the normal lung tissue from the same patients it was 5.6 ± 1.7 (p < 0.001). Adenocarcinomas presented higher methylation levels than squamous cell carcinomas (p < 0,001).

Conclusions

Methylation of WT1 gene is significantly increased in NSCLC. Both histotype and exposure to cigarette smoke heavily influence the pattern of CpG islands which undergo hypermethylation.     

Detection of multiple cytokines in the urine of patients with focal necrotising glomerulonephritis may predict short and long term outcome of renal function

Background

Detection of urinary cytokines in pauci-immune focal segmental necrotizing glomerulonephritis (FSNGN) may provide valuable information about disease pathogenesis and prognosis.

Methods

Epidermal growth factor (EGF), transforming growth factor (TGF-β1) and vascular endothelial growth factor (VEGF) were measured by ELISA, and Interleukins, monocyte chemotactic protein-1 (MCP-1), macrophage inflammatory protein (MIP1β) by a multiplex cytokine assay, in 38 patients with FSNGN. Their levels were correlated with severity of histological findings and renal function outcome in short and long term.

Results

The percentage of crescents in renal biopsy had positive correlation with TGF-β1 (p = 0.004) and IL-15 urinary excretion (p = 0.01), and negative correlation with EGF (p = 0.01). Increased urinary excretion of IL-6, IL-15, VEGF and MIP-1β was associated with poor renal function outcome, but increased levels of EGF, IL-2 and IL-9 predicted a favourable prognosis. In multiple regression analysis IL-6 and VEGF urinary levels were independent predictors of no-response at the acute phase (p = 0.001 and p < 0.0001, respectively), while, IL-6 was the only factor (p = 0.03) predicted worse outcome at the end of follow-up (39.4 ± 45 months).

Conclusion

Increased urinary excretion of IL-6, IL-15, VEGF, TGF-β1, MCP-1 and MIP-1β and reduced EGF, IL-2, IL-9 may be associated with histological damage and influence response to treatment in pauci-immune FSNGN.     

Chronic angiotensin-(1-7) administration improves vascular remodeling after angioplasty through the regulation of the TGF-β/Smad signaling pathway in rabbits

Objective

Angiotensin-(1-7) [ANG-(1-7)] has been reported to attenuate neointimal formation after vascular injury and stent implantation in rats, but the mechanism remains mostly unresolved. Interestingly, the levels of circulating transforming growth factor-beta1 (TGF-β1) after myocardial infarction were suppressed by ANG-(1-7), which suggests a possible downstream target for the anti-remodeling action of ANG-(1-7). Our study focused on the effects of ANG-(1-7) on vascular remodeling, including neointimal formation and collagen synthesis, and determining whether or not these effects were dependent upon the TGF-β signaling pathway.

Methods

Thirty-two New Zealand white rabbits underwent sham surgery or angioplasty in abdominal aorta. The animals were divided into four groups, which were sham, control, ANG-(1-7), and ANG-(1-7) + A-779. Subsequently, an osmotic minipump was implanted to deliver saline, ANG-(1-7) (576 μg kg⁻¹ d⁻¹) or ANG-(1-7) + A-779 (576 μg kg⁻¹ d⁻¹) for 4 weeks.

Results

The ANG-(1-7) group displayed a significant reduction in neointimal thickness (207.51 ± 16.70 μm vs. 448.08 ± 15.30 μm, P < 0.001), neointimal area (0.266 ± 0.009 mm² vs. 0.408 ± 0.002 mm², P < 0.001), and restenosis rate (28.13 ± 2.74% vs. 40.13 ± 2.74%, P < 0.001) when compared to the control group. ANG-(1-7) also inhibited collagen synthesis by significantly decreasing the mRNA expression of Collagen I and Collagen III (vs. Control group: 0.2190 ± 0.0036 vs. 0.3852 ± 0.0212, P < 0.001 and 1.1328 ± 0.0554 vs. 1.7378 ± 0.1164, P < 0.001, respectively). Furthermore, the expression of TGF-β1 and phosphor-Smad2 (p-Smad2) were significantly suppressed by ANG-(1-7) (vs. Control group: 1.21 ± 0.07 vs. 1.54 ± 0.08, P < 0.001 and 0.31 ± 0.01 vs. 0.43 ± 0.02, P < 0.001, respectively), but no effect on p38 phosphorylation was observed. [d-Ala⁷]-ANG-(1-7) (A-779), showed a tendency to attenuate the anti-remodeling effects of ANG-(1-7).

