The thyroid hormone activating enzyme, type 2 deiodinase, induces myogenic differentiation by regulating mitochondrial metabolism and reducing oxidative stress.
Publication Date: 22/05/2019, on Redox biology
by Sagliocchi S, Cicatiello AG, Di Cicco E, Ambrosio R, Miro C, Di Girolamo D, Nappi A, Mancino G, De Stefano MA, Luongo C, Raia M, Ogawa-Wong AN, Zavacki AM, Paladino S, Salvatore D, Dentice M
Thyroid hormone (TH) is a key metabolic regulator that acts by coordinating short- and long-term energy needs. Accordingly, significant metabolic changes are observed depending on thyroid status. Although it is established that hyperthyroidism augments basal energy consumption, thus resulting in an enhanced metabolic state, the net effects on cellular respiration and generation of reactive oxygen species (ROS) remain unclear. To elucidate the effects of augmented TH signal in muscle cells, we generated a doxycycline-inducible cell line in which the expression of the TH-activating enzyme, type 2 deiodinase (D2), is reversibly turned on by the "Tet-ON" system. Interestingly, increased intracellular TH caused a net shift from oxidative phosphorylation to glycolysis and a consequent increase in the extracellular acidification rate. As a result, mitochondrial ROS production, and both the basal and doxorubicin-induced production of cellular ROS were reduced. Importantly, the expression of a set of antioxidant genes was up-regulated, and, among them, the mitochondrial scavenger Sod2 was specifically induced at transcriptional level by D2-mediated TH activation. Finally, we observed that attenuation of oxidative stress and increased levels of SOD2 are key elements of the differentiating cascade triggered by TH and D2, thereby establishing that D2 is essential in coordinating metabolic reprogramming of myocytes during myogenic differentiation. In conclusion, our findings indicate that TH plays a key role in oxidative stress dynamics by regulating ROS generation. Our novel finding that TH and its intracellular metabolism act as mitochondrial detoxifying agents sheds new light on metabolic processes relevant to muscle physiology.
Fatigue in Parkinson's disease: Italian validation of the Parkinson Fatigue Scale and the Fatigue Severity Scale using a Rasch analysis approach.
Publication Date: 22/05/2019, on Parkinsonism & related disorders
by Siciliano M, Chiorri C, De Micco R, Russo A, Tedeschi G, Trojano L, Tessitore A
The Fatigue Severity Scale (FSS-9) and the Parkinson Fatigue Scale (PFS-16) are commonly used for assessing fatigue in Parkinson's disease (PD). Here we validated the Italian version of these scales, assessed their psychometric properties by Rasch analysis, and computed their optimal cut-off scores using clinical diagnosis of PD-related fatigue as the gold standard.
Senescence Phenomena and Metabolic Alteration in Mesenchymal Stromal Cells from a Mouse Model of Rett Syndrome.
Publication Date: 21/05/2019, on International journal of molecular sciences
by Squillaro T, Alessio N, Capasso S, Di Bernardo G, Melone MAB, Peluso G, Galderisi U
Chromatin modifiers play a crucial role in maintaining cell identity through modulation of gene expression patterns. Their deregulation can have profound effects on cell fate and functions. Among epigenetic regulators, the MECP2 protein is particularly attractive. Mutations in the gene are responsible for more than 90% of cases of Rett syndrome (RTT), a progressive neurodevelopmental disorder. As a chromatin modulator, MECP2 can have a key role in the government of stem cell biology. Previously, we showed that deregulated MECP2 expression triggers senescence in mesenchymal stromal cells (MSCs) from (RTT) patients. Over the last few decades, it has emerged that senescent cells show alterations in the metabolic state. Metabolic changes related to stem cell senescence are particularly detrimental, since they contribute to the exhaustion of stem cell compartments, which in turn determine the falling in tissue renewal and functionality. Herein, we dissect the role of impaired MECP2 function in triggering senescence along with other senescence-related aspects, such as metabolism, in MSCs from a mouse model of RTT. We found that MECP2 deficiencies lead to senescence and impaired mitochondrial energy production. Our results support the idea that an alteration in mitochondria metabolic functions could play an important role in the pathogenesis of RTT.
