A novel magnet-based scratch method for standardisation of wound-healing assays.

A novel magnetic scratch method achieves repeatability, reproducibility and geometric control greater than pipette scratch assays and closely approximating the precision of cell exclusion assays while inducing the cell injury inherently necessary for wound healing assays. The magnetic scratch is affordable, easily implemented and standardisable and thus may contribute toward better comparability of data generated in different studies and laboratories.

Meniscus regeneration combining meniscus and mesenchymal stromal cells in a degradable meniscus implant: an in vitro study.

Meniscus regeneration is an unmet clinical need as damage to the meniscus is common and causes early osteoarthritis. The aim of the present study was to investigate the feasibility of a one-stage cell-based treatment for meniscus regeneration by augmenting a resorbable collagen-based implant with a combination of recycled meniscus cells and mesenchymal stromal cells (MSCs). Cell communication and fate of the different cell types over time in co-culture were evaluated by connexin 43 staining for gap junctions and polymerase chain reaction (PCR) to discriminate between meniscus cells and MSCs, based on a Y-chromosome gene. To define optimal ratios, human meniscus cells and bone-marrow-derived MSCs were cultured in different ratios in cell pellets and type I collagen hydrogels. In addition, cells were seeded on the implant in fibrin glue by static seeding or injection. Cellular communication by gap junctions was shown in co-culture and a decrease in the amount of MSCs over time was demonstrated by PCR. 20 : 80 and 10 : 90 ratios showed significantly highest glycosaminoglycan and collagen content in collagen hydrogels. The same statistical trend was found in pellet cultures. Significantly more cells were present in the injected implant and cell distribution was more homogenous as compared to the statically seeded implant. The study demonstrated the feasibility of a new one-stage cell-based procedure for meniscus regeneration, using 20 % meniscus cells and 80 % MSCs seeded statically on the implant. In addition, the stimulatory effect of MSCs towards meniscus cells was demonstrated by communication through gap junctions.

A distal renal tubular acidosis showing hyperammonemia and hyperlactacidemia.

INTRODUCTION: distal renal tubular acidosis (dRTA) presents itself with variable clinical manifestations and often with late expressions that impact on prognosis. CASE REPORT: A 45-day-old male infant was admitted with stopping growth, difficult feeding and vomiting after meals. Clinical tests and labs revealed a type 1 renal tubular acidosis, even if the first blood tests showed ammonium and lactate increase. We had to exclude metabolic diseases before having a certain diagnosis. CONCLUSIONS: blood and urine investigations and genetic tests are fundamental to formulate dRTA diagnosis and to plan follow-up, according to possible phenotypic expressions of recessive and dominant autosomal forms in patients with dRTA.

Kayser-Fleischer ring in Wilson's disease: a cohort study.

AIMS: To evaluate Wilson's disease (WD) features in Sardinian patients with Kayser-Fleischer (KF) ring and to evaluate correlations between modifications in KF and anti-copper therapy and systemic WD evolution. PATIENTS AND METHODS: Sixty-seven WD patients (35 m/32 f; mean age 41 years) were retrospectively studied. At diagnosis and during follow up comprehensive ophthalmologic and neurologic examinations, brain RMN and ECD SPECT, detailed objective laboratory studies and hepatic histological examination were performed on all patients for analysis. All patients were given anti-copper therapy with d-Penicillamine in mono-therapy or in combination with Zinc Salts. RESULTS: At diagnosis, KF was observed in 27% of patients with equal distribution in all age groups. Significant correlations between KF at diagnosis, neuro-psychiatric manifestations and pathologic features in brain RMN and in brain ECD SPECT were found at diagnosis. During follow up, a decrease in, or regression of KF was seen in 14% of patients. Anti-copper therapy leads to KF regression and prevents the appearance of KF. No significant correlations were observed between KF regression and clinical neurological or neuro-imaging improvement nor between KF modifications and clinical hepatic improvement. CONCLUSIONS: Our study highlights the peculiar features of Sardinian WD patients: low representation of KF, its equal distribution in all age groups, significant correlation between KF at diagnosis and clinical neurological manifestations, pathologic brain RMN and brain ECD-SPECT are highlighted by our study. Anti-copper therapy induces KF regression and prevents its onset. Therefore, KF ring does appear to be a predictive factor in the neurological and hepatic evolution of WD.

A comparison of two blood culture procedures for the isolation of staphylococci in a paediatric intensive care unit.