Conclusion

ANG-(1-7) decreases the amount of vascular remodeling, including a reduction in neointimal formation and collagen synthesis, after angioplasty in rabbits. The responsible mechanism may function through the possible down-regulation of TGF-β1 levels and inhibition of the Smad2 pathway.     

Overexpression of apolipoprotein-B improves cardiac function and increases survival in mice with myocardial infarction

Background

The heart produces apolipoprotein-B containing lipoproteins (apoB) whose function is not well understood. The aim of this study was to evaluate importance of myocardial apoB for cardiac function, structure and survival in myocardial infarction (MI) and heart failure (HF).

Methods and results

MI was induced in mice (n = 137) and myocardial apoB content was measured at 30 min, 3, 6, 24, 48, 120 h and 8 weeks post-MI. Transgenic mice overexpressing apoB (n = 27) and genetically matched controls (n = 27) were used to study the effects of myocardial apoB on cardiac function, remodeling, arrhythmias and survival after MI. Echocardiography was performed at rest and stress conditions at baseline, 2, 4 and 6 week post-MI and cumulative survival rate was registered. The myocardial apoB content increased both in the injured and the remote myocardium (p < 0.05) in response to ischemic injury. ApoB mice had 2-fold higher survival rate (p < 0.05) and better systolic function (p < 0.05) post-MI.

Conclusion

Overexpression of apoB in the heart increases survival and improves cardiac function after acute MI. Myocardial apoB may be an important cardioprotective system in settings such as myocardial ischemia and HF.     

Características de la fractura de cadera y posterior recuperación en pacientes mayores de 65 años con historia de caídas recurrentes

Recurrent falls affect between 14.8% and 19% of the elderly population, and are associated with an increased risk of fracture. We know little about the influence the history of recurrent falls may have on recovery after hip fracture.

Methods

Cohort study. The patients included were, over 65 years admitted during a 1 year period to the General University Hospital of Albacete with a hip fracture due to a fall. Recurrent falls were defined as a history of two or more falls within the 6 months prior to the fracture. Variables: demographic data, circumstances of fall, number of falls in the previous 6 months, type of fracture and its repair, comorbidity and drug treatment, cognitive status at admission (Pfeiffer test) and independence for activities of daily living (Barthel Index - BI) were collected. A subsample of patients with pre-fracture BI≥60 and Pfeiffer at admission≤4 was followed up at 3, 6 and 12 months.

Results

A total of 335 patients were admitted. Data were collected on 279 of them, 19.4% of whom had previously suffered two or more falls. The recurrent fallers had a worse mental status on admission, a higher number of associated diseases, a lower percentage of independence in dressing and in bed-chair transferring than patients without history of recurrent falls, all statistically significant. In the 201 patients followed up, the impairment on the BI after 12 months compared to the BI previous to fracture was higher in recurrent fallers (-20.8 ± 31.54 vs -10.73 ± 20.21, P = .04), focusing more on independence in eating (76% vs 91.9%, P < .05), grooming (72% vs 91,9%, P < .01), faecal continence (60% vs 78.7%, p < .05) and walking indoors (80% vs 93.3%, P < .05).

Conclusions

The recovery of independence after hip fracture is significantly lower in the group of recurrent fallers in patients without moderate or severe functional impairment previous to fracture and cognitively stable.     

The relative contribution of environmental and genetic factors to phenotypic variation in familial Mediterranean fever (FMF)

Introduction

Familial Mediterranean fever (FMF) is an autosomal recessive disease, caused by mutations in the FMF gene MEFV (MEditerranean FeVer). It has a large phenotypic diversity even in patients with similar genotypes. Despite evidence that environmental factors (EFs) and genetic factors, including MEFV mutations (such as M694V, E148Q) and background modifier genes (MGs), affect the clinical manifestations of FMF, the relative contribution of each remains unknown.

