Biomedical subjects

Cristina Bianchi

Publications and source records attributed to Cristina Bianchi.

At least 19 recordsLinked to original sources

PubMed2006

Left ventricular function in normotensive young adults with well-controlled type 1 diabetes mellitus.

The aim of this study was to determine whether early myocardial structural and functional systolic and diastolic alterations in asymptomatic and uncomplicated patients with type 1 diabetes mellitus (DM) could be detected using the new highly sensitive echocardiographic techniques of integrated backscatter and color Doppler myocardial imaging. Forty asymptomatic and uncomplicated patients with type 1 DM and 40 gender- and age-matched normal controls were studied. All patients were analyzed by conventional and new echocardiographic techniques (integrated backscatter and color Doppler myocardial imaging). Patients with DM showed reduced systolic function compared with controls, evidenced by significantly lower peak strain, strain rates, and cyclic variation indexes at the septum (p <0.0001, <0.01, and <0.001, respectively) and at the posterior wall level (p <0.0001, <0.0001, and <0.001, respectively). On receiver-operating characteristic curve analysis, systolic strain and the cyclic variation index showed the highest discriminating power for separating patients with DM and control subjects. Neither structural or ultrastructural nor diastolic functional abnormalities were detected. On univariate regression analysis, a significant inverse correlation was found for DM duration with conventional (E/A ratio) and unconventional (tissue Doppler imaging E/A ratio) indexes of diastolic function, in the absence of any correlation for systolic function. In conclusion, highly sensitive ultrasonic techniques demonstrate evidence of left ventricular systolic dysfunction in the early stage of type 1 DM, in the absence of ultrastructural and left ventricular diastolic functional abnormalities.

Adult↗

PubMed2006

Mitochondrial Complex I: structural and functional aspects.

This review examines two aspects of the structure and function of mitochondrial Complex I (NADH Coenzyme Q oxidoreductase) that have become matter of recent debate. The supramolecular organization of Complex I and its structural relation with the remainder of the respiratory chain are uncertain. Although the random diffusion model [C.R. Hackenbrock, B. Chazotte, S.S. Gupte, The random collision model and a critical assessment of diffusion and collision in mitochondrial electron transport, J. Bioenerg. Biomembranes 18 (1986) 331-368] has been widely accepted, recent evidence suggests the presence of supramolecular aggregates. In particular, evidence for a Complex I-Complex III supercomplex stems from both structural and kinetic studies. Electron transfer in the supercomplex may occur by electron channelling through bound Coenzyme Q in equilibrium with the pool in the membrane lipids. The amount and nature of the lipids modify the aggregation state and there is evidence that lipid peroxidation induces supercomplex disaggregation. Another important aspect in Complex I is its capacity to reduce oxygen with formation of superoxide anion. The site of escape of the single electron is debated and either FMN, iron-sulphur clusters, and ubisemiquinone have been suggested. The finding in our laboratory that two classes of hydrophobic inhibitors have opposite effects on superoxide production favours an iron-sulphur cluster (presumably N2) is the direct oxygen reductant. The implications in human pathology of better knowledge on these aspects of Complex I structure and function are briefly discussed.

Animals↗

PubMed2005

Clinical and epidemiological features of oral cancer in a medical school teaching hospital from 1994 to 2002: increasing incidence in women, predominance of advanced local disease, and low incidence of neck metastases.

PURPOSE: Description of clinical and epidemiological characteristics of patients who underwent surgery for oral cancer in a Medical School Teaching Hospital, and determination of differences with respect to other institutions and/or periods of time. METHOD: The charts of patients undergoing surgery for oral cancer from 1994 to 2002 were reviewed. Data were collected in a spreadsheet in order to analyze clinical and epidemiological features.. RESULTS: A total of 374 patients having undergone 406 operations was identified. Their ages varied from 14 to 94 years (mean = 57.4 years), with 255 men (68.2%), and 295 out 366 Caucasian (80.6%). A majority had tumors of the tongue and/or floor of mouth (55.6%), while 20.3% had lip cancer. Squamous cell carcinoma was found in 90.3%, and glandular carcinoma in 4%. T4 tumors in 39.6%, Tis or T1 lesions in 15.2% of all patients. Nearly 62% had no regional metastases, and the relative incidence in young patients (40 years or younger) reached 8.6%. CONCLUSION: In spite of the predominance of locally advanced tumors, a majority of patients had no neck metastases. The 31.8% incidence in females indicates an increasing incidence of oral cavity cancer among women when compared to previous periods at the same institution.

