Generation and Characterization of a Transgenic Mouse Carrying a Functional Human ß -Globin Gene with the IVSI-6 Thalassemia Mutation.

Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. Transgenic mice carrying the ß-globin IVSI-6 mutation (the most frequent in Middle-Eastern regions and recurrent in Italy and Greece) are, at present, not available. We report the production and characterization of a transgenic mouse line (TG-ß-IVSI-6) carrying the IVSI-6 thalassemia point mutation within the human ß-globin gene. In the TG-ß-IVSI-6 mouse (a) the transgenic integration region is located in mouse chromosome 7; (b) the expression of the transgene is tissue specific; (c) as expected, normally spliced human ß-globin mRNA is produced, giving rise to ß-globin production and formation of a human-mouse tetrameric chimeric hemoglobin (mu) α-globin2/(hu) ß-globin2 and, more importantly, (d) the aberrant ß-globin-IVSI-6 RNAs are present in blood cells. The TG-ß-IVSI-6 mouse reproduces the molecular features of IVSI-6 ß-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides targeting the cryptic sites responsible for the generation of aberrantly spliced ß-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for ß-thalassemia.

Universal neonatal screening for sickle cell disease and other haemoglobinopathies in Ferrara, Italy.

BACKGROUND: Sickle cell disease is the commonest haemoglobinopathy in Africa, the Middle East and India. In recent years, its incidence has increased dramatically also in Europe and North America because of the high rate of migration of people from endemic areas. From January 2009 to January 2010 the number of foreign residents in the province of Ferrara (Italy) increased by 12.2%: most of the immigrants were from countries at high risk of sickle cell disease. Since neonatal screening and prophylactic penicillin in early childhood could reduce mortality by 10 years of age to less than 2%, the aim of this study was to establish a neonatal screening programme for haemoglobinopathies in Ferrara. MATERIALS AND METHODS: First we assessed how many pregnant women underwent haemoglobin analysis by high performance liquid chromatography before or during pregnancy and how many of them were carriers of haemoglobinopathies. Subsequently, we verified the feasibility of neonatal screening for sickle cell disease and other haemoglobinopathies, analysing cord blood by high performance liquid chromatography. Neonates found to be positive were managed by a multidisciplinary team to implement all the appropriate prophylactic and therapeutic measures. RESULTS: We found that 59% of women who delivered at the University Hospital of Ferrara, from 2007 to 2009, had undergone high performance liquid chromatography. Of the 41% who were not tested, many were from areas in which sickle cell disease is common. Between September 26th 2010 and January 31st 2012, 1992 neonatal tests were performed and 24 carriers of haemoglobinopathies were identified (16 with HbS, 4 with HbC, 2 with HbE, 1 with HbD Punjab and 1 with HbD-Ouled Rabah); 42.6% of the mothers of these 1,992 neonates had not undergone high performance liquid chromatography during pregnancy. DISCUSSION: Currently prevention of haemoglobinopathies in Italy is provided during the pre-conception period but only to patients with abnormal blood counts. Neonatal screening is useful and cost-effective to ensure early diagnosis and appropriate treatment for infants with sickle cell disease or other haemoglobinopathies.

Multidimensional evaluation of performance with experimental application of balanced scorecard: a two year experience.

BACKGROUND: In today's dynamic health-care system, organizations such as hospitals are required to improve their performance for multiple stakeholders and deliver an integrated care that means to work effectively, be innovative and organize efficiently. Achieved goals and levels of quality can be successfully measured by a multidimensional approach like Balanced Scorecard (BSC). The aim of the study was to verify the opportunity to introduce BSC framework to measure performance in St. Anna University Hospital of Ferrara, applying it to the Clinical Laboratory Operative Unit in order to compare over time performance results and achievements of assigned targets. METHODS: In the first experience with BSC we distinguished four perspectives, according to Kaplan and Norton, identified Key Performance Areas and Key Performance Indicators, set standards and weights for each objective, collected data for all indicators, recognized cause-and-effect relationships in a strategic map. One year later we proceeded with the next data collection and analysed the preservation of framework aptitude to measure Operative Unit performance. In addition, we verified the ability to underline links between strategic actions belonging to different perspectives in producing outcomes changes. RESULTS: The BSC was found to be effective for underlining existing problems and identifying opportunities for improvements. The BSC also revealed the specific perspective contribution to overall performance enhancement. After time results comparison was possible depending on the selection of feasible and appropriate key performance indicators, which was occasionally limited by data collection problems. CONCLUSIONS: The first use of BSC to compare performance at Operative Unit level, in course of time, suggested this framework can be successfully adopted for results measuring and revealing effective health factors, allowing health-care quality improvements.