Methods

To investigate the relative contribution of environmental and genetic factors to the phenotype of FMF, we compared the intra-pair clinical concordance of 10 mono and 7 dizygotic twins with FMF. The part played by EFs was determined by the phenotypic discordance of the monozygous twins, and the MGs effect was determined by deducing the environmental effect, computed for MZ twins, from the phenotypic discordance of the dizygous twins.

Results

The mean ± SD of intra-pair concordance was higher in the MZ than in DZ twin group (88.1 ± 13.2 vs. 70.7 ± 14.1 respectively, P value < 0.05). Based on the concordance in clinical manifestations in MZ and DZ twins, the environmental effect on the phenotype of FMF is estimated as 11.9% ± 6.6% and the MGs effect as 17.4% ± 15.5% in average.

Conclusions

In FMF the phenotype is affected by MEFV mutations, MGs and EFs in an estimated ratio of about 6:1.5:1 respectively.     

Fraxinus excelsior seed extract FraxiPure™ limits weight gains and hyperglycemia in high-fat diet-induced obese mice

Purpose

The aim of this study was to determine whether a Fraxinus excelsior L. seed extract, FraxiPure™ (0.5% in the diet), limits weight gain and hyperglycemia in mice. In a previous report, we identified several secoiridoids in FraxiPure™, some of which activated peroxisome proliferator-activated receptor alpha (PPARα) in vitro and inhibited the differentiation of 3T3-L1 preadipocyte cells. In a separate study, FraxiPure™ reduced glycemia in healthy volunteers, following an oral glucose tolerance test. These findings suggest that FraxiPure™ has antiobesity and antihyperglycemia effects.

Materials and methods

FraxiPure™ was tested in mice that were fed a high-fat diet over 16 weeks and compared with low-fat and high-fat diet controls. Weight gain, omental and retroperitoneal fat, fasting blood glucose, and fasting blood insulin were measured.

Results

FraxiPure™ reduced gains in body weight by 32.30% (p < 0.05), omental fat by 17.92%, and retroperitoneal fat by 17.78%. FraxiPure™ also lowered fasting blood glucose levels by 76.52% (p < 0.001) and plasma insulin levels by 53.43% (p < 0.05) after 16 weeks. Moreover, FraxiPure™ lowered liver weight gains by 63.62% (p < 0.05) and the incidence of fatty livers by 66.67%.

Conclusions

Our novel results demonstrate the antiobesity effects of chronic administration of an F. excelsior seed extract and confirm its ability to regulate glycemia and insulinemia. In addition, this extract, which is rich in secoiridoid glucosides, protects against obesity-related liver steatosis.     

High plasma CXCL10 levels are associated with HCV-genotype 1, and higher insulin resistance, fibrosis, and HIV viral load in HIV/HCV coinfected patients

Background

CXCL10 may contribute to the host immune response against the hepatitis C virus (HCV), liver disease progression, and response to HCV antiviral therapy. The aim of our study was to analyze the relationship among virological, immunological, and clinical characteristics with plasma CXCL10 levels in human immunodeficiency virus (HIV)/HCV-coinfected patients.

Methods

We carried out a cross-sectional study on 144 patients. CXCL10 and insulin were measured using an immunoassay kit. The degree of insulin resistance was estimated for each patient using the homeostatic model assessment (HOMA) method. Insulin resistance was defined as a HOMA index higher than or equal to 3.8. Aspartate aminotransferase (AST) to platelet ratio (APRI), FIB-4, Forns index, HGM1, and HGM2 were calculated.