Cardiotoxicity and pro-inflammatory effects of the immune checkpoint inhibitor Pembrolizumab associated to Trastuzumab.
Publication Date: 17/05/2019, on International journal of cardiology
by Quagliariello V, Passariello M, Coppola C, Rea D, Barbieri A, Scherillo M, Monti MG, Iaffaioli RV, De Laurentiis M, Ascierto PA, Botti G, De Lorenzo C, Maurea N
The immunotherapy has revolutionized the world of oncology in the last decades with considerable advantages in terms of overall survival in cancer patients. The association of Pembrolizumab and Trastuzumab was recently proposed in clinical trials for the treatment of Trastuzumab-resistant advanced HER2-positive breast cancer. Although immunotherapies are frequently associated with a wide spectrum of immune-related adverse events, the cardiac toxicity has not been properly studied.
Characterisation of the dynamic interactions between complex N-glycans and human CD22.
Publication Date: 16/05/2019, on Chembiochem : a European journal of chemical biology
by Di Carluccio C, Crisman E, Manabe Manabey Chem Sci Osaka-U Ac J YMMCSOAJ, Forgione RE, Lacetera A, Amato J, Pagano B, Randazzo A, Zampella A, Lanzetta R, Koichi F, Molinaro A, Crocker PR, Martin-Santamaria S, Marchetti R, Silipo A
CD22 (Siglec-2) is a B-cell surface inhibitory protein able to selectively recognize sialylated glycans, dampening autoimmune responses against self-antigens. We here characterize the dynamic recognition of complex-type N-glycans by human CD22, by means of orthogonal approaches including NMR spectroscopy, computational methods and biophysical assays. We provide novel molecular insights into the binding mode of sialoglycans in complex with h-CD22, highlighting the role of the sialic acid-galactose moieties in the recognition process, elucidating the conformational behaviour of complex-type N-glycans bound to Siglec-2 and dissecting the formation of CD22 homo-oligomers on the B-cell surface. Our results will enable the development of additional therapeutics able to modulate the activity of h-CD22 in autoimmune diseases and B-cell derived malignancies.
Intein-mediated protein trans-splicing expands adeno-associated virus transfer capacity in the retina.
Publication Date: 15/05/2019, on Science translational medicine
by Tornabene P, Trapani I, Minopoli R, Centrulo M, Lupo M, de Simone S, Tiberi P, Dell'Aquila F, Marrocco E, Iodice C, Iuliano A, Gesualdo C, Rossi S, Giaquinto L, Albert S, Hoyng CB, Polishchuk E, Cremers FPM, Surace EM, Simonelli F, De Matteis MA, Polishchuk R, Auricchio A
Retinal gene therapy with adeno-associated viral (AAV) vectors holds promises for treating inherited and noninherited diseases of the eye. Although clinical data suggest that retinal gene therapy is safe and effective, delivery of large genes is hindered by the limited AAV cargo capacity. Protein trans-splicing mediated by split inteins is used by single-cell organisms to reconstitute proteins. Here, we show that delivery of multiple AAV vectors each encoding one of the fragments of target proteins flanked by short split inteins results in protein trans-splicing and full-length protein reconstitution in the retina of mice and pigs and in human retinal organoids. The reconstitution of large therapeutic proteins using this approach improved the phenotype of two mouse models of inherited retinal diseases. Our data support the use of split intein-mediated protein trans-splicing in combination with AAV subretinal delivery for gene therapy of inherited blindness due to mutations in large genes.
Corrigendum to "Mathematical optimization of the green extraction of polyphenols from grape peels through a cyclic pressurization process" [Heliyon 5 (4) (April 2019) e01526].