Adolescent↗

PubMed2005

Supercomplex organization of the mitochondrial respiratory chain and the role of the Coenzyme Q pool: pathophysiological implications.

In this review we examine early and recent evidence for an aggregated organization of the mitochondrial respiratory chain. Blue Native Electrophoresis suggests that in several types of mitochondria Complexes I, III and IV are aggregated as fixed supramolecular units having stoichiometric proportions of each individual complex. Kinetic evidence by flux control analysis agrees with this view, however the presence of Complex IV in bovine mitochondria cannot be demonstrated, presumably due to high levels of free Complex. Since most Coenzyme Q appears to be largely free in the lipid bilayer of the inner membrane, binding of Coenzyme Q molecules to the Complex I-III aggregate is forced by its dissociation equilibrium; furthermore free Coenzyme Q is required for succinate-supported respiration and reverse electron transfer. The advantage of the supercomplex organization is in a more efficient electron transfer by channelling of the redox intermediates and in the requirement of a supramolecular structure for the correct assembly of the individual complexes. Preliminary evidence suggests that dilution of the membrane proteins with extra phospholipids and lipid peroxidation may disrupt the supercomplex organization. This finding has pathophysiological implications, in view of the role of oxidative stress in the pathogenesis of many diseases.

Animals↗

PubMed2005

N- and C-terminal isoforms of Arg quantified by real-time PCR are specifically expressed in human normal and neoplastic cells, in neoplastic cell lines, and in HL-60 cell differentiation.

The human ABL2 (or ARG) gene codes for a nonreceptor tyrosine kinase is involved in translocation with the ETV6 gene in human leukemia and has an altered expression in several human carcinomas. Two isoforms of Arg with different N-termini (1A and 1B) have been described. The C-terminal domain of Arg contains two F-actin-binding sequences that perform a number of actions related to cell morphology and motility by interacting with actin filaments. We have identified different-sized specific cDNAs in hematopoietic, epithelial, nervous, and fibroblastic cells by means of the reverse transcription (RT)-polymerase chain reaction (PCR) analysis of human Arg mRNA. Some of these cDNAs showed an adjunctive alternative splice event involving the 63 bp sequence of exon II, thus leading to four cDNA types with different N-termini: 1A long and short, and 1B long and short. Other cDNAs lacked a 309 bp sequence in the last exon involving one of the C-terminal F-actin binding domains, thus giving rise to two cDNA types: C-termini long and short. Quantified by real-time PCR-quantitative RT-PCR-these Arg transcript isoforms have specific expression patterns not only in different normal and tumor cell types, but also during cell differentiation and growth arrest. These isoforms maintained the open reading frames, and eight putative proteins were predicted. The different C-termini isoforms seem to retain the same quantitative reciprocal ratio of their respective transcripts. The Arg protein isoforms with different C-terminal actin-binding domains and different N-termini might have specific cellular localizations/concentrations, and differently regulated catalytic activity with different implications in normal and neoplastic cells.

Amino Acid Sequence↗

PubMed2005

Prevalence of the metabolic syndrome among Italian adults according to ATP III definition.

