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  1. Article ; Online: Development of immunodiagnostic tools for in situ investigation of Ovis aries papillomavirus 3 (OaPV3).

    Cacciotto, Carla / Dore, Gian Mario / Anfossi, Antonio Giovanni / Tore, Gessica / Varoni, Maria Vittoria / Demontis, Maria Piera / Antuofermo, Elisabetta / Pittau, Marco / Alberti, Alberto

    Veterinary research communications

    2022  Volume 47, Issue 2, Page(s) 641–649

    Abstract: Cutaneous squamous cell carcinoma (cSCC) is a malignant lesion characterized by proliferation and transformation of keratinocytes in the epidermis and infiltrating derma. cSCC is reported in domestic and wild animal species, worldwide. The occurrence and ...

    Abstract Cutaneous squamous cell carcinoma (cSCC) is a malignant lesion characterized by proliferation and transformation of keratinocytes in the epidermis and infiltrating derma. cSCC is reported in domestic and wild animal species, worldwide. The occurrence and development of cSCC rely on synergic multifactorial conditions, most importantly sunlight exposure and Papillomavirus (PV) infection. In sheep, the development of such lesions represents a threat both to animal welfare and milk production. Ovis aries papillomavirus 3 (OaPV3) is the main cSCC viral determinant and oncogenic properties of viral E6 and E7 proteins were preliminarily investigated. However, E6 and E7 role and mechanisms resulting in cSCC have not been fully clarified, mainly due to the lack specific immunological tools, such as antibodies for in situ detection of ovine papillomavirus. This paper reports the development of specific serological tools for the investigation of OaPV3 pathogenicity, and their preliminary use to screen 4 ovine cSSC formalin-fixed paraffin embedded tissues. Relevance of immunological tools to investigation of viral biological properties and diagnosis are also discussed.
    MeSH term(s) Sheep ; Animals ; Carcinoma, Squamous Cell/diagnosis ; Carcinoma, Squamous Cell/veterinary ; Carcinoma, Squamous Cell/pathology ; Sheep, Domestic ; Skin Neoplasms/diagnosis ; Skin Neoplasms/veterinary ; Skin Neoplasms/pathology ; Papillomaviridae ; Sheep Diseases/diagnosis ; Sheep Diseases/pathology
    Language English
    Publishing date 2022-11-04
    Publishing country Switzerland
    Document type Journal Article
    ZDB-ID 406735-6
    ISSN 1573-7446 ; 0165-7380
    ISSN (online) 1573-7446
    ISSN 0165-7380
    DOI 10.1007/s11259-022-10018-5
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: Cadmium and wild boar: Environmental exposure and immunological impact on macrophages.

    Franzoni, Giulia / Ciccotelli, Valentina / Masiello, Lucia / De Ciucis, Chiara Grazia / Anfossi, Antonio Giovanni / Vivaldi, Barbara / Ledda, Mauro / Zinellu, Susanna / Dei Giudici, Silvia / Berio, Enrica / Tiziana, Andreoli / Dellepiane, Monica / Zoppi, Simona / Masotti, Chiara / Crescio, Maria Ines / Oggiano, Annalisa / Ercolini, Carlo / Razzuoli, Elisabetta

    Toxicology reports

    2022  Volume 9, Page(s) 171–180

    Abstract: ... Cadmium ( ... ...

    Abstract Cadmium (Cd
    Language English
    Publishing date 2022-01-29
    Publishing country Ireland
    Document type Journal Article
    ZDB-ID 2805786-7
    ISSN 2214-7500 ; 2214-7500
    ISSN (online) 2214-7500
    ISSN 2214-7500
    DOI 10.1016/j.toxrep.2022.01.009
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  3. Article ; Online: Lipoprotein(a) and family history of cardiovascular disease in children with familial dyslipidemias.

    Guardamagna, Ornella / Abello, Francesca / Anfossi, Giovanni / Pirro, Matteo

    The Journal of pediatrics

    2011  Volume 159, Issue 2, Page(s) 314–319

    Abstract: Objective: To investigate in children and adolescents with familial dyslipidemias the association between lipoprotein(a) [Lp(a)] level and family history of cardiovascular disease (CVD), and whether this association is independent of the disturbed lipid ...