Publication Date: 14/05/2019, on Heliyon
by Gallo M, Formato A, Giacco R, Riccardi G, Luongo D, Formato G, Amoresano A, Naviglio D
[This corrects the article DOI: 10.1016/j.heliyon.2019.e01526.].
Exploring cellular uptake, accumulation and mechanism of action of a cationic Ru-based nanosystem in human preclinical models of breast cancer.
Publication Date: 07/05/2019, on Scientific reports
by Piccolo M, Misso G, Ferraro MG, Riccardi C, Capuozzo A, Zarone MR, Maione F, Trifuoggi M, Stiuso P, D'Errico G, Caraglia M, Paduano L, Montesarchio D, Irace C, Santamaria R
According to WHO, breast cancer incidence is increasing so that the search for novel chemotherapeutic options is nowadays an essential requirement to fight neoplasm subtypes. By exploring new effective metal-based chemotherapeutic strategies, many ruthenium complexes have been recently proposed as antitumour drugs, showing ability to impact on diverse cellular targets. In the framework of different molecular pathways leading to cell death in human models of breast cancer, here we demonstrate autophagy involvement behind the antiproliferative action of a ruthenium(III)-complex incorporated into a cationic nanosystem (HoThyRu/DOTAP), proved to be hitherto one of the most effective within the suite of nucleolipidic formulations we have developed for the in vivo transport of anticancer ruthenium(III)-based drugs. Indeed, evidences are implicating autophagy in both cancer development and therapy, and anticancer interventions endowed with the ability to trigger this biological response are currently considered attractive oncotherapeutic approaches. Moreover, crosstalk between apoptosis and autophagy, regulated by finely tuned metallo-chemotherapeutics, may provide novel opportunities for future improvement of cancer treatment. Following this line, our in vitro and in vivo preclinical investigations suggest that an original strategy based on suitable formulations of ruthenium(III)-complexes, inducing sustained cell death, could open new opportunities for breast cancer treatment, including the highly aggressive triple-negative subtype.
Coping Strategies in Migraine without Aura: A Cross-Sectional Study.
Publication Date: 05/05/2019, on Behavioural neurology
by Russo A, Santangelo G, Tessitore A, Silvestro M, Trojsi F, De Mase A, Garramone F, Trojano L, Tedeschi G
In the context of a causal relationship between stress and migraine, coping strategies are aimed at managing stressful life events and reducing the distressing emotions connected to them.
Epigenetic targeting of autophagy for cancer prevention and treatment by natural compounds.
Publication Date: 02/05/2019, on Seminars in cancer biology
by Vidoni C, Ferraresi A, Secomandi E, Vallino L, Dhanasekaran DN, Isidoro C
Despite the undeniable progress made in the last decades, cancer continues to challenge the scientists engaged in searching for an effective treatment for its prevention and cure. One of the malignant hallmarks that characterize cancer cell biology is the altered metabolism of sugars and amino acids. Autophagy is a pathway allowing the macromolecular turnover via recycling of the substrates resulting from the lysosomal degradation of damaged or redundant cell molecules and organelles. As such, autophagy guarantees the proteome quality control and cell homeostasis. Data from in vitro, in animals and in patients researches show that dysregulation of autophagy favors carcinogenesis and cancer progression, making this process an ineluctable target of cancer therapy. The autophagy process is regulated at genetic, epigenetic and post-translational levels. Targeting autophagy with epigenetic modifiers could represent a valuable strategy to prevent or treat cancer. A wealth of natural products from terrestrial and marine living organisms possess anti-cancer activity. Here, we review the experimental proofs demonstrating the ability of natural compounds to regulate autophagy in cancer via epigenetics. The hope is that in the near future this knowledge could translate into effective intervention to prevent and cure cancer.
Amino acid response by Halofuginone in Cancer cells triggers autophagy through proteasome degradation of mTOR.
