Diabete mellito, aspetti biochimico-molecolari. AM-UniMi 2 Diabete mellito Un gruppo eterogeneo di malattie caratterizzate da un metabolismo anormale.

Presentazione sul tema: "Diabete mellito, aspetti biochimico-molecolari. AM-UniMi 2 Diabete mellito Un gruppo eterogeneo di malattie caratterizzate da un metabolismo anormale."— Transcript della presentazione:

1 Diabete mellito, aspetti biochimico-molecolari

2 AM-UniMi 2 Diabete mellito Un gruppo eterogeneo di malattie caratterizzate da un metabolismo anormale dei CARBOIDRATI, causato da un DEFICT DI INSULINA assoluto (tipo 1) o relativo (tipo 2), che provoca IPERGLICEMIA

3 AM-UniMi 3 Rischi associati al diabete mellito malattiarischio rispetto ai non diabetici Cecità20 volte Insufficienza renale25 volte Amputazione40 volte Infarto miocardico2 – 5 volte Ictus2 – 3 volte Nathan, N Engl J Med 1993

4 AM-UniMi 4 14.2 17.5 23 % 15.6 22.5 44 % 26.5 32.9 24 % 84.5 132.3 57 % 9.4 14.1 50 % 1.0 1.3 33 % 2000: 151 milioni 2010: 221 milioni Incremento 46 % Prevalenza e incremento dei diabetici nel mondo

5 AM-UniMi 5 Epidemiologia Prevalenza in Italia:circa 2 milioni (3,5 %) Tipo 1: circa 5 % Tipo 2: circa 90 % IncidenzaNord-Italia: 5-6/100.000 nuovi casi/anno Prevalenza nel mondo 2001: circa 140.000.000 2025: circa 300.000.000

6 AM-UniMi 6 Frequenza del diabete e dellintolleranza al glucosio in funzione delletà etàdiabete diagnosticato diabete non diagnosticato intolleranza al glucosio 45 – 543,81,34,4 55 – 649,51,86,4 65 – 7410,05,010,0 oltre i 7511,35,019,4 Garancini et al, 1995

7 AM-UniMi 7 t.adiposo fegato t.adiposo muscolo GLUCOSIO

8 AM-UniMi 8 Euglicemia: Glucosio 3,5-6,0 mmol/L

9 AM-UniMi 9 Segni, sintomi e conseguenze dellipoglicemia Morte Danni cerebrali permanenti Convulsioni Coma Letargia Sintomi da neuroglicopenia Controregolazione Sfumata sintomatologia neurologica

10 AM-UniMi 10 Diabete tipo 1, sviluppo Markers - antigeni HLA classe II DR3, DR4 (DR2: protettivo) - anticorpi anticellule - anticorpi antiinsulina induzione attivazione e "homing" sulle cellule distruzione

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12 AM-UniMi 12 LOSS OF FIRST PHASE INSULIN RESPONSE TIME Stages in Development of Type 1 Diabetes BETA CELL MASS DIABETES PRE- DIABETES GENETIC PREDISPOSITION INSULITIS BETA CELL INJURY NEWLY DIAGNOSED DIABETES MULTIPLE ANTIBODY POSITIVE GENETICALLY AT RISK

13 AM-UniMi 13 Portatori di alleli HLA DR3-DR4: rischio 6-7 % Parente di I grado (senza conoscere genotipo): rischio 3-6 % Parente di I grado (con assetto genetico noto):rischio 6-16 % Diabete di tipo 1

14 AM-UniMi 14 Possibile patogenesi del Diabete di tipo 2 Ambiente

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16 AM-UniMi 16 Caratteristiche cliniche del LADA (Latent Autoimmune Diabetes of the Adult) Prevalenza: circa il 10 % del diabete delladulto Età desordio generalmente superiore ai 35 anni Quadro desordio lento od attenuato Sviluppo graduale di insulino-dipendenza Frequente presenza di anticorpi antiGAD

17 AM-UniMi 17 Prevalenza: oltre 5 % delle gravidanze Definizione: Intolleranza ai CHO di vario grado e severità, con inizio o primo riscontro durante la gravidanza Screening: OGCT (50g) Diagnosi: OGTT (100 o 75g) Classificazione dopo il parto: NGT, IFG, IGT, Diabete Diabete Gestazionale Lapolla, 2001