    Abstract Objective: To investigate in children and adolescents with familial dyslipidemias the association between lipoprotein(a) [Lp(a)] level and family history of cardiovascular disease (CVD), and whether this association is independent of the disturbed lipid profile.
    Study design: Lp(a) level, lipid profile, and a 2-generation genealogic tree to detect cardiovascular events were evaluated in 231 patients with familial dyslipidemias. Lp(a) levels were stratified according to presence, age of occurrence, and number and type of cardiovascular events in the patient's kindreds.
    Results: Lp(a) and other plasma lipid fractions did not differ between patients with and those without a family history of cardiovascular events. However, the percentage of patients with elevated Lp(a) level (≥85th percentile) was higher in those with a positive family history for early cardiovascular events (P = .01). Lp(a) level was a significant independent predictor of the number of premature cardiovascular events (β = 0.17; P = .01) and of cerebrovascular events in kindreds (OR, 2.5; 95% CI, 1.05-6.03; P = .039), independent of plasma lipid fractions and other cardiovascular risk factors.
    Conclusions: In children and adolescents with familial dyslipidemias, the overall association between Lp(a) level and family history of early CVD may be due to a threshold effect in those with the highest Lp(a) levels. However, multiple cardiovascular events and cerebrovascular events are predicted by any increase in plasma Lp(a) level, independent of other cardiovascular risk factors.
    MeSH term(s) Adolescent ; Cardiovascular Diseases/blood ; Cardiovascular Diseases/epidemiology ; Cardiovascular Diseases/etiology ; Child ; Child, Preschool ; Dyslipidemias/blood ; Dyslipidemias/complications ; Dyslipidemias/genetics ; Female ; Genetic Predisposition to Disease ; Humans ; Incidence ; Italy/epidemiology ; Lipoprotein(a)/blood ; Male ; Prognosis ; Risk Factors
    Chemical Substances Lipoprotein(a)
    Language English
    Publishing date 2011-08
    Publishing country United States
    Document type Comparative Study ; Journal Article
    ZDB-ID 3102-1
    ISSN 1097-6833 ; 0022-3476
    ISSN (online) 1097-6833
    ISSN 0022-3476
    DOI 10.1016/j.jpeds.2011.01.038
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  4. Article ; Online: Comparative phenotypic and functional analyses of the effects of autologous plasma and recombinant human macrophage-colony stimulating factor (M-CSF) on porcine monocyte to macrophage differentiation.

    Franzoni, Giulia / Bonelli, Piero / Graham, Simon Paul / Anfossi, Antonio Giovanni / Dei Giudici, Silvia / Pilo, Giovannantonio / Pittau, Marco / Nicolussi, Paola / Oggiano, Annalisa

    Veterinary immunology and immunopathology

    2017  Volume 187, Page(s) 80–88

    Abstract: Porcine monocyte-derived macrophages (moMΦ) have been employed as a model cell in numerous studies of the porcine immune system. However, the lack of a standardized method for moMΦ differentiation hampers the comparison of results coming from the use of ... ...

    Abstract Porcine monocyte-derived macrophages (moMΦ) have been employed as a model cell in numerous studies of the porcine immune system. However, the lack of a standardized method for moMΦ differentiation hampers the comparison of results coming from the use of different laboratory protocols. In this study we compared the use of varying concentrations of autologous plasma (10, 20 and 30% v/v) or recombinant human macrophage-colony stimulating factor (hM-CSF; 50, 100, and 200ng/ml) to differentiate porcine monocytes into macrophages. Changes in cell morphology and surface marker expression were assessed by confocal microscopy and flow cytometry. Macrophage differentiation was evaluated by analysing TNF-α response to LPS stimulation and determining cytokine secretion patterns under both basal conditions and after classical and alternative activation. The effects of the differentiation methods on metabolic activity and susceptibility to infection with the myelotropic African swine fever virus (ASFV) were also evaluated. Monocytes cultured using the different culture conditions tested augmented in dimension and cellular complexity, but increasing porcine plasma concentrations resulted in a dose dependent enhancement in granularity and a marked pleomorphism. As expected, CD163, MHC class II DR and CD203a expression were up-regulated in both hM-CSF (M-CSF-moMΦ) and autologous plasma cultured macrophages (AP-moMΦ), although a lower percentage of CD163
    MeSH term(s) Animals ; Cell Differentiation/drug effects ; Macrophage Colony-Stimulating Factor/pharmacology ; Macrophages/drug effects ; Macrophages/physiology ; Monocytes/drug effects ; Monocytes/physiology ; Phenotype ; Recombinant Proteins ; Swine/immunology
    Chemical Substances Recombinant Proteins ; Macrophage Colony-Stimulating Factor (81627-83-0)
    Language English
    Publishing date 2017-05
    Publishing country Netherlands
    Document type Comparative Study ; Journal Article
    ZDB-ID 754160-0
    ISSN 1873-2534 ; 0165-2427
    ISSN (online) 1873-2534
    ISSN 0165-2427
    DOI 10.1016/j.vetimm.2017.04.006
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  5. Article ; Online: Ovis aries Papillomavirus 3 in Ovine Cutaneous Squamous Cell Carcinoma.