Publication Date: 02/05/2019, on Cell communication and signaling : CCS
by Follo C, Vidoni C, Morani F, Ferraresi A, Seca C, Isidoro C
In the event of amino acid starvation, the cell activates two main protective pathways: Amino Acid starvation Response (AAR), to inhibit global translation, and autophagy, to recover the essential substrates from degradation of redundant self-components. Whether and how AAR and autophagy (ATG) are cross-regulated and at which point the two regulatory pathways intersect remain unknown. Here, we provide experimental evidence that the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) specifically located at the lysosome level links the AAR with the autophagy pathway.
A rapid, safe, and quantitative in vitro assay for measurement of uracil-DNA glycosylase activity.
Publication Date: 01/05/2019, on Journal of molecular medicine (Berlin, Germany)
by Squillaro T, Finicelli M, Alessio N, Del Gaudio S, Di Bernardo G, Melone MAB, Peluso G, Galderisi U
Base excision repair (BER) is a frontline repair mechanism that operates through the G phase of the cell cycle, which ensures the genome integrity by repairing thousands of DNA lesions due to endogenous and exogenous agents. Its correct functioning is fundamental for cell viability and the health of the organism. Uracil is one of the most prevalent lesions that appears in DNA arising by spontaneous or enzymatic deamination of cytosine or misincorporation of the deoxyuridine 5'-triphosphate nucleotide (dUTP) in place of deoxythymidine 5'-triphosphate (dTTP) during DNA replication. In the first pathway, the uracil will preferentially pair with adenine, leading to C:G → T:A transition. When uracil in DNA arises from misincorporation of dUTP instead of dTTP, this process will result in A:U pairs. Organisms counteract the mutagenic effects of uracil in DNA using the BER repair system, which is mediated by a member of the uracil-DNA glycosylase (UDG) superfamily. Several assays evaluating the in vitro BER enzyme activity have been described so far. Some of these measure the BER activity by an oligonucleotide incision assay using radiolabeled duplex oligo. Others use circular double-stranded DNA substrates containing a defined lesion. The novelty of our method resides in its rapidity and safety (radioactive free detection) as well as in the possibility of having a reliable quantitative determination of UDG activity in both cell and tissue extracts. We also demonstrated the effectiveness of our method in assessing UDG activity in cell lines with a reduced DNA repair capacity and in different kinds of tissues. KEY MESSAGES: • Base excision repair is a fundamental repair mechanism ensuring the genome integrity. • Uracil is one of the most prevalent lesions that appears in DNA. • The mutagenic effects of uracil in DNA are mitigated by the uracil-DNA glycosylase. • Several assays evaluating the in vitro BER activity have been described so far. • A safe and quantitative assay evaluating the in vitro UDG activity is required.
Efficacy of the "first wave" Direct Acting antivirals against HCV infection: results from the Italian LINA (Liver Network Activity) cohort.
