Results of the first international round robin for the quantification of urinary and plasma hepcidin assays: need for standardization

The recently discovered iron regulatory peptide hormone hepcidin holds promise as a novel biomarker in iron metabolism disorders. To date, various mass spectrometry and immunochemical methods have been developed for its quantification in plasma and urine. Differences in methodology and analytical performance hinder the comparability of data. As a first step towards method harmonization, several hepcidin assays were compared. Worldwide eight laboratories participated in a urinary and plasma round robin in which hepcidin was analyzed. For both urine and plasma: (i) the absolute hepcidin concentrations differed widely between methods, (ii) the between-sample variation and the analytical variation of the methods are similar. Importantly, the analytical variation as percentage of the total variance is low for all methods, indicating their suitability to distinguish hepcidin levels of different samples. Spearman correlations between methods were generally high. The round robin results inform the scientific and medical community on the status and agreement of the current hepcidin methods. Ongoing initiatives should facilitate standardization by exchanging calibrators and representative samples.     

Immunolocalization of cystinosin,the protein defective in cystinosis

Cystinosis is an autosomal recessive disorder associated with excessive lysosomal cystine accumulation secondary to defective lysosomal cystine efflux. CTNS, the gene mutated in cystinosis, codes for the lysosomal membrane protein cystinosin. Antisera were raised in rabbits to a carboxy-terminal oligopeptide sequence from cystinosin. Antisera were screened by Western blotting and immunocytochemical analyses of transfected COS-7 cells expressing either human wild-type cystinosin, a wild-type cystinosin-green fluorescent protein (GFP) fusion protein, or a fusion protein of GFP and mutant human cystinosin with a carboxy-terminal deletion. In Western blots, bands corresponding to cystinosin or cystinosin-GFP were observed in transfected cells but no signal was detected in cells expressing the carboxy-terminal mutant; preimmune sera yielded negative results in all three cases. In transfected cells expressing wild-type cystinosin, immunoreactivity appeared in subcellular vesicles. In cells expressing the wild-type cystinosin-GFP fusion protein, immunoreactivity colocalized with GFP fluorescence. Previous studies demonstrated that GFP fluorescence from this construct colocalized with immunostaining for a known lysosomal membrane protein, i.e., lysosome-associated membrane protein 2. In immunohistochemical analyses, cystinosin localized to tubule epithelia in three normal human kidneys, with a pattern similar to that of lysosome-associated membrane protein 2; cystinosin immunoreactivity was absent in kidneys from patients with a CTNS deletion. For the first time, antisera have been raised that localize cystinosin in cells in vitro and in vivo.     

Prevalence of Factor V Leiden and other thrombophilic traits among Cretan children with malignancy

BACKGROUND: The prevalence of thrombophilic traits, which might further enhance the risk of thrombotic complications in children treated for cancer, varies significantly among different populations. OBJECTIVE: To evaluate the prevalence of common thrombophilic traits of the East Mediterranean Region, among native Cretan children treated for malignancy. METHODS: Blood samples were consecutively collected from 31 native Cretan children treated for acute lymphoblastic leukaemia (n = 19) or other malignancies (n = 12) over 3 years. A molecular diagnosis based on the presence of Factor V Leiden (FVL), as well as on PT G20210A and MTHFR C677T mutation (in 14 patients) using PCR was applied. Patients who had central venous catheters (n = 29) were treated with an intensified thromboprophylaxis protocol that had been previously established in our institution. RESULTS: The prevalence of the FVL mutation was 19.4% (95% CI = 5-32). The allele frequency is estimated at 11.3% (95% CI: 3.5-19.1) which is higher than that reported for the population of the mainland of Greece. The prevalence of the PT G20210A and MTHFR C677T mutation was 14.3 and 71.4%, respectively (corresponding allele frequencies 7.1 and 50%, respectively). Only one patient developed thrombosis, having although no thrombophilic trait. CONCLUSIONS: Thrombophilic traits were relatively common in this group of native Cretan children treated for malignancy. Thromboprophylaxis should be considered in Cretan children in the presence of known acquired risk factors for thrombosis, but a larger prospective to study is first needed.     