    Vitiello, Veronica / Burrai, Giovanni P / Agus, Mariagrazia / Anfossi, Antonio G / Alberti, Alberto / Antuofermo, Elisabetta / Rocca, Stefano / Cubeddu, Tiziana / Pirino, Salvatore

    Veterinary pathology

    2017  Volume 54, Issue 5, Page(s) 775–782

    Abstract: Squamous cell carcinoma (SCC) is a common malignancy affecting humans and other animals. Papillomaviruses (PVs) are frequently reported as causal agents of cutaneous benign and malignant epithelial lesions in different animal species, but only few ... ...

    Abstract Squamous cell carcinoma (SCC) is a common malignancy affecting humans and other animals. Papillomaviruses (PVs) are frequently reported as causal agents of cutaneous benign and malignant epithelial lesions in different animal species, but only few studies have investigated their role in ovine SCC. In this study, we explore the possible involvement of the Ovine aries PVs (OaPV1, OaPV2, OaPV3) in cutaneous SCC using an integrated histological and molecular approach. Forty cutaneous SCCs from different anatomical locations of Sardinian sheep and 40 matched non-SCC samples were evaluated histologically and by polymerase chain reaction (PCR) to assess the presence of ovine PVs. In addition, DNA in situ hybridization (ISH) and reverse transcription-polymerase chain reaction (RT-PCR) were carried out to evaluate the cellular localization and viral transcriptional activity, respectively. OaPV3 DNA was detected in 26 of 40 (65%) SCCs and in 12 of 40 (30%) non-SCC samples using PCR. OaPV1 and OaPV2 were not detected. OaPV3 viral DNA was observed by ISH in malignant epithelial squamous cells of 18 of 40 (45%) SCCs. In addition, the viral transcriptional activity was identified in 24 of 40 (60%) SCCs by RT-PCR. Notably, a higher viral positivity was observed in SCCs compared with non-SCC samples. The considerable infection rate of OaPV3 in the most common skin tumor of the sheep suggests that PV could represent a key factor in the onset of ovine SCC.
    Language English
    Publishing date 2017-09
    Publishing country United States
    Document type Journal Article
    ZDB-ID 188012-3
    ISSN 1544-2217 ; 0300-9858
    ISSN (online) 1544-2217
    ISSN 0300-9858
    DOI 10.1177/0300985817705171
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  6. Article: Pathophysiology of platelet resistance to anti-aggregating agents in insulin resistance and type 2 diabetes: implications for anti-aggregating therapy.

    Anfossi, Giovanni / Trovati, Mariella

    Cardiovascular & hematological agents in medicinal chemistry

    2006  Volume 4, Issue 2, Page(s) 111–128

    Abstract: The insulin resistance syndrome, which presents among its many facets obesity and type 2 diabetes mellitus, is a major risk factor for cardiovascular events. Thus, therapeutic guidelines recommend multifactorial treatment programs including, especially ... ...