18 AM-UniMi 18 Altre condizioni patologiche a rischio di evoluzione a diabete mellito Ridotta tolleranza al glucosio (IGT: impaired glucose tolerance) Alterata glicemia a digiuno (IFG: impaired fasting glucose)

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20 AM-UniMi 20 Glicemia - variazioni giornaliere M. Luzzana, 1999

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23 AM-UniMi 23 Sticking to membranes. Hexokinases associate with various cellular membranes, and this association affects their activity. These enzymes are not only involved in glucose sensing and metabolism but also in signal transduction. This duality is achieved by switching between a bound and unbound form that interacts with different proteins, such as regulatory DNA-protein complexes in the nucleus. Receptors for hexokinases (purple) must be present to enable differential targeting of these enzymes to different subcellular locations. Hexokinases associate with membranes of subcellular compartments, such as the endoplasmic reticulum (ER) and mitochondria. Frommer et al, Science 2003

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30 AM-UniMi 30 Purpose of review Glucose homeostasis must be finely regulated. Changes in glucose levels elicit a complex neuroendocrine response that prevents or rapidly corrects hyper- or hypoglycemia. It is well established that different parts of the brain, particularly the hypothalamus and the brain stem, are important centres involved in the monitoring of glucose status and the regulation of feeding. The pioneering work of Mayer, including his proposal of the glucostatic theory, has recently received experimental support from the molecular, electro-physiological and physiological fields. Recent findings Making the analogy with the cell of the islet of Langerhans, it has been proposed that glucose sensing could be assured in some cells of the brain by proteins such as glucose transporter 2, glucokinase and the ATP-dependent potassium channel. Furthermore, some pathological conditions such as diabetes and obesity have been shown to alter this glucose sensing system. Summary These findings could lead to a better understanding of metabolic disorders, with hypoglycemia possibly being the most deleterious.Brain glucose sensing mechanism and glucose homeostasis. Brain glucose sensing mechanism and glucose homeostasis Luc Pénicaud, Corinne Leloup, Anne Lorsignol, Thierry Alquier and Elise Guillod Current Opinion in Clinical Nutrition and Metabolic Care 2002, 5:539±543

31 AM-UniMi 31 trasportatori del glucosio GLUT1: RBC/epatociti492 aa1p35-31.3 GLUT2: -cell/fegato524 aa3q26 GLUT3: cervello496 aa12p13 GLUT4: insulin-resp.509 aa17p13 GLUT5: intestino501 aa1p31 GLUT6: pseudogene GLUT7: reticolo endoplasmico epatociti

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33 AM-UniMi 33 Sci. STKE, Vol. 2003, Issue 169, pp. pe5, 11 February 2003 A Long Search for Glut4 Activation Konstantin V. Kandror * Boston University School of Medicine, Boston, MA 02118, USA. Summary: Insulin stimulates glucose transport in its target cells by translocation of the glucose transporter isoform 4 (Glut4) from an intracellular storage pool to the plasma membrane. A large body of evidence indicates that activity of Glut4 at the plasma membrane may vary. Recent findings suggest that p38 MAPK may be involved in regulation of the intrinsic activity of the transporter.

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39 AM-UniMi 39 Figure 1 (A) Schematic representation of human PG. Tissue-specific post-translational processing of PG in the pancreas (B) and small intestine (C). The numbers indicate positions of amino acid residues and cleavage sites. Relative presence of glucagon and GLP-1 derived from the post-translational processing of preproglucagon molecule in the pancreas (D) and small intestine (E). In the pancreas, PG is cleaved to produce GRPP, glucagon, IP-1 and MPGF. All of these products are present in approximately equimolar amounts and are secreted synchronously. In addition to these predominant products, small amounts of a peptide corresponding to the GLP-1 domain are also formed. This molecule, which is probably biologically inactive, corresponds to PG(72±107), but small amounts of PG(72±108) are also formed. GLP-1 regulates glucose homeostasis 719 EUROPEAN JOURNAL OF ENDOCRINOLOGY (2000) 143 www.eje.org

40 AM-UniMi 40 Figure 2 Schematic representation of GLP-1 action on target tissues. The role of GLP-1 on muscle and adipose tissues is represented with a question mark next to the proposed enhancement of insulin sensitivity based on the yet controversial findings reported.