Publication Date: 29/04/2019, on The new microbiologica
by Gentile I, Buonomo AR, Coppola C, Staiano L, Amoruso DC, Saturnino MR, Maraolo AE, Portunato F, De Pascalis S, Martini S, Crispo M, Macera M, Pinchera B, Zappulo E, Scotto R, Coppola N
Approximately 71 million people are chronically infected with HCV worldwide. Recently, interferon-free therapies effective against HCV became available and nowadays, therapeutic strategies include a combination of two or three drugs with different mechanisms of action. In the present study, we reported real-life SVR rates in a large cohort of four prescribing centers in a high-endemic area of Southern Italy. We conducted a prospective multicenter study among all the patients with chronic HCV infection, who received therapy with the first available interferon-free therapies between March 2015 and December 2017 and who referred to one of the 4 DAA-prescribing centers in Campania, Southern Italy. Patients with Child C cirrhosis, a diagnosis of active HCC at the baseline or who refused the consent form, were excluded. Nine-hundred fifty-three patients were enrolled. Most of the enrolled patients had HCV genotype 1b infection (66.4%), were older than 65 years (64.1%) and had advanced liver fibrosis (Metavir > F4) (73.5%). The overall SVR12 rate was 98.5%. Patients with clinical cirrhosis had a similar SVR12 rate compared to those without cirrhosis (97.8% vs 99.2%, p=0.09), while patients with decompensated cirrhosis had a significantly lower rate of SVR12 compared to those without decompensated disease (95.3% vs 99.0%, p<0.05). Patients aged more than 65 years had a similar rate of SVR12 compared with patients aged ≤ 65 years (98.6% vs 98.0%, p=0.57). Among patients >65 years, those with clinical cirrhosis, as well as those with advanced liver fibrosis, had a similar SVR12 rate compared with the patients with a Metavir score < F4 (98.3% vs 99.0%, p=0.70 and 98.6% vs 98.6%, p=1.00, respectively). In the present, real-life study, DAA regimens are effective and safe in patients with chronic HCV infection, regardless of age and stage of liver disease, providing very high rates of SVR12 (98.5%).
Structure, stability and aggregation propensity of a Ribonuclease A-Onconase chimera.
Publication Date: 24/04/2019, on International journal of biological macromolecules
by Esposito L, Donnarumma F, Ruggiero A, Leone S, Vitagliano L, Picone D
Structural roles of loop regions are frequently overlooked in proteins. Nevertheless, they may be key players in the definition of protein topology and in the self-assembly processes occurring through domain swapping. We here investigate the effects on structure and stability of replacing the loop connecting the last two β-strands of RNase A with the corresponding region of the more thermostable Onconase. The crystal structure of this chimeric variant (RNaseA-ONC) shows that its terminal loop size better adheres to the topological rules for the design of stabilized proteins, proposed by Baker and coworkers . Indeed, RNaseA-ONC displays a thermal stability close to that of RNase A, despite the lack of Pro at position 114, which, due to its propensity to favor a cis peptide bond, has been identified as an important stabilizing factor of the native protein. Accordingly, RNaseA-ONC is significantly more stable than RNase A variants lacking Pro114; RNaseA-ONC also displays a higher propensity to form oligomers in native conditions when compared to either RNase A or Onconase. This finding demonstrates that modifications of terminal loops should to be carefully controlled in terms of size and sequence to avoid unwanted and/or potentially harmful aggregation processes.
The role of G-quadruplex structures of LIGS-generated aptamers R1.2 and R1.3 in IgM specific recognition.
Publication Date: 22/04/2019, on International journal of biological macromolecules
by Moccia F, Platella C, Musumeci D, Batool S, Zumrut H, Bradshaw J, Mallikaratchy P, Montesarchio D
Exploiting a variant of SELEX called "Ligand-Guided Selection" (LI-GS), we recently identified two novel truncated G-rich aptamers, called R1.2 and R1.3, specific for membrane-bound IgM (mIgM), the hallmark of B cells. Herein, the conformational behaviour of these aptamers has been analysed by multiple biophysical methods. In order to investigate their functional secondary structures, these studies have been carried out in pseudo-physiological buffers mimicking different cellular environments. Both aptamers proved to be highly polymorphic, folding into stable, unimolecular G-quadruplex structures in K-rich buffers. In turn, in buffered solutions containing Na/Mg ions, R1.2 and R1.3 formed mainly duplex structures. Remarkably, these aptamers were able to effectively bind mIgM on B-cell lymphoma exclusively in the presence of potassium ions. These findings demonstrate the key role of G-quadruplex folding in the molecular recognition and efficient binding of R1.2 and R1.3 to mIgM expressed in lymphoma and leukemia cells, providing a precious rational basis for the design of effective aptamer-based biosensors potentially useful for the detection of cancer-relevant biomarkers.