Therapeutic concentrations of mitotane (o,p'-DDD) inhibit thyrotroph cell viability and TSH expression and secretion in a mouse cell line model.

Mitotane therapy is associated with many side effects, including thyroid function perturbations mimicking central hypothyroidism, possibly due to laboratory test interference or pituitary direct effects of mitotane. We investigated whether increasing concentrations of mitotane in the therapeutic range might interfere with thyroid hormone assays and evaluated the effects of mitotane on a mouse TSH-producing pituitary cell line. TSH, free T(4), and free T(3) levels do not significantly change in sera from hypo-, hyper-, or euthyroid patients after addition of mitotane at concentrations in the therapeutic window. In the mouse TalphaT1 cell line, mitotane inhibits both TSH expression and secretion, blocks TSH response to TRH, and reduces cell viability, inducing apoptosis at concentrations in the therapeutic window. TRH is not capable of rescuing TalphaT1 cells from the inhibitory effects of mitotane on TSH expression and secretion, which appear after short time treatment and persist over time. Our results demonstrate that mitotane does not interfere with thyroid hormone laboratory tests but directly reduces both secretory activity and cell viability on pituitary TSH-secreting mouse cells. These data represent a possible explanation of the biochemical picture consistent with central hypothyroidism in patients undergoing mitotane therapy and open new perspectives on the direct pituitary effects of this drug.

Multidimensional evaluation of performance: experimental application of the balanced scorecard in Ferrara university hospital.

BACKGROUND AND AIMS: One of the best-known performance planning and evaluation techniques utilising both monetary and non-monetary data is the Balanced Scorecard (BSC). This is a means of rationalising the global activity of a business in the attempt to create value, and to translate the company vision into a set of tactical objectives and measurable strategies. The aim of this study was to implement and evaluate the use of BSC in two departments of the St. Anna University Hospital, Ferrara: the Analysis Laboratory and Digestive Endoscopy operating units (OU). MATERIALS AND METHODS: With the collaboration of the health workers involved, a precise methodological programme was pursued: Definition of the strategic map from 4 perspectives, according to Kaplan and Norton, Definition of the Key Performance Areas (KPA), or macro-objectives, Identification of the cause-effect relationships between KPAs, Identification of the sub-objectives of each KPA, Definition of the Key Performance Indicators (KPI), Definition of the weight/importance of each objective in the global evaluation. RESULTS: The information gathered permitted the definition of macro- and sub-objectives for each perspective, as well as determining the relevant indicators, standards, weights, frequency of detection and means of acquisition. Strategic maps showing the cause/effect relationships in each OU were created, as were 'evaluation panels', which describe the global performance of each department. For each perspective, the fundamental data were summarised in one table. Evaluation of each perspective yielded a positive result for the majority of the objectives, and the global result (including all 4 perspectives) was found to be satisfactory. DISCUSSION-CONCLUSION: The Balanced Scorecard was implemented in the abovementioned OUs of St. Anna University Hospital, Ferrara, after the health workers themselves realised the need for change.In our research the employees were pleased to be evaluated, not only for the financial outcomes, but also for the satisfaction of improving internal procedure, relationships with the community and their own growth/learning. BSC is an ideal point of contact between the financial and clinical dimensions of management. However, difficulties in its application were faced, among these, at least in the initial phase, the lack of information systems able to drive it, and the complexity of the research for specific indicators needed to be overcome. The time factor (on average, at least two years are required) and the availability of technological resources were also limiting factors.The rapid diffusion of BSC among the principal international profit and non-profit organisations is testament to its great potential. This project could be seen as a preparatory phase in the strategical analysis of a subsequent business plan.

Surface plasmon resonance and biosensor technology for real-time molecular diagnosis of beta o 39 thalassemia mutation.

BACKGROUND: Biospecific interaction analysis (BIA) employing surface plasmon resonance (SPR) and biosensor technologies is of interest in clinical genetics. However, few data are available on its use in hereditary diseases caused by genetic mutations. AIM: The primary aim of this study was the refinement of BIA technology for use in identifying the beta o 39 mutation of the beta-globin gene, a mutation which causes a common type of beta o thalassemia. METHODS: Target-biotinylated PCR products were immobilized on streptavidin-coated sensor chips and diagnosed using SPR-based BIA performed by injecting specific oligonucleotide probes into the sensor chip. RESULTS: We demonstrated that the beta o 39 mutation can be easily and reproducibly identified during the association phase. CONCLUSIONS: This should be considered a pilot study demonstrating the ability of SPR-based BIA to detect point mutations in the beta-globin gene by real-time monitoring of hybridization between oligonucleotide probes and target-biotinylated PCR products generated from genomic DNA from normal, heterozygous individuals and homozygous beta o thalassemia patients.