BACKGROUND AND AIM: To evaluate the prevalence of the metabolic syndrome as defined by NCEP ATP III criteria in an Italian cohort of adult subjects. METHODS AND RESULTS: A total of 2100 subjects aged 19 years or more, were randomly selected from the general population of the Lucca area. Metabolic syndrome was defined by the clustering of three or more of the following abnormalities: waist circumference greater than 102 cm in men and 88 cm in women; serum triglycerides level of at least 150 mg/dl (1.69 mmol/l); high-density lipoprotein cholesterol less than 40 mg/dl (1.04 mmol/l) in men and 50 mg/dl (1.29 mmol/l) in women; blood pressure greater than 130/85 mmHg; or serum glucose greater than 110 mg/dl (6.1 mmol/l). The prevalence of the metabolic syndrome was 18% in women and 15% in men. The prevalence increased from 3% among subjects aged 20-29 years to 25% in subjects aged 70 years or older. Application of this estimated prevalence data to the Italian adult population suggests that 3.6 million women and 3 million men may have the metabolic syndrome. CONCLUSIONS: The prevalence of metabolic syndrome in a cohort likely representative of the Italian adult population is high. The recognition of the syndrome may represent an important challenge for physicians and healthcare and requires immediate strategies aimed to reduce level of individual metabolic traits.

Adult↗

PubMed2004

The mitochondrial respiratory chain is partially organized in a supercomplex assembly: kinetic evidence using flux control analysis.

The model of the respiratory chain in which the enzyme complexes are independently embedded in the lipid bilayer of the inner mitochondrial membrane and connected by randomly diffusing coenzyme Q and cytochrome c is mostly favored. However, multicomplex units can be isolated from mammalian mitochondria, suggesting a model based on direct electron channeling between complexes. Kinetic testing using metabolic flux control analysis can discriminate between the two models: the former model implies that each enzyme may be rate-controlling to a different extent, whereas in the latter, the whole metabolic pathway would behave as a single supercomplex and inhibition of any one of its components would elicit the same flux control. In particular, in the absence of other components of the oxidative phosphorylation apparatus (i.e. ATP synthase, membrane potential, carriers), the existence of a supercomplex would elicit a flux control coefficient near unity for each respiratory complex, and the sum of all coefficients would be well above unity. Using bovine heart mitochondria and submitochondrial particles devoid of substrate permeability barriers, we investigated the flux control coefficients of the complexes involved in aerobic NADH oxidation (I, III, IV) and in succinate oxidation (II, III, IV). Both Complexes I and III were found to be highly rate-controlling over NADH oxidation, a strong kinetic evidence suggesting the existence of functionally relevant association between the two complexes, whereas Complex IV appears randomly distributed. Moreover, we show that Complex II is fully rate-limiting for succinate oxidation, clearly indicating the absence of substrate channeling toward Complexes III and IV.

Adenosine Triphosphate↗

PubMed2004

Yessotoxin, a shellfish biotoxin, is a potent inducer of the permeability transition in isolated mitochondria and intact cells.

The diarrhetic poisoning by bivalve molluscs, diarrhetic shellfish poisoning, is due to consumption of mussels containing biotoxins produced by some Dinoflagellate species. Toxic effects of yessotoxin (YTX) include morphological alterations of mitochondria from heart and liver but the biochemical basis for these alterations is completely unknown. This paper demonstrates that YTX is a very powerful compound that opens the permeability transition pore (PTP) of the inner mitochondrial membrane of rat liver mitochondria at nanomolar concentrations. The effect requires the presence of a permissive level of calcium, by itself incapable of opening the pore. The direct effect of YTX on PTP is further confirmed by the inhibition exerted by cyclosporin A (CsA) that is known as a powerful inhibitor of PTP opening. Moreover, YTX induces membrane depolarization as shown by the quenching of tetramethylrhodamine methyl ester (TMRM), also prevented by the addition of CsA. YTX caused PTP opening in Morris Hepatoma 1C1 cells, as shown by the occurrence of CsA-sensitive depolarization within minutes of the addition of submicromolar concentrations of the toxin. These results provide a biochemical basis for the mitochondrial alterations observed in the course of intoxication with YTX, offering the first clue into the pathogenesis of diseases caused by YTX, and providing a novel tool to study the PTP in situ.

Animals↗

PubMed2004

The mitochondrial production of reactive oxygen species in relation to aging and pathology.