    Abstract The insulin resistance syndrome, which presents among its many facets obesity and type 2 diabetes mellitus, is a major risk factor for cardiovascular events. Thus, therapeutic guidelines recommend multifactorial treatment programs including, especially in the presence of type 2 diabetes, antiplatelet drugs. Few data, however, are available about the protective effect of antiplatelet therapy in both obese and type 2 diabetic patients. Furthermore, some reports showed a decreased sensitivity to the platelet antiaggregating effect of acetylsalicylic acid in diabetic patients. In the first part of this review, we focused our attention to alterations of platelets from insulin resistant subjects with or without type 2 diabetes, underlining that platelet hyperactivation is explained, at least in part, by: i) a reduced sensitivity to agents exerting an inhibitory modulation of platelet responses, ii) an altered intracellular milieu with elevated cytosolic Ca2+, iii) an enhanced thromboxane A2 synthesis, and iv) an increased number and/or function of GPIIb/IIIa complexes on platelet membranes. Furthermore, oxidative stress, which increases isoprostane production from arachidonic acid, may be involved in platelet hyperactivation, since isoprostanes activate platelets by interplaying with thromboxane receptors. These defects explain why antiplatelet therapy for both chronic atherosclerotic vascular disease and acute coronary syndromes should be specifically tailored in obese, insulin resistant subjects, especially in the presence of type 2 diabetes mellitus. Thus, in the second part of this review we carried out a critical overview of the clinical trials in subjects with metabolic syndrome and type 2 diabetes mellitus with or without macroangiopathy.
    MeSH term(s) Blood Platelets/drug effects ; Blood Platelets/physiology ; Diabetes Mellitus, Type 2/blood ; Diabetes Mellitus, Type 2/physiopathology ; Humans ; Insulin Resistance ; Metabolic Syndrome/blood ; Metabolic Syndrome/physiopathology ; Platelet Aggregation Inhibitors/pharmacology ; Thrombosis/blood ; Thrombosis/physiopathology
    Chemical Substances Platelet Aggregation Inhibitors
    Language English
    Publishing date 2006-03-28
    Publishing country Netherlands
    Document type Journal Article ; Research Support, Non-U.S. Gov't ; Review
    ZDB-ID 2215710-4
    ISSN 1875-6182 ; 1871-5257
    ISSN (online) 1875-6182
    ISSN 1871-5257
    DOI 10.2174/187152506776369908
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  7. Article ; Online: Host cell tropism, genome characterization, and evolutionary features of OaPV4, a novel Deltapapillomavirus identified in sheep fibropapilloma.

    Tore, Gessica / Cacciotto, Carla / Anfossi, Antonio Giovanni / Dore, Gian Mario / Antuofermo, Elisabetta / Scagliarini, Alessandra / Burrai, Giovanni Pietro / Pau, Salvatore / Zedda, Maria Teresa / Masala, Gerolamo / Pittau, Marco / Alberti, Alberto

    Veterinary microbiology

    2017  Volume 204, Page(s) 151–158

    Abstract: Investigating papillomavirus (PV) diversity is crucial to fully comprehend pathogenicity, genetic features, and evolution of taxa hosted by domestic and wild animal species. This study reports the identification of OaPV4, a novel ovine PV type within ... ...

    Abstract Investigating papillomavirus (PV) diversity is crucial to fully comprehend pathogenicity, genetic features, and evolution of taxa hosted by domestic and wild animal species. This study reports the identification of OaPV4, a novel ovine PV type within Deltapapillomaviruses 3. The study of OaPV4 genomic features combined to in situ hybridization and immunohistochemistry investigations allowed extrapolating several general biological features of ovine PVs, such as their cellular tropism, pathogenicity, and evolutionary history. Based on results, ovine PVs can be grouped into a polyphyletic ancient group of viruses, which splits in two main subgroups having peculiar cellular tropism and pathogenicity. Results add up to animal PV diversity and are crucial to future studies aimed to investigate the correlation between animal PV and cutaneous benign and malign proliferations.
    Language English
    Publishing date 2017-05
    Publishing country Netherlands
    Document type Journal Article
    ZDB-ID 753154-0
    ISSN 1873-2542 ; 0378-1135
    ISSN (online) 1873-2542
    ISSN 0378-1135
    DOI 10.1016/j.vetmic.2017.04.024
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  8. Article ; Online: A short-term incubation with high glucose impairs VASP phosphorylation at serine 239 in response to the nitric oxide/cGMP pathway in vascular smooth muscle cells: role of oxidative stress.

