Diabete mellito, aspetti biochimico-molecolari

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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

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Rischi associati al diabete mellito

malattia rischio rispettoai non diabetici

Cecità 20 volte

Insufficienza renale 25 volte

Amputazione 40 volte

Infarto miocardico 2 – 5 volte

Ictus 2 – 3 volte

Nathan, N Engl J Med 1993

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14.217.523%

15.622.544%

26.532.924% 84.5

132.3

57%

9.4

14.150%

1.0

1.3

33%2000: 151 milioni 2010: 221 milioni Incremento 46 %

Prevalenza e incremento dei diabetici nel mondo

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Epidemiologia

Prevalenza in Italia: circa 2 milioni (3,5 %)

Tipo 1: circa 5 %

Tipo 2: circa 90 %

Incidenza Nord-Italia: 5-6/100.000 nuovi casi/anno

Prevalenza nel mondo

2001: circa 140.000.000

2025: circa 300.000.000

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Frequenza del diabete e dell’intolleranza al glucosio in funzione dell’età

età diabetediagnosticato

diabetenon diagnosticato

intolleranza alglucosio

45 – 54 3,8 1,3 4,4

55 – 64 9,5 1,8 6,4

65 – 74 10,0 5,0 10,0

oltre i 75 11,3 5,0 19,4

Garancini et al, 1995

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t.adiposo

fegato

t.adiposo

muscolo

GLUCOSIO

GLUCOSIO

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

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Segni, sintomi e conseguenze dell’ipoglicemia

0102030405060708090

Morte

Danni cerebrali permanentiConvulsioni

Coma

Letargia

Sintomi da neuroglicopenia

Controregolazione

Sfumata sintomatologia neurologica

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Diabete tipo 1, sviluppo

D is tru z ion e ce llu la reT-c ito toss ic im ac ro fag i

B -lin foc it i (A b )

p resen taz ion e A gce llu le T-h e lp er

E sp ress ion e d i an tig en im em b ran e b e ta ce llu le

G en i d i su sce tt ib ilità(fa tto ri am b ien ta li)

Markers- antigeni HLA classe II DR3, DR4 (DR2: protettivo)- anticorpi anticellule- anticorpi antiinsulina

induzione

attivazione e "homing" sulle cellule

distruzione

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LOSS OF FIRST PHASE LOSS OF FIRST PHASE INSULIN RESPONSE INSULIN RESPONSE

TIMETIME

Stages in Development of Type 1 DiabetesStages in Development of Type 1 Diabetes B

ET

A C

EL

L M

AS

SB

ET

A C

EL

L M

AS

S

DIABETES

“PRE”-DIABETES

GENETICPREDISPOSITION

INSULITISBETA CELL INJURY

NEWLY DIAGNOSED DIABETES

MULTIPLE ANTIBODY MULTIPLE ANTIBODY POSITIVEPOSITIVE

GENETICALLY AT RISK

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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

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Possibile patogenesi del Diabete di tipo 2

C aren za re la tiva d i in su lin a

D ife tto d e l g lu corece tto re

R id o tta m assab e ta ce llu la re

IP E R G L IC E M IA

In su lin o res is ten za

Ip erin su lin em ia

O b es ità

Ip e ra lim en taz ion e

Ambiente

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Diabete mellito (classificazione eziologica) I. Diabete di tipo 1 (caratterizzato da distruzione delle -cellule,

solitamente comportante un deficit assoluto di insulina) A. Immuno-mediato B. Idiopatico (LADA)

II. Diabete di tipo 2 (può variare da predominantemente insulino-resistente e relativamente insulino-deficente, a predominantemente insulino-deficente e relativamente poco insulino-resistente)

III. Altri tipi specifici A. Difetti genetici della funzionalità -cellulare B. Difetti genetici dell’azione insulinica C. Malattie del pancreas esocrino D. Endocrinopatie E. Malattie indotte da farmaci o sostanze chimiche F. Infezioni G. Rare forme di diabete immuno-mediato H. Altre sindromi genetiche a volte associate al diabete

IV. Diabete mellito gestazionale

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Caratteristiche cliniche del LADA(Latent Autoimmune Diabetes of the Adult)

• Prevalenza: circa il 10 % del diabete dell’adulto

• Età d’esordio generalmente superiore ai 35 anni

• Quadro d’esordio lento od attenuato

• Sviluppo graduale di insulino-dipendenza

• Frequente presenza di anticorpi antiGAD

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• 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

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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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Screening del diabete mellito in soggetti presumibilmente sani. Su tutti gli adulti al di sopra dei 45 anni di età glicemia a digiuno ogni 3 anni, a meno che il soggetto abbia già una diagnosi di diabete mellito. La misura della glicemia a digiuno si deve fare in età inferiore e con cadenza più ravvicinata nei soggetti che presentino: Obesità (? 120 % del peso corpoeo desiderabile o indice di massa

corporeo ? 27 kg/m2) Parentela di primo grado con un paziente diabetico Appartenenza ad un gruppo etnico ad alto rischioa Precedente esperienza di diabete gestazionale o parto di neonato

sovrapeso (> 4.1 kg) Ipertensione (? 140/90) Bassa concentrazione del colesterolo-HDL nel siero (< 35 mg/dL) Elevata concentrazione di trigliceridi nel siero (> 200 mg/dL) Precedente storia di IFG od IGT

a Afro-americani, ispano-americani, nativi americani, asio-americani

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Glicemia - variazioni giornaliere