Results

The variables associated with log₁₀ CXCL10 levels by univariate analysis were age (b = 0.013; p = 0.023), prior AIDS-defining condition (b = 0.127; p = 0.045), detectable plasma HIV viral load (b = 0.092; p = 0.006), log₁₀ HOMA (b = 0.216; p = 0.002), HCV-genotype 1 (b = 0.114; p = 0.071), and liver fibrosis assessed by all non-invasive indexes (log₁₀ APRI (b = 0.296; p = 0.001), log₁₀ FIB-4 (b = 0.436; p < 0.001), log₁₀ Forns index (b = 0.591; p < 0.001), log₁₀ HGM1 (b = 0.351; p = 0.021), and log₁₀ HGM2 (b = 0.215; p = 0.018)). However, in multivariate analysis, CXCL10 levels were only associated with HOMA, detectable plasma HIV viral load, HCV-genotype 1 and FIB-4 (R-square = 0.235; p < 0.001).

Conclusion

Plasma CXCL10 levels were influenced by several characteristics of patients related to HIV and HCV infections, insulin resistance, and liver fibrosis, indicating that CXCL10 may play an important role in the pathogenesis of both HCV and HIV infections.     

Evaluation impact of long-term usage of inhaled fluticasone propionate on ocular functions in children with asthma

Objective

Although systemic, topical, and periocular corticosteroid administration have long been associated with ocular side effects, there has been little evidence to suggest that long-term inhaled corticosteroids can cause ocular side effects. The aim of this study was to evaluate the effects of long term treatment inhaled fluticasone propionate spray usage the recommended dose on some ocular functions in pediatric patients with asthma.

Methods

The study group consisted of 266 prepubertal children with asthma who had used inhaled fluticasone propionate spray at 3-6 years intermittently. One hundred and sixty children who were newly diagnosed with asthma without any treatment made up the control group. Schirmer test results, central corneal thickness, visual acuity, intraocular pressure, cataract formation, keratometry and tear break-up time compared between study and control groups.

Results

The ages of the 266 study patients (150 male) were between 7 and 11 years. The average age (±SEM) was 8.2 ± 1.7 years, and the mean (±SEM) a daily dose of 323 μg (range 250-450 μg) inhaled fluticasone propionate spray, with 865.2 ± 215 g total steroid use during treatment. Eye functions including cataract formation, corneal ectasia, ocular hypertension or glaucoma, and dry eye were not observed in any of the patients in the study group and were not correlated with total steroid dosage (t = 0.150, p = 0.384).

Conclusion

Our findings suggest that long-term intermittent treatment for 3-6 years with inhaled fluticasone propionate spray, as much as average 320 μg daily, in children with asthma seems to be safe for some eye functions.     

Association between the epidermal growth factor rs4444903 G/G genotype and advanced fibrosis at a young age in chronic hepatitis C

Background

The epidermal growth factor (EGF) rs4444903 A > G polymorphism has been associated with the development of liver cancer, which commonly complicates cirrhosis of viral origin; however, whether this polymorphism might be associated with fibrosis progression in chronic viral hepatitis is unknown. The present study was performed to assess the allelic and genotypic frequencies of the rs4444903 A > G polymorphism in patients with chronic hepatitis C virus HCV infection and to ascertain whether this polymorphism might be an independent predictor of the degree of fibrosis.

Methods

An RFLP-PCR technique was used to genotype 645 patients (211 with cirrhosis); 528 were referred for the diagnosis and treatment of chronic hepatitis C, and 117 were transplanted for HCV-related end stage liver disease. A group of 428 healthy subjects served as a control. All the subjects were of Caucasian ethnicity.

Results

The EGF rs4444903 A > G polymorphism genotype frequencies in HCV chronic infected patients were as follows: A/A = 227 (35.3%), A/G = 328 (50.9%), and G/G = 90 (14.8%). Genotype frequencies were found to differ between patients with an Ishak staging score ? 2 (A/A = 117, A/G = 157, G/G = 34) and patients with a score > 2 (A/A = 110, A/G = 171, G/G = 56, p = 0.038). A highly significant linear relationship between increasing stage scores and EGF genotype was detected in younger patients (A/A: 2.02 ± 0.18, A/G: 2.55 ± 0.17, G/G: 3.00 ± 0.32, p = 0.008). However, no significant association was detected between the stage score and EGF genotype in older patients (A/A: 3.79 ± 0.19, A/G: 3.64 ± 0.15, G/G: 3.98 ± 0.30 p = 0.579).