Mitochondria are known to be strong producers of reactive oxygen species (ROS) and, at the same time, particularly susceptible to the oxidative damage produced by their action on lipids, proteins, and DNA. In particular, damage to mtDNA induces alterations to the polypeptides encoded by mtDNA in the respiratory complexes, with consequent decrease of electron transfer, leading to further production of ROS and thus establishing a vicious circle of oxidative stress and energetic decline. This deficiency in mitochondrial energetic capacity is considered the cause of aging and age-related degenerative diseases. Complex I would be the enzyme most affected by ROS, since it contains seven of the 13 subunits encoded by mtDNA. Accordingly, we found that complex I activity is significantly affected by aging in rat brain and liver mitochondria as well as in human platelets. Moreover, due to its rate control over aerobic respiration, such alterations are reflected on the entire oxidative phosphorylation system. We also investigated the role of mitochondrial complex I in superoxide production and found that the one-electron donor to oxygen is most probably the Fe-S cluster N2. Short chain coenzyme Q (CoQ) analogues enhance ROS formation, presumably by mediating electron transfer from N2 to oxygen, both in bovine heart SMP and in cultured HL60 cells. Nevertheless, we have accumulated much evidence of the antioxidant role of reduced CoQ(10) in several cellular systems and demonstrated the importance of DT-diaphorase and other internal cellular reductases to reduce exogenous CoQ(10) after incorporation.

Aging↗

PubMed2004

Impairment of mitochondrial oxidative phosphorylation in rat fatty liver exposed to preservation-reperfusion injury.

BACKGROUND/AIMS: As the impairment of the cellular energy metabolism contributes to the failure of fatty liver grafts after transplantation, we aimed to determine whether steatosis affects the oxidative phosphorylation activity during preservation. METHODS: Rat normal and fatty livers were preserved for 18 h and then reperfused with warm oxygenated solution. The oxidative phosphorylation, the F(0)F(1)-ATPase and the Complex I activities were assessed in isolated mitochondria before and after preservation, and during reperfusion. The ALT release and portal pressure were monitored during reperfusion. RESULTS: The baseline phosphorylation activity was similar in normal and steatotic mitochondria. After cold preservation, the respiratory control index and state 3 respiration decreased significantly only in steatotic livers. Reperfusion induced a further deterioration in either group. Contrary to normal liver, uncoupling of fatty liver mitochondria allowed the recovery of the maximal respiration rate only using succinate (Complex II-dependent substrate), but not glutamate-malate (Complex I-dependent). Complex I dysfunction was confirmed spectrophotometrically. The ATPase activity was also significantly lower in fatty livers. Finally, ALT release and portal pressure were greater in steatotic livers. CONCLUSIONS: The alteration of the oxidative phosphorylation activity during preservation is greatly exacerbated by fatty infiltration likely resulting from damage of the respiratory chain Complex I and of the F(0)F(1)-ATP synthase.

Adenosine Triphosphate↗

PubMed2004

Evaluation of cellular energetics by the pasteur effect in intact cardiomyoblasts and isolated perfused hearts.

This work aims at exploring changes in cellular energetics by exploiting the Pasteur effect. We assumed that lactate overproduction arising from antimycin A-induced inhibition of mitochondrial respiration (delta-lactate = stimulated [lactate] -basal [lactate]) is indicative of the energy provided aerobically by the cell. Rat embryonal cardiomyocytes (H9c2), incubated with 2 micromol/L antimycin A, increased about 6 fold their lactate production in a manner linear with time and cell number. Antimycin A was also delivered to Langendorff-perfused rat hearts under control aerobic conditions or after 20 min-ischemia and 30 min-reperfusion. The test started at the end of each perfusion and lactate was measured into perfusate collected for further 25 min. A cardioplegic solution was also delivered during the test to exclude that lactate production was influenced by cardiac contraction. Control delta-lactate was 20.9 +/- 2.31 (S.E.M.) microg/mL and markedly decreased after reperfusion (7.66 +/- 0.51, p < 0.001), showing that energy production was impaired of about 70%. The determination of oxygen consumption by mitochondria isolated from reperfused hearts also suggested that the damage to the respiratory chain was similar to that evaluated by lactate overproduction (Respiratory Control Index: 75% lower than control, p < 0.001). Moreover, when delta-lactate was referred to the estimated cells which remained viable at the end of reperfusion (49.9%), it was 25% lower than control (p < 0.05). Therefore, we proposed this test as a tool for quantifying both physiological and pathological energetic modifications in living intact cardiomyocytes and in isolated and perfused hearts.