41 AM-UniMi 41 SEVERAL BIOLOGICAL FEATURES of glucagon-like peptide 1 (GLP-1) have led to propose this peptide hormone as an ideal candidate for the treatment of diabetes(1). GLP-1 lowers postprandial hyperglycemia via three independent mechanisms: increases insulin secretion, inhibits glucagon release, and inhibits gastrointestinal motility. Perhaps even more important is the observation that the insulin secretory action of GLP-1 is regulated by the plasma concentration of glucose, virtually preventing the possibility of developing reactive hypoglycemia while inducing the release of insulin (2). Finally, it is of significant clinical relevance the observation that GLP-1 retains its glucose lowering activity in patients with diabetes, even many years after clinical onset of the disease, when islet -cells are no longer responsive to other pharmacological insulin-secreting agents (3). secretion. Indeed, GLP-1 also affects the expression of insulin and other -cell-specific genes whose products are involved in the regulation of glucose utilization (4, 5). The mechanism by which GLP-1 modulates the -cell- specific gene expression has only in part been elucidated, and it is known to require the activation of the homeodomain transcription factor IDX-1 (6).

42 AM-UniMi 42 Endocrinology 2003;144(12):5149– 5158 FIG. 1. Islets cell morphology. Human islets were cultured for 1, 3, and 5 d in M199 medium, with 6 mM glucose, 10% FCS, and 0.1 mM diprotin-A and in the presence, or absence, of GLP-1 (10 nM, added every 12 h). Human islets on d 1, control (A) and GLP-1-treated islets (B); d 3, control (C) and GLP-1 treated (D); and d 5, control (E) and GLP-1 treated (F). Pictures are representative of islets morphology as observed by culturing human islets from three independent donors.

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44 AM-UniMi 44 Trasduzione segnale insulinico

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46 AM-UniMi 46 pp-120 IRS-1 IRS-2 IRS-3 IRS-4 Glut-4 Prot SH2 PI-3 chinasi MAPK PKB SgK Trasduzione del segnale insulinico

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49 AM-UniMi 49 Kulkarni, Science 2004

50 AM-UniMi 50 J. Risley, 1999 - http://opbs.okstate.edu/

51 AM-UniMi 51 J. Risley, 1999 - http://opbs.okstate.edu/ 1370aa 2, 2 NPEY motif > 60 mutazioni note

52 AM-UniMi 52 J. Risley, 1999 - http://opbs.okstate.edu/

53 AM-UniMi 53 Phenotype Nucleotide substit. Micro- lesions Gross lesions Leprechaunism1952 Insulin resistance2530 Insulin resistance A300 Diabetes, NIDDM600 Fibre-type disproportion myopathy, congenital 100 Acanthosis nigricans001 Acanthosis nigricans, insulin related001 Mutations in this gene were first reported in 1988 Kadowaki (1988) Science 240, 787 Yoshimasa (1988) Science 240, 784 Moller (1988) N Engl J Med 319, 1526

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55 AM-UniMi 55 Basi biochimiche delle complicanze diabetiche

56 AM-UniMi 56 Brownlee M. Biochemistry and molecular cell biology of diabetic complications. Nature. 414(6865):813-20, 2001 Dec 13 Diabetes-specific microvascular disease is a leading cause of blindness, renal failure and nerve damage, and diabetes- accelerated atherosclerosis leads to increased risk of myocardial infarction, stroke and limb amputation. Four main molecular mechanisms have been implicated in glucose-mediated vascular damage. All seem to reflect a single hyperglycaemia-induced process of overproduction of superoxide by the mitochondrial electron-transport chain. This integrating paradigm provides a new conceptual framework for future research and drug discovery.

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58 AM-UniMi 58 A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115 Collagene Cristallino Albumina Emoglobina LDL

59 AM-UniMi 59 A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115

60 AM-UniMi 60 A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115

61 AM-UniMi 61 Possibile utilizzo degli AGE Misura del controllo glico-metabolico a lungo termine –early glycation Glicoalbumina HbA 1c –intermediate glycation Metil-gliossale Misura del grado di modificazioni tissutali in relazione al rischio delle complicanze –AGE Pentosidina libera Misura effetto terapie

62 AM-UniMi 62 Alcuni prodotti di glicazione intermedi e tardivi (AGE)

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64 AM-UniMi 64 Administering the soluble form of the glycation product receptor seems to stop the accelerated atherosclerosis. Nature 4: 1025, 1998.