    Russo, Isabella / Viretto, Michela / Doronzo, Gabriella / Barale, Cristina / Mattiello, Luigi / Anfossi, Giovanni / Trovati, Mariella

    BioMed research international

    2014  Volume 2014, Page(s) 328959

    Abstract: A reduction of the nitric oxide (NO) action in vascular smooth muscle cells (VSMC) could play a role in the vascular damage induced by the glycaemic excursions occurring in diabetic patients; in this study, we aimed to clarify whether a short-term ... ...

    Abstract A reduction of the nitric oxide (NO) action in vascular smooth muscle cells (VSMC) could play a role in the vascular damage induced by the glycaemic excursions occurring in diabetic patients; in this study, we aimed to clarify whether a short-term incubation of cultured VSMC with high glucose reduces the NO ability to increase cGMP and the cGMP ability to phosphorylate VASP at Ser-239. We observed that a 180 min incubation of rat VSMC with 25 mmol/L glucose does not impair the NO-induced cGMP increase but reduces VASP phosphorylation in response to both NO and cGMP with a mechanism blunted by antioxidants. We further demonstrated that high glucose increases radical oxygen species (ROS) production and that this phenomenon is prevented by the PKC inhibitor chelerythrine and the NADPH oxidase inhibitor apocynin. The following sequence of events is supported by these results: (i) in VSMC high glucose activates PKC; (ii) PKC activates NADPH oxidase; (iii) NADPH oxidase induces oxidative stress; (iv) ROS impair the signalling of cGMP, which is involved in the antiatherogenic actions of NO. Thus, high glucose, via oxidative stress, can reduce the cardiovascular protection conferred by the NO/cGMP pathway via phosphorylation of the cytoskeleton protein VASP in VSMC.
    MeSH term(s) Acetophenones/pharmacology ; Animals ; Antioxidants/metabolism ; Benzophenanthridines/pharmacology ; Cell Adhesion Molecules/metabolism ; Cells, Cultured ; Cyclic GMP/metabolism ; Glucose/metabolism ; Male ; Microfilament Proteins/metabolism ; Muscle, Smooth, Vascular/metabolism ; Myocytes, Smooth Muscle/metabolism ; NADPH Oxidases/antagonists & inhibitors ; NADPH Oxidases/metabolism ; Nitric Oxide/metabolism ; Oxidative Stress/physiology ; Phosphoproteins/metabolism ; Phosphorylation/physiology ; Protein Kinase C/antagonists & inhibitors ; Protein Kinase C/metabolism ; Rats ; Rats, Zucker ; Reactive Oxygen Species/metabolism ; Serine/metabolism
    Chemical Substances Acetophenones ; Antioxidants ; Benzophenanthridines ; Cell Adhesion Molecules ; Microfilament Proteins ; Phosphoproteins ; Reactive Oxygen Species ; vasodilator-stimulated phosphoprotein ; Nitric Oxide (31C4KY9ESH) ; Serine (452VLY9402) ; acetovanillone (B6J7B9UDTR) ; chelerythrine (E3B045W6X0) ; NADPH Oxidases (EC 1.6.3.-) ; Protein Kinase C (EC 2.7.11.13) ; Cyclic GMP (H2D2X058MU) ; Glucose (IY9XDZ35W2)
    Language English
    Publishing date 2014-03-23
    Publishing country United States
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2698540-8
    ISSN 2314-6141 ; 2314-6133
    ISSN (online) 2314-6141
    ISSN 2314-6133
    DOI 10.1155/2014/328959
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  9. Article ; Online: Contribution of insulin resistance to vascular dysfunction.

    Anfossi, Giovanni / Russo, Isabella / Doronzo, Gabriella / Trovati, Mariella

    Archives of physiology and biochemistry

    2009  Volume 115, Issue 4, Page(s) 199–217

    Abstract: Insulin is a vascular hormone, able to influence vascular cell responses. In this review, we consider the insulin actions on vascular endothelium and on vascular smooth muscle cells (VSMC) both in physiological conditions and in the presence of insulin ... ...

