Cell Line and DNA Biobank From Patients Affected by Genetic Diseases

The Bioresource, presently storing 10,279 biospecimens, was initially established in 1976 as a private laboratory-collection to maintain rare mutant cell lines from genetic-metabolic diseases. Shortly afterwards, however, data from the sample collection was organised in a database and the sample collection was released to the scientific community. The Biobank has received Telethon grants since 1993, as individual facility, and from 2008 as part of the Telethon Network of Genetic Biobanks (www.biobanknetwork.org).In 2010, the Biobank has obtained official recognition from Regione Liguria. The Biobank has always provided essential services by establishing, analysing, maintaining, and distributing biospecimens from patients affected by rare genetic diseases. Up to now, the contribution of the Biobank to the scientific community has been expressed in acknowledgement notes in 145 scientific manuscripts

Defects in mitochondrial energetic function compels Fanconi Anaemia cells to glycolytic metabolism

Energetic metabolism plays an essential role in the differentiation of haematopoietic stem cells (HSC). In Fanconi Anaemia (FA), DNA damage is accumulated during HSC differentiation, an event that is likely associated with bone marrow failure (BMF). One of the sources of the DNA damage is altered mitochondrial metabolism and an associated increment of oxidative stress. Recently, altered mitochondrial morphology and a deficit in the energetic activity in FA cells have been reported. Considering that mitochondria are the principal site of aerobic ATP production, we investigated FA metabolism in order to understand what pathways are able to compensate for this energy deficiency. In this work, we report that the impairment in mitochondrial oxidative phosphorylation (OXPHOS) in FA cells is countered by an increase in glycolytic flux. By contrast, glutaminolysis appears lower with respect to controls. Therefore, it is possible to conclude that in FA cells glycolysis represents the main pathway for producing energy, balancing the NADH/NAD+ratio by the conversion of pyruvate to lactate. Finally, we show that a forced switch from glycolytic to OXPHOS metabolism increases FA cell oxidative stress. This could be the cause of the impoverishment in bone marrow HSC during exit from the homeostatic quiescent state. This is the first work that systematically explores FA energy metabolism, highlighting its flaws, and discusses the possible relationships between these defects and BMF

Defects in mitochondrial energetic function compels Fanconi Anaemia cells to glycolytic metabolism

Energetic metabolism plays an essential role in the differentiation of haematopoietic stem cells (HSC). In Fanconi Anaemia (FA), DNA damage is accumulated during HSC differentiation, an event that is likely associated with bone marrow failure (BMF). One of the sources of the DNA damage is altered mitochondrial metabolism and an associated increment of oxidative stress. Recently, altered mitochondrial morphology and a deficit in the energetic activity in FA cells have been reported. Considering that mitochondria are the principal site of aerobic ATP production, we investigated FA metabolism in order to understand what pathways are able to compensate for this energy deficiency. In this work, we report that the impairment in mitochondrial oxidative phosphorylation (OXPHOS) in FA cells is countered by an increase in glycolytic flux. By contrast, glutaminolysis appears lower with respect to controls. Therefore, it is possible to conclude that in FA cells glycolysis represents the main pathway for producing energy, balancing the NADH/NAD+ratio by the conversion of pyruvate to lactate. Finally, we show that a forced switch from glycolytic to OXPHOS metabolism increases FA cell oxidative stress. This could be the cause of the impoverishment in bone marrow HSC during exit from the homeostatic quiescent state. This is the first work that systematically explores FA energy metabolism, highlighting its flaws, and discusses the possible relationships between these defects and BMF

Treatment of FANCA cells with resveratrol and N-acetylcysteine: a comparative study.

Fanconi anemia (FA) is a genetic disorder characterised by chromosome instability, cytokine ipersensibility, bone marrow failure and abnormal haematopoiesis associated with acute myelogenous leukemia. Recent reports are contributing to characterize the peculiar FA metabolism. Central to these considerations appears that cells from complementation group A (FANCA) display an altered red-ox metabolism. Consequently the possibility to improve FA phenotypical conditions with antioxidants is considered. We have characterized from the structural and biochemical point of view the response of FANCA lymphocytes to N-acetyl-cysteine (NAC) and resveratrol (RV). Surprisingly both NAC and RV failed to revert all the characteristic of FA phenotype and moreover their effects are not super imposable. Our data suggest that we must be aware of the biological effects coming from antioxidant treatment

Impaired immune response to Candida albicans in cells from Fanconi anemia patients

Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure and cancer predisposi-tion. Several studies show alterations of the immunological status of FA patients including defects inperipheral blood lymphocyte subsets, serum immunoglobulin levels, and inflammatory cytokines.However scanty information is available on the response of FA cells to specific infectious antigens. In thiswork we examined the response of FA cells to different immunological stimuli and found a defectiveresponse of IL-1b, TNF-aand IL-17 toCandida albicansstimulation thus pointing to a potentially impairedresponse to fungal infections of FA patients

Morphological changes of mitochondria in FANCA lymphocytes and healthy controls.

<p><b>A.</b> Representative transmission electron microscopy images of morphologically altered mitochondria from untreated (a,c,e,g) and treated FANCA (b,d,f,h,i) with different antioxidants. Mitochondria of untreated lymphocytes from FANCA patients 1, 2, 3 (a,c,e) appeared swollen (black arrows) with increased size, disrupted cristae (black asterisks) and rarefacted matrix (black arrowheads). On the contrary, mitochondria in untreated lymphocytes from FANCA4 patient were small, oval and round with dark matrix and swollen internal cristae (black stars). (b) Mitochondria of FA1 lymphocytes showed worsened alteration such as more enlarged shape (white arrow) after NAC treatment. (d) Mitochondria of FANCA2 lymphocytes appeared indistinguishable from untreated (b) in overall aspect, after NAC treatment (white arrowheads). (f) Mitochondria of FANCA3 and FANCA4 lymphocytes showed a striking rescue after NAC or RV treatments (white asterisks). (Scale bars, 1 µm) <b>B.</b> Representative micrographs of morphologically normal mitochondria (arrowheads) in control lymphocytes at day 1 and day 5 before and after NAC or RV treatments. (Scale bars 1µm).</p

Biochemical parameters in FANCA and wild-type (wt) cells untreated or treated with 500 µm nac or 10 µm rv for 5 days.

<p>A. Table reports values for oxygen consumption (µM O₂/min/mg) in FANCA and wild type (wt) primary fibroblasts treated NAC or RV after induction with pyruvate/malate (Complexes I, III and IV), or succinate (Complexes II+III+IV). B. Assay of the electron transfer from Complex I to Complex III on wt and FANCA samples. Electron transfer between Complex I and III was measured following the reduction of cytochrome c at 550 nm after the addition of 0.7 mM NADH, the substrate of Complex I. Data are reported as µmol reduced Cytochrome c/min/mg. C. Figure shows the ratio among ATP and AMP concentration in wt and FANCA samples. D. Histogram reports a comparison of the ATP-AMP phosphotransferase activity (AK1+AK2 white columns) and GTP-AMP phosphotransferase (AK3, black columns) activity on wt and FANCA samples. The activity is expressed as µmol of ADP produced/min/mg. Data are expressed as mean ± SD. Each panel is representative of at least five experiments.</p