Malgorzata Klimek

Metal-specific effects on metallothionein gene induction and riboflavin content in coelomocytes of Allolobophora chlorotica.

Metal pollution affects earthworm coelomocytes, including their differential counts, riboflavin content and metallothioneins (MT) involved in metal homoeostasis and detoxification. The present work shows effects of Ni, Cu, Zn, Cd, and Pb at the same molarity (1mM) on coelomocytes of Allolobophora chlorotica after 2-day worm dermal exposure to metal chlorides. Numbers of coelomocytes/eleocytes extruded by electric shock and amounts of riboflavin in coelomocyte lysates were significantly decreased in Cu-exposed worms, less diminished in response to Ni, Zn, Cd, and unaffected by Pb. In sharp contrast, real-time PCR revealed a very strong (272 fold) MT-mRNA induction in response to Cd only. The induction was very low in response to Zn, Cu, Pb, and Ni ions (2.6, 2.1, 1.4, and 1.3-fold, respectively). In conclusion, decreased cell counts and riboflavin content are molecular biomarkers of Cu exposure while induction of MT-mRNA is a molecular biomarker of worm Cd exposure.

Correlation between early neonatal diet and atopic symptoms up to 5-7 years of age in very low birth weight infants: follow-up of randomized, double-blind study

The influence of early feeding on the risk of atopic diseases has been studied in full‐term newborns, not in very low birth weight infants (VLBW). The study evaluated effect of early feeding of VLBW infants with either cow’s milk‐based formula (CMF) or extensively hydrolyzed milk formula (HF) on incidence of atopic diseases and markers of atopy at 5‐7 years of age. This was a follow‐up of the randomized, double‐blind study evaluating the influence of different enteral feeding protocols on the early morbidity of VLBW infants. In the original study 80 children were randomly allocated into 2 groups receiving during first month of life HF (experimental group) or CMF (control group). At the age of 5‐7 years, 62 children among 74 available (84%) with mean birthweight 1124g were evaluated according to standardized ISAAC (International Study of Asthma and Allergies in Childhood) protocol. Total IgE level, specific IgE, lymphocyte CD4+CCR4+/CD4+CXCR3+ ratio and skin prick tests (SPT) were done. Prevalence of obvious allergic diseases was not significantly different between the studied groups (HF: 12/33; CMF: 6/29; RR [relative risk] HF vs CMF: 1.76; 95%CI [confidence interval]: 0.76–4.09). Comparison of atopic status across groups revealed similar rate of positive markers of atopy: IgE (RR: 2.57 95%CI: 0.91–8,08), SPT (RR: 5.13; 95%CI: 0.93–31.6), lymphocyte CD4+CCR4+/CD4+CXCR3+ ratio (OR: 2.32; 95%CI: 0.78–7.53) in the both studied groups. Based on the carried out follow‐up study we were unable to confirm the usefulness of hydrolyzed formula in prevention of allergy in an unselected cohort of very low birth weight infants.

Characteristics of immune-competent amoebocytes non-invasively retrieved from populations of the sentinel earthworm Lumbricus rubellus (Annelida; Oligochaeta; Lumbricidae) inhabiting metal polluted field soils

Lumbricus rubellus is a cosmopolitan earthworm devoid of riboflavin-storing eleocytes; its immune competent coelomocytes are predominantly amoebocytes. Our aim was to determine whether amoebocyte cytometrics in L. rubellus are robust biomarkers for innate immunological responses to environmental pollutants. Investigations were conducted on populations inhabiting three unpolluted and five metalliferous (mainly Pb+Zn+Cd) habitats in the UK and Poland. Inter-population differences in worm mass and amoebocyte numbers did not consistently reflect soil or tissue metal concentrations. Flow cytometry indicated that autofluorescence of the amoebocytes differs between cells from the unpolluted and metal-polluted worms, and pinocytosis of neutral red by amoebocytes was lower (especially at 15 versus 60 min incubation) in worms from the polluted Poland site compared with the reference population. To conclude, amoebocyte cytometrics and functionality are potentially useful for environmental diagnostics; deployment is contingent on better understanding potential confounders.

Nephrectomy or nephron-sparing surgery – how to decide?