Blood culture results obtained between January 2000 and July 2003 were reviewed for 1360 patients in a paediatric intensive care unit (PICU). The BacT/Alert FA aerobic medium was used with a blood volume of 1.5 mL for the first 23 months, and the BacT/Alert PF paediatric medium was used with a 0.5-mL volume for the remaining 18 months. The isolation rates were similar during both periods (13.4% vs. 13.1%), and staphylococci were the most common isolates (72.8%). There was a shorter time to detection of staphylococci with the smaller-volume (PF) procedure, which thus seems suitable for use in the diagnosis of staphylococcal bacteraemia in the PICU.

Use of the E test to assess synergy of antibiotic combinations against isolates of Burkholderia cepacia-complex from patients with cystic fibrosis.

Treatment of Burkholderia cepacia-complex infections in cystic fibrosis patients is problematic, since the microorganism is often resistant to most antimicrobial agents. In this study, the Epsilometer test, or E test, was used to assess the activity of antimicrobial combinations against Burkholderia cepacia-complex. In a preliminary evaluation, the E test was compared to the checkerboard method using 10 test organisms. Synergy testing by the E test was then performed on 131 clinical isolates of Burkholderia cepacia-complex using various combinations of antimicrobial agents. Agreement between the E test and the checkerboard method was 90%. The rate of resistance to individual agents ranged from 48% for meropenem to 100% for tobramycin, chloramphenicol, and rifampin. In 71.6%, 15.6%, and 12.6% of the test evaluations performed, the combinations tested resulted in additivity/indifference, synergism, and antagonism, respectively. The highest rates of synergy were observed with combinations of ciprofloxacin-piperacillin (44%), rifampin-ceftazidime (33%), chloramphenicol-ceftazidime (22%), cotrimoxazole-piperacillin/tazobactam (22%), and ciprofloxacin-ceftazidime (21%). Rates of antagonism for cotrimoxazole and chloramphenicol in combination with beta-lactam agents were higher than those observed for ciprofloxacin plus beta-lactam agents. These results suggest that the E test is a valuable and practical method to be considered for improving the identification of possible therapeutic options in cystic fibrosis patients infected with organisms belonging to the Burkholderia cepacia-complex.

[Epidemiology and sensitivity to antibiotics in paediatric respiratory infections over a 4-year period. Retrospective study of 460 H. influenzae, S. pneumoniae and M. catarrhalis strains] / Epidemiologia e sensibilita agli antibiotici in infezioni respiratorie di pazienti pediatrici in un periodo di 4 anni. Studio retrospettivo su 460 ceppi di H. influenzae, S. pneumoniae e M. catarrhalis.

H. influenzae, S. pneumoniae and M. catarrhalis are the most common bacterial pathogens causing respiratory infections in children. Resistance to antibiotics may vary according to the geographical area. It is therefore important that the resistance pattern of such pathogens is determined by surveillance studies carried out both on a national scale and by individual laboratories. In this study, we determined retrospectively the prevalence of H. influenzae, S. pneumoniae and M. catarrhalis in upper respiratory tract infections involving subjects of paediatric age, with reference to the type of clinical sample (pharingeal swab and nasal swab), symptomatology and age group. Moreover, for the above micro-organisms the pattern of sensitivity to antibiotics was assessed. In the observation period (January 1996-December 1999), at the day hospital of the Paediatric Pneumology Division of the Gaslini Institute in Genova, in 476 patients between 0 and 15 years of age a total of 460 respiratory pathogens were isolated: 164 S. pneumoniae strains, 163 of H. influenzae (96 belonging to type B and 67 non-attributable to any type) and 133 of M. catarrhalis. As regards sensitivity to antibiotics, ceftriaxone and amoxycillin/clavulanic acid proved to be the most active molecules in all the studied strains.

Variations of cardiac performance and inotropism in healthy postmenopausal women treated with estroprogestin replacement therapy.