Recovery during high-intensity intermittent anaerobic exercise in boys, teens, and men

PURPOSE: This study examined the effects of age on recovery of peak torque of knee extensors (PTEX) and flexors (PTFL), and total work (TW) during high-intensity intermittent 30-s (HI30) and 60-s (HI60) exercise in boys (N=19; age, 11.4+/-0.5 yr), teens (N=17; age, 14.7+/-0.4 yr), and men (N=18; age, 24.1+/-2.0 yr). METHODS: Each age group's subjects were subdivided to participate in an HI30 or an HI60 protocol. The HI30 involved 4x18 maximal knee extensions and flexions (1-min rest between sets), and the HI60 comprised of 2x34 reps (2-min rest). PTEX (N.m.kg), PTFL (N.m.kg), and TW (J.kg) were recorded at each set. The percent recovery of PTEX, PTFL, and TW was calculated as percent of the value achieved in the first set. RESULTS: In HI60, the percent recovery for PTEX, PTFL, and TW after the first set was higher in boys compared with teens and men (P<0.01). In HI30, the percent recovery for PTEX, PTFL, and TW was higher in boys compared with men in all sets (P<0.01), and in teens compared with men in the last two sets (P<0.05). The percent recovery of PTFL and TW was higher in boys compared with teens in the last two sets (P<0.05). Lactate increase was most pronounced in men, less pronounced in teens, and least pronounced in boys (P<0.01). Heart rate recovered faster in boys compared with teens and men in both protocols (P<0.05). CONCLUSIONS: The recovery was faster in boys than in teens and men during HI30 and HI60, as evident by the greater percent recovery in boys for a given time. Furthermore, it appears that the rate of recovery during HI30 and HI60 anaerobic exercise is maturity dependent.     

Dose reduction in maxillofacial imaging using low dose Cone Beam CT

OBJECTIVES: (a) To measure the absorbed dose at certain anatomical sites of a RANDO phantom and to estimate the effective dose in radiographic imaging of the jaws using low dose Cone Beam computed tomography (CBCT) and (b) to compare the absorbed and the effective doses between thyroid and cervical spine shielding and non-shielding techniques. STUDY DESIGN: Thermoluminescent dosimeters (TLD-100) were placed at 14 sites in a RANDO phantom, using a Cone Beam CT device (Newtom, Model QR-DVT 9000, Verona, Italy). Dosimetry was carried out applying two techniques: in the first, there was no shielding device used while in the second one, a shielding device (EUREKA!, TRIX) was applied for protection of the thyroid gland and the cervical spine. Effective dose was estimated according to ICRP(60) report (E(ICRP)). An additional estimation of the effective dose was accomplished including the doses of the salivary glands (E(SAL)). A Wilcoxon Signed Ranks Test was used for statistical analysis. RESULTS: In the non-shielding technique the absorbed doses ranged from 0.16 to 1.67 mGy, while 0.32 and 1.28 mGy were the doses to the thyroid and the cervical spine, respectively. The effective dose, E(ICRP), was 0.035 mSv and the E(SAL) was 0.064 mSv. In the shielding technique, the absorbed doses ranged from 0.09 to 1.64 mGy, while 0.18 and 0.95 mGy were the respective values for the thyroid and the cervical spine. The effective dose, E(ICRP), was 0.023 mSv and E(SAL) was 0.052 mSv. CONCLUSIONS: The use of CBCT for maxillofacial imaging results in a reduced absorbed and effective dose. The use of lead shielding leads to a further reduction of the absorbed doses of thyroid and cervical spine, as well as the effective dose.     

Intracellular and plasma cytokine profile in neonates born to non-atopic parents: the impact of breast feeding

The aim of this study was to profile the changes in intracellular and plasma cytokines during the neonatal period and evaluate the impact of breast feeding on these parameters. For this purpose, we measured the interleukin (IL)-2 and IL-4 producing CD3+/CD69+ T-cells using flow cytometry and plasma concentrations of interferon (IFN)-gamma and IL-4 using ELISA, in 122 healthy term neonates, aged 6–12 h, born to non-atopic parents, and 25 healthy children aged 1–12 years. A total of 42/122 neonates exclusively breast-fed (BF) and 39/122 formula fed (FF) were studied again on the 30th day of life for the above parameters. Finally, a clinical evaluation for the presence of atopic disease was conducted at 2 years of age. We found that at birth, the percentage of CD3+/CD69+/IL-4+ T-cells (median = 15.8%, range = 4.4%–49%) and plasma concentrations of IL-4 (median = 0.22 pg/ml, range = 0.18–0.25 pg/ml) were significantly higher (P<0.0001) compared to those of children (median = 1.6%, range = 0.16%–2.7% for CD3+/CD69+/IL-4+ and median = 0.17 pg/ml, range = 0.13–0.26 pg/ml for IL-4), whereas plasma concentrations of IFN-gamma were significantly lower in neonates (median = 0.42 pg/ml, range = 0.3–1.5 pg/ml) than in children (median = 1.2 pg/ml, range = 0.3–2.6 pg/ml, P<0.0001). During the neonatal period, only the CD3+/CD69+/IL-4+ T-cells increased significantly in both BF and FF groups. Comparison between BF and FF groups revealed no significant difference in any of the parameters measured. Moreover, no difference in the development of atopy during the first 2 years of life was found between BF and FF infants. Conclusion: our findings demonstrate that during the entire neonatal period type 2 immunity dominates, regardless of the mode of feeding, whereas type 1 immunity dominates during childhood. Moreover, in the absence of family history of atopy, the mode of feeding per se does not play a crucial role in the development of atopy within the first 2 years of life.Abbreviations BF breast-fed - BFA brefeldin A - FITC fluorescein isothiocyanate - FF Formula-fed - IFN interferon - IL interleukin - MoAbs monoclonal antibodies - PBS-BSA phosphate buffered saline bovine serum albumin - PE-Cy 5 phycoerythrin-cyanin 5 - PMA phorbol 12-myristate 13-acetate - Tc T-cytotoxic - Th T-helper     