Aim of the study Radical nephrectomy in the treatment of renal cell carcinoma (RCC) remains the gold standard, but nephron-sparing surgery (NSS) is still increasing in importance. The main goal of this study was to compare the diagnostic accuracy of ultrasound and multi-detector computed tomography in RCC staging and its influence on deciding about further patient treatment. Material and methods 87 patients (age range 27–90 years; median 61.5) underwent ultrasound (US) scan and contrast-enhanced computed tomography (CE-CT) of the abdomen and pelvis. 28 patients were qualified for NSS. The remaining group of patients underwent nephrectomy. Results NSS was performed more frequently among patients with lesions in the lower pole of the kidney and there was no infiltration to the calyx and renal pelvis. Radical nephrectomy (RN) was pursued in cases with lesions in the central or upper pole. Lesion diameter in patients qualified for NSS was smaller than in patients qualified for radical nephrectomy. Conclusions Determining the relationship between tumour and adjacent structures is not a simple matter. According to our study, 50% of CT results differ from histopathology assessment. Tumour diameter determined in CT examination is larger than in ultrasound and histopathological measurements while US scanning tends to underestimate tumour size in relation to histopathological assessment.

From a regional cohort of extremely low birth weight infants: cardiac function at the age of 7 years.

Background: The long-term impact of prematurity on cardiac structure and function has not yet been fully discovered. Objectives: To assess long-term cardiac complications in the regional cohort of extremely low birth weight (ELBW) children born in 2002-2004. Material and Methods: Eighty-one children born as ELBW infants (91% of the available cohort) with a median birth weight of 890 g (25-75th percentile: 760-950) were evaluated at the mean age of 6.7 years. The control group included 40 children born full-term, selected from one general practice in the district. Echocardiography and 24-hour ambulatory blood pressure measurements (ABPM) were performed. The primary outcome variable was the presence of cardiac complications such as left ventricular hypertrophy (LVH), diastolic dysfunction or systolic dysfunction. Results: LVH was diagnosed in 4/81 ELBW children and 2/40 control children (p = 1.0). Concentric remodeling was detected in 8 (10%) subjects from the ELBW group and in 2 (5%) from the control group (p = 0.49). There were no patients with diastolic or systolic dysfunction in either group. After having expressed the results of M-mode echocardiography as z-scores for body surface area (BSA), statistically significant differences were observed for right-ventricle dimension in diastole (-1.49 ± 1.25 vs. -0.31 ± 0.91; p < 0.001), LV inner dimension in diastole (-0.53 ± 1.26 vs. 0.13 ± 0.94; p = 0.01) and left atrium (-0.93 ± 1.07 vs. -0.15 ± 1.02; p < 0.01). Heart rate (HR) was significantly faster in ELBW children (92.9 ± 8.4 vs. 86.7 ± 7.4 bpm; p = 0.01 adjusted for BSA) and they also had significantly higher night-time blood pressure [mean (z-score): 1.15 vs. 0.2; p = 0.02] without nocturnal dipping (night-time dipping <10%: 13 (16.7%) vs. 2 (5.2%), p = 0.13). Conclusions: No differences were found between the groups in the occurrence of cardiac complications. Ex-preterm ELBW children at age 6 may have a faster HR, smaller cardiac dimensions on echocardiography and higher nocturnal blood pressure. The clinical relevance of these findings is unknown.

Relationship between Proton Magnetic Resonance Spectroscopy of Frontoinsular Gray Matter and Neurodevelopmental Outcomes in Very Low Birth Weight Children at the Age of 4

Very low birth weight is associated with long term neurodevelopmental complications. Macroscopic brain abnormalities in prematurity survivors have been investigated in several studies. However, there is limited data regarding local cerebral metabolic status and neurodevelopmental outcomes. The purpose of this study was to characterize the relationship between proton magnetic resonance spectra in basal ganglia, frontal white matter and frontoinsular gray matter, neurodevelopmental outcomes assessed with the Leiter scale and the Developmental Test of Visual Perception and selected socioeconomic variables in a cohort of very low birth weight children at the age of four. Children were divided in three groups based on the severity of neurodevelopmental impairment. There were no differences in spectroscopy in basal ganglia and frontal white matter between the groups. Lower concentrations of N-acetylaspartate (NAA), choline (Cho) and myoinositol (mI) were observed in the frontoinsular cortex of the left hemisphere in children with neurodevelopmental impairment compared to children with normal neurodevelopmental outcomes. Higher parental education, daycare attendance and breastfeeding after birth were associated with more favorable neurodevelopmental prognosis, whereas rural residence was more prevalent in children with moderate and severe impairment. Our study demonstrates the role of long term neurometabolic disruption in the left frontoinsular cortex and selected socioeconomic variables in determination of neurodevelopmental prognosis in prematurity survivors.