Conclusions

The EGF rs4444903 A > G polymorphism may facilitate liver fibrosis progression in Caucasian patients with chronic hepatitis C, especially in younger patients.     

Leaf press juice from Bryophyllum pinnatum (Lamarck) Oken induces myometrial relaxation

Aims

The use of preparations from Bryophyllum pinnatum (Lamarck) Oken (Kalanchoe pinnata (Lamarck) Persoon) in tocolysis is supported by clinical evidence. We studied here the effect of B. pinnatum leaf press juice and its chemical fractions on the response of human myometrial strips. No data are available if the influence on myometrial strips of the juice differs from that of its components in the chemical fractions, in order to increase the pharmacological effect.

Methodology

In vitro study to test the effect of repeated addition of B. pinnatum leaf press juice (BPJ) and its chemical components in several dilutions (undiluted, 1-10%) on myometrium strips hang up in a myograph chamber. Chemical analysis is including HPLC, MPLC with Sephadex LH-20 and TLC.

Results

All test solutions are inhibiting contractility by reducing the amplitude and the area under the curve (AUC) of the contractions. Undiluted BPJ and its undiluted chemical fraction 4 are reducing most effective these two parameters: the amplitude was at 78% of the baseline (95% CI (77-89); p < 0.05) at the second addition of the BPJ and at 70% (95% CI (50-90); p < 0.05) of the first addition of fraction 4; the AUC was at 82% (95% CI (69-95); p < 0.05) of the baseline at the first addition of the press juice and at 51% (95% CI (27-74); p < 0.05) at the first addition of fraction 4. The BPJ decreased amplitude and AUC significantly faster and increased frequency significantly faster than the control. Fractions could be tentatively assigned to bufadienolids, flavonoids and cinnamic acids. Fraction 4, accounted for flavonoids, increased the frequency of the contractions most effectively: 557% of the baseline (95% CI (316-797); p < 0.05) at the first addition.

Conclusion

Leaf juice of B. pinnatum and its flavonoid fraction are most effective in relaxing myometrial strips by inducing frequency.     

The effect of zoo visitors on the behaviour and faecal cortisol of the Mexican wolf (Canis lupus baileyi)

The effect of zoo visitors on the behaviour and physiological responses of the Mexican wolf has not been documented but is worthy of investigation since it is a critically endangered native species and most of the population live in zoos. The effect of the number of visitors was assessed in 12 wolves (five males and seven females) in three zoos of central Mexico. Each wolf was observed for a total of 72 h on Saturdays, Sundays, Mondays and Tuesdays (6 h/day) during a three-week period. Scan sampling was used to obtain individual time budgets. A faecal sample was collected on the following morning of each observation day from 10 wolves to determine cortisol level (RIA). A repeated measures analysis of variance revealed that there was an effect of day on the proportion of time lying (P < 0.01), eating (P < 0.01) and in locomotion (P < 0.01). Wolves spent less time lying on Saturdays and Sundays (0.2980 ± 0.012 and 0.3266 ± 0.012, respectively) than on Mondays and Tuesdays (0.3821 ± 0.012 and 0.4075 ± 0.012, respectively), as well as less time eating on weekend days (Saturdays 0.1214 ± 0.015, Sundays 0.0556 ± 0.015) than in the other 2 days (Mondays 0.816 ± 0.015 and Tuesdays 0.0915 ± 0.015). Mean faecal cortisol was 203.7 ng/g of DM and ranged from 113.10 to 314.48 ng/g. There was an effect of day on the faecal cortisol response (P < 0.01) with wolves having higher cortisol response on Sundays (242.50 ng/g ± 8.48) compared to the other 3 days (Saturdays 192.71 ± 8.48, Mondays 170.73 ng/g ± 7.80, Tuesdays 183.82 ng/g ± 8.48). This is the first study that measures faecal cortisol of Mexican wolves. These results indicate that the amount of visitors in zoos influence the behaviour and adrenal activity of these animals which could be undesirable for ex situ conservation efforts of this endangered species.