National Institute of Pharmaceutical Education and Research, Hyderabad

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ENDOPLASMIC RETICULUM STRESS IN DIABETIC COMPLICATIONS

Diabetes is among the most occurring metabolic disorders.

Insulin injection other therapy have been established.

Need of more cellular targets

ER stress have link with other organ dysfunction

Targeting ER stress is among the upcoming novel targets for diabetic therapy

Non communicable diseases deaths are projected to increase by 15% globally between 2010 and 2020.

Transportation of proteins and other carbohydrates to another organelle

Manage enzymatic processes

Various post-translational modifications, including glycosylation and disulphide bond formation

Secretory proteins folding, modification, and assembling into multiprotein complexes

Vital role in the synthesis of proteins, lipids, glycogen and steroids.

Meusser B, Hirsch C, Jarosch E, Sommer T. 2011. ERAD & autophagy: the long road to destruction. Nat. Cell Biol. 7:766–72

ERAD(ER-associated degradation):Sometimes protein-folding machines fraction of secretory proteins normally fails to fold properly in the ER

Secretory proteins have crucial signalling roles ( cell surface receptors or polypeptide hormones)

Incompletely folded forms are not tolerated

Unfolded proteins are removed to the cytosol for subsequent ubiquitination and degradation by the 26S proteasome

AutophagyAnother mechanism for disposing unfolded secretory proteins.Under normal condition normal proteins are secreted.

Depletion of SPP1 and increased intracellular S1P triggers ER stress and the UPR by activation of ATF6, IRE1α, and PERK, resulting in autophagy.

PERK signaling also activates Akt, which protect cells from apoptosis

Meusser B, Hirsch C, Jarosch E, Sommer T. 2011. ERAD & autophagy: the long road to destruction. Nat. Cell Biol. 7:766–72

Conditions whereby cell encounter environmental challenges during which protein-folding demand in the ER exceeds capacity.Imbalanced state of secretory proteins Secretory proteins start to accumulate in incompletely modified and unfolded forms at high levels within the ER.

Mutant forms of secretory proteins causes overproduction of

secretory proteins itself generate ER stress

Involves four distinct processes:

Increase in the protein folding capacity through transcriptional induction of ER chaperones

Reduction of the biosynthetic load by translational attenuation

Degradation of the misfolded proteins by ERAD

Apoptosis, at last to eliminate infected cells and maintain homeostasis.

Translational attenuation

Translational attenuation

Degradation by ERAD

Degradation by ERAD

Apoptosis

Unfolded Protein

Transcriptional activation(Chaperones)

Transcriptional activation(Chaperones)

Ron D, Walter P. 2007. Signal integration in the endoplasmic reticulum unfolded protein response. Nat. Rev. Mol. Cell Biol. 8:519–29

UPR:(unfolded protein response)

The presence of unfolded proteins in the ER during stress triggers

a set of intracellular signalling pathways.

Alert to cell for UPR

Protein kinase RNA (PKR)-like ER kinase (PERK)

Activating transcription factor-6 (ATF6)

Inositol-requiring enzyme-1 (IRE1a)

IRE1IRE1

bZIPbZIP

PERKPERK

Cell fate outcomeCell fate outcome

KinaseKinase ER LUMEN

KinaseKinase

ATF6

T. Kudo, S. Kanemoto, H. Hara et al., “A molecular chaperone inducer protects neurons from ER stress,” Cell Death Differentiation, vol. 15, no. 2, pp. 364–375, 2008.

ER stress

Three signalling proteins become activated through direct or indirect

binding of unfolded proteins

Signals from IRE1a, PERK, and ATF6 initially trigger transcriptional

programs .

Activation of ER chaperones, oxidoreductases, and ERAD components.

By increasing the complement of ER protein-folding and quality control

enzymes, the UPR enhances the cell’s capacity.

Allow pre existing proteins to fold before new ones are made.

Successful UPR restores homeostasis managing cell ER stress.

Condition whereby ER stress persists at high levels, the UPR

promotes programmed cell death, usually through apoptosis.

Vigorous apoptopic process results into organ failure

Caused by an insufficient mass of functioning cells.

Degeneration of insulin-producing pancreatic islet β cells

causes diabetes.

Endoplasmic Reticulum Stress and Type 2 Diabetes Sung Hoon Back and Randal J. Kaufman Reviews in Advance,12 March,2014, 16.2-16.19

25% of cellular ROS originates from the ER through the process of oxidative protein folding.

Acts through protein disulfide isomerases (PDIs)

The ERO1 oxido-reductase transfers electrons from cysteine sulfhydryl groups on ER-translocated proteins to molecular oxygen, generating H2O2 as an ROS by product.

The b-cell ER needs to produce three disulfides for each molecule of pro-insulin it synthesizes(i.e., 3 million disulfides/min per cell).