Relationship between Stereoscopic Vision, Visual Perception, and Microstructure Changes of Corpus Callosum and Occipital White Matter in the 4-Year-Old Very Low Birth Weight Children

Aim. To assess the relationship between stereoscopic vision, visual perception, and microstructure of the corpus callosum (CC) and occipital white matter, 61 children born with a mean birth weight of 1024 g (SD 270 g) were subjected to detailed ophthalmologic evaluation, Developmental Test of Visual Perception (DTVP-3), and diffusion tensor imaging (DTI) at the age of 4. Results. Abnormal stereoscopic vision was detected in 16 children. Children with abnormal stereoscopic vision had smaller CC (CC length: 53 ± 6 mm versus 61 ± 4 mm; p < 0.01; estimated CC area: 314 ± 106 mm2 versus 446 ± 79 mm2; p < 0.01) and lower fractional anisotropy (FA) values in CC (FA value of rostrum/genu: 0.7 ± 0.09 versus 0.79 ± 0.07; p < 0.01; FA value of CC body: 0.74 ± 0.13 versus 0.82 ± 0.09; p = 0.03). We found a significant correlation between DTVP-3 scores, CC size, and FA values in rostrum and body. This correlation was unrelated to retinopathy of prematurity. Conclusions. Visual perceptive dysfunction in ex-preterm children without major sequelae of prematurity depends on more subtle changes in the brain microstructure, including CC. Role of interhemispheric connections in visual perception might be more complex than previously anticipated.

Regional lung ventilation pattern in preschool children with bronchopulmonary dysplasia is modified by bronchodilator response

Bronchopulmonary dysplasia (BPD) remains a significant long‐term complication of prematurity. A standardized method of pulmonary function testing is still not available in preschool children with BPD. We investigated the feasibility of Electrical Impedance Segmentography (EIS) monitoring in this group and the impact of bronchodilator response (BDR) to salbutamol on the pattern of lung ventilation.

Physical state of human papillomavirus type 16 in cervical intraepithelial lesions and cancers determined by two different quantitative real-time PCR methods.

The aim of this study was to analyse the correlation between a new multiplex qPCR assay and a reference qPCR assay for assessment of the human papillomavirus (HPV16) load and the viral genome status. The study was performed on 100 HPV16 positive samples containing premalignant lesions and carcinomas. HPV16 E2 and E6 gene loads were assessed by two PCR methods. The load of E2 and E6 was normalized to the cell number by qPCR targeting the RNase P open reading frame. The physical state of the viral genome was determined as a ratio of E2/E6 copies number per cell. Among 100 samples analysed, there were no statistically significant differences in the E2 and E6 viral load evaluated by multiplex qPCR and qPCR, the correlation coefficients were 0.98 and 0.97, respectively. There were 19% of samples with the integrated, 73% with mixed and 8% with episomal state of viral genome detected by multiplex qPCR and 17%, 79%, 4%, respectively, found by qPCR. Prevalence of integrated and episomal forms estimated by multiplex qPCR was higher than the one obtained by qPCR (Chi2, p < 0.0001), but in samples with premalignant and malignant diagnoses no significant differences were demonstrated regardless of the methods used. Sensitivity and specificity of multiplex qPCR were 93.7% and 100% as compared with qPCR, the positive predictive value was 100%. In summary, the multiplex qPCR assay in respect of HPV16 load and the frequency of viral genome status was shown to be a sensitive and specific reference method. Simultaneous estimation of E2 and E6 genes in one reaction tube reduces the cost of testing.

Two Siblings with Phenotypes Mimicking Peroxisomal Disorders but with Discordant Biochemical Findings

The authors present a report on two siblings with a nearly identical phenotype mimicking peroxisomal disorder but with totally discordant biochemical findings. In an attempt to confirm the diagnosis of a peroxisomal disorder, plasma levels of very long chain fatty acids (VLCFA) and phytanic acid were determined. In addition, fibroblasts profile of VLCFA and plasmalogen levels were evaluated for Case 2. Pathomorphological examinations represented by lymph node and kidney biopsies as well as postmortem examinations of liver, adrenals and brain were performed in Case 1. The phenotypes of both siblings showed marked resemblance to those observed in children with Zellweger syndrome. The results of biochemical tests performed in Case 1, together with abundant peroxisomes in the liver and the wide spectrum of clinical abnormalities suggest pseudo-Zellweger or Zellweger-like syndromes, but no conclusive data have been found. The plasmalogen level in fibroblasts as well as the fibroblasts profile of VLCFA noted in Case 2 were within the normal range, thus excluding the possibility of a peroxisomal disorder. The striking phenotype resemblance of the siblings mimicking Zellweger, together with discordant biochemical findings, are difficult to explain on the basis of currently available tests.