Rare manifestations of sirenomelia syndrome: a report of five cases

Five cases of sirenomelia presented with rare manifestations are discussed. Three neonates were born alive and died within 2 to 12 hours after birth. One case was the offspring of a triple in vitro fertilization pregnancy with history of early intrauterine death of one of the triplets. The main features included fusion of lower extremities (five of five), renal agenesis (three of five), polycystic renal dysplasia (two of five), anal atresia with large bowel hypoplasia (three of five), pulmonary hypoplasia (four of five), and single umbilical artery (five of five). Other features that have only rarely been associated with sirenomelia included concurrence of congenital heart disease and neuroblastoma, gallbladder agenesis, and upper extremity defects.     

Prospective assessment of emotional distress, cognitive function, and quality of life in patients with cancer treated with chemotherapy

BACKGROUND: The current study sought to delineate prospectively the rates and clinical course of emotional distress, cognitive impairment, and quality of life (QOL) in chemotherapy-naive patients with cancer and to consider the determinants of global QOL. METHODS: Patients who consented to participate were administered the European Organization for Research and Treatment of Cancer QLQ-C30 questionnaire, the Mini-Mental State Examination (MMSE), and the Hospital Anxiety and Depression Scale before and at the end of treatment (EOT). RESULTS: Of the 102 patients initially assessed, 80 (78.4%) completed the study. Most aspects of QOL did not change considerably over time. At EOT, patients reported only significant increases in fatigue and significant decreases in sleep disturbance. Although no significant changes emerged in the rates of anxiety or depression throughout chemotherapy, nearly one-third of the patients experienced severe emotional distress at both points in time. In addition, the authors observed neither significant alteration in the cognitive performance over time nor reliable associations between scores on the MMSE and subjective cognitive function, emotional distress, or QOL. Finally, depression proved to be the leading predictor of global QOL at baseline and at EOT. CONCLUSIONS: The results indicated that a significant proportion of Greek patients with cancer experienced intense anxiety and depression throughout chemotherapy and confirmed the importance of depression as a strong predictor of global QOL. Routine screening of emotional distress across all phases of cancer is mandatory because it will contribute to the identification of patients who are in need of pharmaceutical and/or psychologic intervention.     

Delivery outcomes following loop electrosurgical excision procedure for microinvasive (FIGO stage IA1) cervical cancer

OBJECTIVE: The goal of this study was to report the delivery outcomes in women who had loop electrosurgical excision procedure (LEEP) for microinvasive cervical cancer (stage IA1 without lymphovascular invasion) and became pregnant and progressed beyond 24 weeks. METHODS: A case-control study was performed. Twenty-eight women who were managed exclusively with LEEP for microinvasive cervical carcinoma had at least one pregnancy beyond 24 weeks and were the cases. Each case was matched with one woman who delivered at the same department without prior treatment of her cervix (controls) and their delivery outcomes were compared with those of the cases. Known risk factors for preterm delivery were used as matching factors. RESULTS: There was no statistically significant difference (P > 0.05) between cases and controls in the duration of pregnancy (37.6-38.4 weeks respectively), birth weight (3212-3315 g), cesarean section rate (17.8-32.1%), neonatal unit admission rate (21.4-10.7%), and precipitate labor rate (13-10.5%). The duration of labor was significantly shorter in cases (5.5-7.1 h, P = 0.032). After LEEP the relative risk for preterm delivery is 3.67 (95% confidence interval, 0.97-20.27), for low birth weight infant 0.67 (0.06-5.8), for precipitate labor 1 (0.05-3.88), and for delivery by cesarean section 0.5 (0.17-4.46). CONCLUSIONS: Women treated for microinvasive cancer with LEEP did not have significantly more delivery complications compared with controls apart from shorter duration of labor. There was a possible non-statistically significant trend toward shorter duration of pregnancy in cases. While caution should be advised when selecting and treating women with microinvasive carcinoma by LEEP, the apparent safety of the management and the satisfactory delivery outcome seem to justify this approach in many cases.