M. Luzzana, 1999

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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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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

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trasportatori del glucosio

GLUT1: RBC/epatociti 492 aa 1p35-31.3

GLUT2: -cell/fegato 524 aa 3q26

GLUT3: cervello 496 aa 12p13

GLUT4: insulin-resp. 509 aa 17p13

GLUT5: intestino 501 aa 1p31

GLUT6: pseudogene

GLUT7: reticolo endoplasmico epatociti

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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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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

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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.

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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).

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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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pp-120IRS-1IRS-2IRS-3IRS-4

Glut-4

Prot SH2PI-3 chinasi

MAPKPKBSgK

Trasduzione del segnale insulinico

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AM-UniMi 49Kulkarni, Science 2004

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

AM-UniMi 51J. Risley, 1999 - http://opbs.okstate.edu/

1370aa2, 2 NPEY motif

> 60 mutazioni note

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

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PhenotypeNucleotide

substit.Micro-lesions

Gross lesions

Leprechaunism 19 5 2

Insulin resistance 25 3 0

Insulin resistance A 3 0 0

Diabetes, NIDDM 6 0 0

Fibre-type disproportion myopathy, congenital

1 0 0

Acanthosis nigricans 0 0 1

Acanthosis nigricans, insulin related 0 0 1

Mutations in this gene were first reported in 1988

Kadowaki (1988) Science 240, 787Yoshimasa (1988) Science 240, 784

Moller (1988) N Engl J Med 319, 1526

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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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A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115

CollageneCristallinoAlbuminaEmoglobinaLDL

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A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115

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A. Lapolla et al. / Clinical Biochemistry 38 (2005) 103–115

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Possibile utilizzo degli AGE

• Misura del controllo glico-metabolico a lungo termine– “early glycation”

• Glicoalbumina

• HbA1c

– ”intermediate glycation”• Metil-gliossale

• Misura del grado di modificazioni tissutali in relazione al rischio delle complicanze– AGE

• Pentosidina “libera”

• Misura effetto terapie

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Alcuni prodotti di glicazione intermedi e tardivi (AGE)

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

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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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… 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 rats96, and vitamin C improved endothelium-dependent vasodilation in diabetic patients97, low-dose vitamin E failed to alter the risk of cardiovascular and renal disease in patients with diabetes…

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Marcatori genetici del diabete di tipo 2

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Cambiamenti morofologici nel pancreas in diverse condizioni

Rhodes CJ, Science 2005;307

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Modello dinamico per i cambiamenti della massa beta cellulare

t1/2 delle beta-cell: 60 d

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Fattori che scatenano l’apoptosi delle beta-cellule nel DM t2

• Iperglicemia cronica

• Iperlipidemia cronica

• Stress ossidativo

• Diverse citochine

• Sviluppo di stress del reticolo endoplasmico

• Alterazioni di IRS-2

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Meccanismi potenziali che scatenano la degradazione di IRS-2 e l’apoptosi delle beta-cellule

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IRS-2 ha importanza rilevante nel pathway insulinico nei tessuti

insulino-responsivi.

Una sua diminuzione causa insulino-resistenza.

C’è correlazione tra i meccanismi molecolari che controllano la sensibilità insulinica e quelli che promuovono la sopravvivenza β cellulare.

…CONCLUDENDO:

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• 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 (PPAR2: peroxisome-proliferator-activated receptor)

• Riduzione della attivazione della eNOS

Insulino-resistenza

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Some developing countries face the paradox of families in which thechildren are underweight and the adults are overweight. This combination has been attributed by some people to intrauterine growthretardation and resulting low birth weight, which apparently confera predisposition to obesity later in life through the acquisition ofa “thrifty” phenotype that, when accompanied by rapid childhoodweight gain, is conducive to the development of insulin resistanceand the metabolic syndrome.

n engl j med 356;3 www.nejm.org january 18, 2007

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O’Rahilly, Science 2005

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Loci associati senzaevidenze 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-1HNF-3HNF-4Insulina

IRS-1

ISL-1 NEUROD1

NIDDM2

NOsintasi endoteliale

Promotore dell’insulina (fattore 1)

Protein-fosfatasi 1 (subunità muscolo-specifica)

Ras associato a diabete

Recettore 3-adrenergico

Recettore del glucagone

Recettore della sulfonilurea

Recettore dell’insulina

Recettore tipo B della colecistochinina

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References recenti

Raeder et al. Mutations in the CEL VNTR causes a syndromeof diabetes and pancreatic exocrine disfunction. Nature Genetics 2006;38:54-62

Grant et al. Variant of transcription 7-like 2 (TCF7L2) geneconfers risk of type 2 diabetes. Nature Genetics 2006.

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