OBJECTIVE: The aim of the present study was to evaluate possible variations in cardiac hemodynamic parameters related to the natural changes of ovarian estrogen production. METHODS: Forty postmenopausal women aged 52.7 +/- 4.6 years, randomized into two groups (20 patients in each group) according to the administration (group A) or not (group B) of estroprogestin replacement therapy (ERT), were examined using thoracic electrical bioimpedence. RESULTS: After 6 months of therapy, we observed the following: (1) the mean end-diastolic index was significantly higher in group A than in group B (70.27 and 57.13 mL/m2, respectively) (p < 0.05); (2) the mean acceleration index, indicator of heart contractility, and the mean cardiac index rate, indicators of cardiac performance, were significantly higher in group A than in group B (mean, 1.35 vs. 0.76 s [p < 0.01] and mean, 3.22 vs. 2.34 L/min/m2 [p < 0.05], respectively); and (3) the patients treated with ERT showed systemic vascular resistance index values significantly lower than the controls (mean, 2280 vs. 3150 fOhm/m2 [p < 0.01]), achieving standard levels after 6 months of therapy. Furthermore, the acceleration index showed a significant increase, within group A, between the third and sixth month of ERT (0.91 vs. 1.35 s [p < 0.05]). CONCLUSIONS: Our findings suggest that postmenopausal women treated with a 6-month course of ERT have significantly improved end-diastolic index, heart contractility index, cardiac index, and systemic vascular resistance, whereas 3 months of ERT does not seem to induce the same effects. In our study, thoracic electrical bioimpedence was shown to be a sensitive and specific method of analysis with a very low cost of administration.

Early detection of the anthracycline-induced cardiotoxicity. A non-invasive haemodynamic study.

Anthracyclines are the most frequent cause of iatrogenic congestive heart failure ranging from acute reversible minor, irreversible reduction in the left ventricular ejection fraction and death despite preventive measures. Sensitive methods are needed to detect earliest preclinical cardiotoxicity along with the development of new protective agents. Thirty breast cancer patients were randomly treated with q 21 120 mg/m2 Epirubicin (EPI) x 3, alone (10 patients), or + ICRF-187 (1000 mg/m2) (10 patients) or + C0Q10 (50 mg/day) (10 patients) and monitored by Thoracic Electrical Bioimpedance (TEB) cardiography before (T0) and at the end of chemotherapy (T1), then at 1, 3, 6 months of follow up (F1, F2, F3). a) The group treated with EPI alone showed, between F1-F2, a significant (p < 0.05) decrease in Stroke Index (S1). Acceleration Index (ACI) and a significant (p < 0.05) increase in Systemic Vascular Resistance Index (SVRI), while between F2 and F3 it showed a significant (p < 0.05) recovery in S1 and ACI. b) The group treated with EPI + ICRF-187 showed, between F1 and F2 a significant decrease in S1 and ACI (p < 0.05, p < 0.01 respectively) and a significant (p < 0.05) increase in SVRI: between F2-F3 ACI had a significant (p < 0.05) recovery: c) The group treated with EPI +C0Q10 showed no modification in Sl, ACI, and SVRI during the study. The ejection Fraction (EF) remained unchanged during the study in all the groups. C0Q10 seems to prevent early decreases in cardiac performance and contractiling, thus avoiding an SVRI increase, while ICRF-187 did not. Since ICRF-187 acts by binding iron, we deem that the earliest cardiac involvement, may occur before iron overload; therefore the role of ICRF-187 and C0Q10 in acute or chronic heart toxicity was correlated with high-dose anthracycline and needs to be further investigated.

Primary shunt hyperbilirubinemia associated with Gilbert's syndrome.

A patient with primary shunt hyperbilirubinemia associated with Gilbert's syndrome is described. The laboratory findings of unconjugated hyperbilirubinemia, mild reticulocytosis, normoblastic erythroid hyperplasia of bone marrow, increased plasma iron turnover, and normal peripheral red blood cell survival were consistent with increased intramedullary hemolysis. Nevertheless, unconjugated bilirubin level exceeding 4 mg/100 mL, in the absence of appreciable anemia and of biochemical evidence of hepatic dysfunction, suggested the coexistence of defective hepatic bilirubin clearance. The mode of inheritance and the effect of fasting and phenobarbital on bilirubin concentrations were indicative of associated Gilbert's syndrome.

Glucose-6-phosphate dehydrogenase deficiency: genetic heterogeneity in Sardinia.

Glucose-6-phosphate dehydrogenase (G6PD) was partially purified from red blood cells of 44 Sardinian males with severe enzymatic deficiency and the variants characterized chromatographically and biochemically to see if there is genetic heterogeneity of the enzyme in Southern Sardinia. Three different variants associated with severe G6PD deficiency were identified: the common G6PD Mediterranean variant; G6PD Sassari, recently described by Testa et al. (1980), and a new variant that we have designated G6PD Cagliari. Family studies were carried out to demonstrate that these variants are due to the presence of different alleles.