A large load of ROS is necessarily produced that needs to be continually disposed of

D. Ron and P. Walter, “Signal integration in the endoplasmic reticulum unfolded protein response,” Nature Reviews Molecular Cell Biology, vol. 8, no. 7, pp. 519–529, 2007

Diabetes-hyperglycemia and hyperlipidemiaDiabetes-hyperglycemia and hyperlipidemia

Altered cardiac metabolism &

lipotoxicity

Altered cardiac metabolism &

lipotoxicity

inflammationinflammation

Plasma glucose level

Plasma glucose level FFAFFA

AutophagyAutophagy Insulin resistanceApoptosisApoptosis

ER STRESS

Cardiac myopathy

T. Minamino and M. Kitakaze, “ER stress in cardiovascular disease,” Journal of Molecular and Cellular Cardiology, vol. 48, no. 6, pp. 1105–1110, 2010

ER STRESSER STRESS

Apoptopic β cellApoptopic β cell

Inflammatory responses(IFN ,IL 1β)

Nitric oxide

ATF3 SERCA 2b

CHOP

D. Ron and P. Walter, “Signal integration in the endoplasmic reticulum unfolded protein response,” Nature Reviews Molecular Cell Biology, vol. 8, no. 7, pp. 519–529, 2007

ER ER STRESSSTRESS

ApoptopicApoptopic β cell

ApoptopicApoptopic β cell

FFA hyperinsulinimia

Insulin resistance

IRE1

p-junk CHOP p-akt

p-GSK-3βSerine

p IRS-1

Kahn SE, Hull RL, Utzschneider KM. 2006. Mechanisms linking obesity toinsulin resistance and type2 diabetes. Nature 444:840–46

β cells contain highly developed ER

Each β cell produces approximately 1million molecules of insulin every minute

Complex series of molecular biosynthetic processes

The insulin precursors prepro-insulin( translocated into the ER lumen )

Signal sequence is removed

Oxido-reductases

Insulin precursor Pre pro-insulin ( translocated into the ER lumen )

Formation of three intramolecular disulfides Pro-insulin folds( native state in the ER)

Trafficking to downstream Golgi and secretory granules endoproteases.

Removal of C-peptide

Generation of mature insulin

Pro-insulin

Performed with unnatural ER stress agents causing apoptosis. Measuring ER stress in β cells of living humans is currently not feasibleBecause the pancreatic islet β cell mass and insulin secretory function during life cannot be studied simultaneously. Pancreatic autopsy series from type 2 diabetic patients technique used Show a reduction in β -cell mass with activation in UPR apoptotic markers.

Human type 2 diabetic islets contain protein aggregates in the form of amyloid Islet .Amyloid is composed of a 37-residue amyloidogenic polypeptide called islet amyloid polypeptide (IAPP). IAPP spontaneously forms ER membrane-damaging sheets of amyloid IAPP could be another ER stress link promoting b-cell death through activation of the pro-apoptotic transcription factor CHOP

DRUG MECHANISM POTENTIAL INDICATION

Bip inducer X Induction of GRP 78 Heart failure, stroke

Cs-866 (VESINO INDUSTRIAL CO., LTD.)

Reduction of ER stress by pressure overload

Heart failure, cardiac hypertrophy

Curcumin Induction of GRP 94 Diabetic cardiomyopathy

Salubrinal (Cayman Chemical)

Prevention of elf2a dephosphorylation

Cardiac hypertrophy

Tauroursodeoxycholic acid(TUCD) (Bio-Gen, Turkey)

Chemical chaperone Heart failure, atherosclerosis

TNFα antibody Prevention of CHOP induction

Cardiac hypertrophy

Sunitinib IRE1 activation Heart failure, cardiac hypertrophy

Pioglitazone Decrease ER stress CardioprotectinCohen FE, Kelly JW. Therapeutic approaches to protein-misfoldingb diseases. Nature. 2013 3426:905–909

Role of hyperglycemia or hyperlipidemia in β-cell failure in T2D.

Molecular mechanism of glucose level-dependent differential activation of PERK and IREα in pancreatic β-cells

Lipid intermediates activate or inhibit the UPR

Mechanism of activation of UPR in β-cells by high glucose .

Is there any direct evidence of ER stress?

Molecular mechanisms involved in the formation of the toxic hIAPP oligomers and UPR induction by hIAPP oligomers

• Further investigation is needed to identify effective chemical chaperones that can improve ER function to prevent hyperactivation of the UPR.

ER maintains protein folding by UPR in manageable manner if it exceeds capacity causes misfolding of proteins.

Results into activation of apoptopic pathways and other dysfunction.

ROS generation, mitochondrial dysfunction results into β cell failure.

Islet amyloid polypeptide (IAPP is important mechanism in diabetes.

Targeting pre-apoptopic pathways is essential to achieve more specific actions.

L. Hiebert, J. Han, A.K. Mandal, Glycosaminoglycans, hyperglycemia and disease, Antioxid. Redox Signal. (2014 Feb 4)T.Miki, S. Yuda, H. Kouzu, T.Miura, Diabetic cardiomyopathy: pathophysiology and clinical features, Heart Fail. Rev. 18 (2013) 149–166.Ron D, Walter P. Signal integration in the endoplasmic reticulum unfolded protein response. Nat Rev Mol Cell Biol. 2007;8:519 –529. Jweied EE, McKinney RD, Walker LA, et al. Depressed cardiac myofilament function in human diabetes mellitus Am J Physiol Heart Circ Physiol 2005; 289: H2478–H2483