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67 AM-UniMi 67 Ruboxistaurin is an orally active, specific PKC inhibitor which seems to be well tolerated and normalises retinal blood flow in diabetic patients with retinopathy.

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71 AM-UniMi 71 … interrupting the overproduction of superoxide by the mitochondrial electron-transport chain would normalize polyol pathway flux, AGE formation, PKC activation, hexosamine pathway flux and NF- B activation. But it might be difficult to accomplish this using conventional antioxidants, as these scavenge reactive oxygen species in a stoichiometric manner. Thus, although long-term administration of a multi- antioxidant diet inhibited the development of early diabetic retinopathy in rats 96, and vitamin C improved endothelium- dependent vasodilation in diabetic patients 97, low-dose vitamin E failed to alter the risk of cardiovascular and renal disease in patients with diabetes… 96 97

72 AM-UniMi 72 Marcatori genetici del diabete di tipo 2

73 AM-UniMi 73 Cambiamenti morofologici nel pancreas in diverse condizioni Rhodes CJ, Science 2005;307

74 AM-UniMi 74 Modello dinamico per i cambiamenti della massa beta cellulare t 1/2 delle beta-cell: 60 d

75 AM-UniMi 75 Fattori che scatenano lapoptosi delle beta- cellule nel DM t2 Iperglicemia cronica Iperlipidemia cronica Stress ossidativo Diverse citochine Sviluppo di stress del reticolo endoplasmico Alterazioni di IRS-2

76 AM-UniMi 76 Meccanismi potenziali che scatenano la degradazione di IRS-2 e lapoptosi delle beta-cellule

77 AM-UniMi 77 IRS-2 ha importanza rilevante nel pathway insulinico nei tessuti insulino-responsivi. Una sua diminuzione causa insulino-resistenza. Cè correlazione tra i meccanismi molecolari che controllano la sensibilità insulinica e quelli che promuovono la sopravvivenza β cellulare. …CONCLUDENDO:

78 AM-UniMi 78 Tutti i geni di proteine coinvolte nella trasduzione del segnale insulinico sono potenzialmente coinvolti –Mutazioni del recettore –Varianti IRS-1 –Mutazioni PI-3 chinasi (?) –Ridotta attivazione della via IR/IRS/PI3K Mutazioni di altri fattori (PPAR 2: peroxisome-proliferator-activated receptor) Riduzione della attivazione della eNOS Insulino-resistenza

79 AM-UniMi 79 Some developing countries face the paradox of families in which the children are underweight and the adults are overweight. This combination has been attributed by some people to intrauterine growth retardation and resulting low birth weight, which apparently confer a predisposition to obesity later in life through the acquisition of a thrifty phenotype that, when accompanied by rapid childhood weight gain, is conducive to the development of insulin resistance and the metabolic syndrome. n engl j med 356;3 www.nejm.org january 18, 2007

80 AM-UniMi 80 ORahilly, Science 2005

81 AM-UniMi 81 Loci associati senza evidenze contraddittorie Apolipoproteina A1 Apolipoproteina A2 Apolipoproteina A4 Apolipoproteina B Apolipoproteina C3 Calpaina Convertasi 2 D11S935 (cromosoma 11) Lipasi ormono-sensibile Ormone della crescita PEP carbossichinasi Proteina C-reattiva Recettore 2 -adrenergico Loci controversi Amilina D1S191 (cromosoma 1) D20S197 (cromosoma 20q) Glicogeno sintasi Glucochinasi GLUT-2 HNF-1 HNF-3 HNF-4 Insulina IRS-1 ISL-1 NEUROD1 NIDDM2 NOsintasi endoteliale Promotore dellinsulina (fattore 1) Protein-fosfatasi 1 (subunità muscolo-specifica) Ras associato a diabete Recettore 3 -adrenergico Recettore del glucagone Recettore della sulfonilurea Recettore dellinsulina Recettore tipo B della colecistochinina

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83 AM-UniMi 83 References recenti Raeder et al. Mutations in the CEL VNTR causes a syndrome of diabetes and pancreatic exocrine disfunction. Nature Genetics 2006;38:54-62 Grant et al. Variant of transcription 7-like 2 (TCF7L2) gene confers risk of type 2 diabetes. Nature Genetics 2006.

84 AM-UniMi 84 Frayling T. Nature Review Genetics September 2007