    Abstract Insulin is a vascular hormone, able to influence vascular cell responses. In this review, we consider the insulin actions on vascular endothelium and on vascular smooth muscle cells (VSMC) both in physiological conditions and in the presence of insulin resistance. In particular, we focus the relationships between activation of insulin signalling pathways of phosphatidylinositol-3 kinase (PI3-K) and mitogen-activated protein kinase (MAPK) and the different vascular actions of insulin, with a particular attention to the insulin ability to activate the pathway nitric oxide (NO)/cyclic GMP/PKG via PI3-K, owing to the peculiar relevance of NO in vascular biology. We also discuss the insulin actions mediated by the MAPK pathway (such as endothelin-1 synthesis and secretion and VSMC proliferation and migration) and by the interactions between the two pathways, both in insulin-sensitive and in insulin-resistant states. Finally, we consider the influence of free fatty acids, cytokines and endothelin on vascular insulin resistance.
    MeSH term(s) Blood Vessels/enzymology ; Blood Vessels/metabolism ; Blood Vessels/physiopathology ; Humans ; Insulin/metabolism ; Insulin/physiology ; Insulin Resistance ; Mitogen-Activated Protein Kinases/metabolism ; Phosphatidylinositol 3-Kinases/metabolism ; Signal Transduction
    Chemical Substances Insulin ; Phosphatidylinositol 3-Kinases (EC 2.7.1.-) ; Mitogen-Activated Protein Kinases (EC 2.7.11.24)
    Language English
    Publishing date 2009-10
    Publishing country England
    Document type Journal Article ; Research Support, Non-U.S. Gov't ; Review
    ZDB-ID 1238320-x
    ISSN 1744-4160 ; 1381-3455
    ISSN (online) 1744-4160
    ISSN 1381-3455
    DOI 10.1080/13813450903136791
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  10. Article ; Online: The cardiovascular effects of metformin: further reasons to consider an old drug as a cornerstone in the therapy of type 2 diabetes mellitus.

    Anfossi, Giovanni / Russo, Isabella / Bonomo, Katia / Trovati, Mariella

    Current vascular pharmacology

    2009  Volume 8, Issue 3, Page(s) 327–337

    Abstract: Cardiovascular events occurring in type 2 diabetes (T2DM) are a major problem in clinical practice. In particular, the risk of myocardial infarction (MI) presented by patients affected by T2DM without previous cardiac events is similar to that of non- ... ...

    Abstract Cardiovascular events occurring in type 2 diabetes (T2DM) are a major problem in clinical practice. In particular, the risk of myocardial infarction (MI) presented by patients affected by T2DM without previous cardiac events is similar to that of non-diabetic patients with previous MI. To reduce the elevated cardiovascular risk associated with T2DM, tight glycemic control and aggressive therapy against all known cardiovascular risk factors are strictly required. Despite the role played by hyperglycemia in the pathogenesis of cardiovascular events, studies showing an improvement of cardiovascular outcomes by anti-hyperglycemic or hypoglycemic agents are not conclusive. The United Kingdom Prospective Diabetes Study (UKPDS) demonstrated that in obese type 2 diabetic patients metformin reduces the risk of MI more than sulphonylureas or insulin. This observation identified metformin as the first-line treatment for T2DM. The vasoprotective role of metformin is largely independent of its hypoglycemic action and has been ascribed to pleiotropic effects. The present review considers the putative beneficial action exerted by metformin on arterial vessels by evaluating its effects on lipids, inflammation, hemostasis, endothelial and platelet function and vessel wall abnormalities. Furthermore, the molecular mechanisms of the beneficial metabolic and vascular effects of metformin will be considered, with a particular attention for its ability to activate AMP-activated protein kinase.
    MeSH term(s) Animals ; Cardiovascular Diseases/drug therapy ; Cardiovascular Diseases/etiology ; Diabetes Mellitus, Type 2/complications ; Diabetes Mellitus, Type 2/drug therapy ; Humans ; Hypoglycemic Agents/therapeutic use ; Metformin/therapeutic use ; Prospective Studies ; Risk Factors
    Chemical Substances Hypoglycemic Agents ; Metformin (9100L32L2N)
    Language English
    Publishing date 2009-04-01
    Publishing country United Arab Emirates
    Document type Journal Article ; Research Support, Non-U.S. Gov't ; Review
    ZDB-ID 2192362-0
    ISSN 1875-6212 ; 1570-1611
    ISSN (online) 1875-6212
    ISSN 1570-1611
    DOI 10.2174/157016110791112359
    Database MEDical Literature Analysis and Retrieval System OnLINE

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