Animals↗

PubMed2004

Is oral tolerance correlated with IgE levels and mast cell numbers?

A triple genetic association between serum IgE levels, mast cell numbers, and susceptibility to oral tolerance was observed in mice genetically selected to extreme phenotypes of oral tolerance (susceptibility and resistance), suggesting a coadaptation of genes controlling these traits.

Animals↗

PubMed2003

Structural and functional organization of Complex I in the mitochondrial respiratory chain.

Metabolic flux control analysis of NADH oxidation in bovine heart submitochondrial particles revealed high flux control coefficients for both Complex I and Complex III, suggesting that the two enzymes are functionally associated as a single enzyme, with channelling of the common substrate, Coenzyme Q. This is in contrast with the more accepted view of a mobile diffusable Coenzyme Q pool between these enzymes. Dilution with phospholipids of a mitochondrial fraction enriched in Complexes I and III, with consequent increased theoretical distance between complexes, determines adherence to pool behavior for Coenzyme Q, but only at dilution higher than 1:5 (protein:phospholipids), whereas, at lower phospholipid content, the turnover of NADH cytochrome c reductase is higher than expected by the pool equation.

Animals↗

PubMed2003

Structural organization of the mitochondrial respiratory chain.

Two models exist of the mitochondrial respiratory chain: the model of a random organization of the individual respiratory enzyme complexes and that of a super-complex assembly formed by stable association between the individual complexes. Recently Schägger, using digitonin solubilization and Blue Native PAGE produced new evidence of preferential associations, in particular a Complex I monomer with a Complex III dimer, and suggested a model of the respiratory chain (the respirasome) based on direct electron channelling between complexes. Discrimination between the two models is amenable to kinetic testing using flux control analysis. Experimental evidence obtained in beef heart SMP, according to the extension of the Metabolic Control Theory for pathways with metabolic channelling, showed that enzyme associations involving Complex I and Complex III take place in the respiratory chain while Complex IV seems to be randomly distributed, with cytochrome c behaving as a mobile component. Flux control analysis at anyone of the respiratory complexes involved in aerobic succinate oxidation indicated that Complex II and III are not functionally associated in a stable supercomplex. A critical appraisal of the solid-state model of the mitochondrial respiratory chain requires its reconciliation with previous biophysical and kinetic evidence that CoQ behaves as a homogeneous diffusible pool between all reducing enzyme and all oxidizing enzymes: the hypothesis can be advanced that both models (CoQ pool and supercomplexes) are true, by postulating that supercomplexes physiologically exist in equilibrium with isolated complexes depending on metabolic conditions of the cell.

Animals↗

PubMed2002

The expression of the non-receptor tyrosine kinases Arg and c-abl is differently modulated in B lymphoid cells at different stages of differentiation.

The products of the human ARG gene and the human ABL gene characterize the Abelson family of non-receptor tyrosine protein kinases. Both genes are ubiquitously expressed. The interactions of these two similar protein kinases are still not well known, although it has been suggested that they could cooperate, with redundant actions, to provide intracellular signals in the cells. Lymphopenia occurs in mice with homozygous disruption of c-abl, indicating that in certain tissues Arg is unable to substitute c-abl functions. In B and T lymphoid cell lines at different stages of differentiation, we studied, by a reverse transcriptase-competitive polymerase chain reaction and Western blotting, Arg and c-abl in order to evaluate whether the expression pattern of the two genes could give insight as to why they do not exhibit overlapping roles in lymphocytes and whether the product levels of the two genes are related to lymphoid differentiation. The data showed that their expression is differently modified in lymphoid B cell lines. The highest Arg transcript and protein levels are in the mature B cells.

Actins↗

PubMedDate not supplied

Drop of connexin 43 in replicative senescence of human fibroblasts HEL-299 as a possible biomarker of senescence.

The expression of connexin 43 (cx43) and cell-cell communication were studied in replicative senescence of cultured HEL-299 fibroblasts. A progressive decrease in fluorescent dye transfer was detected by a scrape-loading technique in aging fibroblasts. This change was accounted for by a marked decrease in the amount of cx43 in aging cells, as detected by western blot analysis (cell extracts) and indirect fluorescence (cells in culture). However, semiquantitative RT-PCR assays of cx43 mRNA did not reveal appreciable changes, which suggests several possible explanations for the mechanism(s) underlying the decrease of cx43 in aging cells. These findings support the idea that the reduced expression of cx43 might be a biomarker of cell senescence.

Biomarkers↗

PubMedDate not supplied

Primary cell cultures arising from normal kidney and renal cell carcinoma retain the proteomic profile of corresponding tissues.

Renal cell carcinoma (RCC) tissue is composed of a mixture of neoplastic and normal cells, which complicate proteome analysis. The aim of our study was to investigate whether it is feasible to establish primary cell cultures of RCC and of renal cortex maintaining the tissue phenotype along with a more homogeneous and enriched cytological material. Fourteen (82.3%) primary cultures from 17 surgical cases were established and characterized by morphology, growth rate, immunocytochemistry, and molecular analysis performed by Real-time PCR, Western blotting, two-dimensional electrophoresis (2-DE), and mass spectrometry. Cultures showed >90% cytokeratine-positive epithelial cells. In primary tumor cultures, the molecular phenotype of manganese superoxide dismutase and heat shock protein 27 was the same as that found in tumor tissues with overexpression and increased number of isoforms. Moreover, 27 out 28 specific proteins and their isoforms, present in spots excised from 2-DE gel of cortex or RCC cultures, corresponded to those identified on the 2-DE tissue cortex reference map, suggesting that these primary cultures retain the proteomic profile of the corresponding tissues.

Adult↗

PubMedDate not supplied

The role of quantitative polymerase chain reaction in the management of follicular lymphoma patients.

AIMS AND BACKGROUND: In order to increase the prognostic significance of polymerase chain reaction (PCR) data it has been suggested that quantitative PCR can be used to measure tumor burden. However, this option has not yet been definitely supported or refuted in patients with follicular lymphoma (FL). We decided to evaluate whether knowledge of the quantitative level of minimal residual disease and its variations can be of use in the management of FL patients. METHODS: We used qualitative and competitive PCR to study 11 patients with refractory or relapsed FL harboring the t(14;18) translocation who underwent autologous (nine patients) or allogeneic (two patients) stem cell transplantation (SCT). Competitive PCR was performed with a multiple competitor carrying specific sequences including Bcl2/IgH MBR and mcr, and the beta-globin gene. RESULTS: After a median post-SCT follow-up of 44 months (range, 12-62), overall survival was 91% and disease-free survival 82%. The quantitative PCR data showed that: 1) effective chemotherapy before SCT substantially (1-2 log) reduced the tumor burden in the bone marrow (BM); 2) the increase in rearranged DNA detected in BM was associated with disease progression and relapse; 3) a PCR-negative autograft seemed to lead to lasting molecular remission even when it was performed in patients with a low level of BM infiltration before transplant; and 4) allo-SCT made and maintained the BM PCR negative even in the presence of a greater tumor burden before SCT. Six of the nine patients having CR after SCT (four auto and two allo) are in continuous molecular remission. CONCLUSIONS: In FL patients qualitative and quantitative PCR may provide data that can be helpful for the prognostic evaluation of tumor progression and the early detection of impending relapse by highlighting biological features such as the quality of the infused material, the tumor burden at transplant, and the behavior of tumor cells after transplant.

Adult↗