Скачать презентацию Peripheral actions of the stress hormone Corticotropin Releasing Скачать презентацию Peripheral actions of the stress hormone Corticotropin Releasing

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Peripheral actions of the stress hormone Corticotropin Releasing Hormone (CRH): focus on its immunomodulatory Peripheral actions of the stress hormone Corticotropin Releasing Hormone (CRH): focus on its immunomodulatory effects

The Hypothalamic-Pituitary Adrenal (HPA) axis Hypothalamus CRH + VP + Anterior pituitary ACTH Adrenal The Hypothalamic-Pituitary Adrenal (HPA) axis Hypothalamus CRH + VP + Anterior pituitary ACTH Adrenal cortex Glucocorticoid

Sites of CRH synthesis BRAIN SPINAL CORD IMMUNE SYSTEM GI TRACT PLACENTA T-lymphocyte LUNG Sites of CRH synthesis BRAIN SPINAL CORD IMMUNE SYSTEM GI TRACT PLACENTA T-lymphocyte LUNG

CRH Receptors CRH RECEPTOR 1 (CRHR 1): Brain, anterior pituitary, immune system, GI tract, CRH Receptors CRH RECEPTOR 1 (CRHR 1): Brain, anterior pituitary, immune system, GI tract, adrenal gland, skin CRH RECEPTOR 2 (CRHR 2): Brain, pituitary, immune system, GI tract, adrenal gland, heart, skin, skeletal muscle

CRH AND INFLAMMATION CRH INFLAMMATION ACTH GC PERIPHERAL CRH CRH AND INFLAMMATION CRH INFLAMMATION ACTH GC PERIPHERAL CRH

Dual effects of CRH on the immune /inflammatory response * suppressive, by central CRH Dual effects of CRH on the immune /inflammatory response * suppressive, by central CRH *stimulatory, by peripheral CRH

Signaling pathways mediating the immunomodulatory effects of CRH ? ? Signaling pathways mediating the immunomodulatory effects of CRH ? ?

n. s. CRH 10 -7 M Control l NF- B p 50 Ctrl p n. s. CRH 10 -7 M Control l NF- B p 50 Ctrl p 65 CRH Ctrl CRH

NF- B n. s. 10 7 M l g /m M h ut CRH NF- B n. s. 10 7 M l g /m M h ut CRH an t c 10 -6 om M co pe m tit pe or tit or H/ CR H CR 10 rl S LP Ct

I B CRH 30 min CRH 15 min CONTROL I B CRH 30 min CRH 15 min CONTROL

p 38 (5 min) p 42/44 (5 min) (10 -9 M) CRH (10 -8 p 38 (5 min) p 42/44 (5 min) (10 -9 M) CRH (10 -8 M) CRH (10 -7 M) CRH (10 -6 M) EGF (10 n g/m l) CRH trol Con (10 -9 M) CRH (10 -8 M) CRH (10 -7 M) CRH (10 -6 M) TNF (1 0 ng /ml) CRH trol Concentration-dependent effects of CRH on MAPK activation in human leukocytes JNK (30 min)

Is there an important physiological role for peripheral CRH itself in the regulation of Is there an important physiological role for peripheral CRH itself in the regulation of the inflammatory process ?

The Crh-deficient animal model Normal lifespan Normal weight Normal food intake CRH Blunted/absent circadian The Crh-deficient animal model Normal lifespan Normal weight Normal food intake CRH Blunted/absent circadian rhythmicity Low basal corticosterone Impaired ACTH and corticosterone responses to multiple stressors ACTH GC Muglia et al. Nature, 1995

Sources of peripheral CRH Hypothalamus CRH Pituitary leukocytes ACTH Nerve fibers ? Inflammatory sites Sources of peripheral CRH Hypothalamus CRH Pituitary leukocytes ACTH Nerve fibers ? Inflammatory sites Adrenals glucocorticoid

GLUCOCORTICOID INFLAMMATION EPINEPHRINE CRH GLUCOCORTICOID INFLAMMATION EPINEPHRINE CRH

CYTOKINE RESPONSE TO TURPENTINE Crh+/+ turpentine 2400 Crh-/- turpentine IL-6 (pg/m. L) 1800 1200 CYTOKINE RESPONSE TO TURPENTINE Crh+/+ turpentine 2400 Crh-/- turpentine IL-6 (pg/m. L) 1800 1200 600 0 1 4 8 16 Time (hr) 24 30

HORMONAL RESPONSES TO TURPENTINE Corticosterone (ug/d. L) Crh +/+ saline Crh+/+ turpentine Crh -/- HORMONAL RESPONSES TO TURPENTINE Corticosterone (ug/d. L) Crh +/+ saline Crh+/+ turpentine Crh -/- saline Crh-/- turpentine ACTH (pg/m. L) 600 500 * 400 300 * 200 100 0 1 4 8 16 24 Time (hr) 30 48 60 45 30 15 0 1 4 8 16 24 Time (hr) 30 48

Crh +/+ saline Crh+/+ turpentine Crh -/- saline Crh-/- turpentine 0. 30 *# 0. Crh +/+ saline Crh+/+ turpentine Crh -/- saline Crh-/- turpentine 0. 30 *# 0. 25 0. 20 0. 15 * *# * 0. 10 0. 05 1 2 3 4 5 Days following the injection Body weight (% of control) Food intake (gr)/mouse weight (gr) METABOLIC RESPONSES TO TURPENTINE 108 * * 104 * # 100 # # 96 92 1 2 3 4 5 6 Day following the injection

SYSTEMIC INFLAMMATION: Is there an important physiological role for peripheral CRH itself on the SYSTEMIC INFLAMMATION: Is there an important physiological role for peripheral CRH itself on the regulation of the inflammatory process, or its effects are linked to its role in stimulating the release of glucocorticoid ?

900 750 * 600 *$ 450 300 150 0 Saline LPS Saline. LPS Crh+/+ 900 750 * 600 *$ 450 300 150 0 Saline LPS Saline. LPS Crh+/+ Crh-/- Corticosterone ( g/dl) ACTH (pg/ml) HPA axis response 1 hr after LPS or saline 80 * 60 * 40 20 0 Saline LPS Saline. LPS Crh+/+ *: P<0. 05 between treatments in the same genotype $: P<0. 05 between genotypes following the same treatment Crh-/-

35 0 30 * 0 20 0 10 0 0 *$ Saline LPS Crh+/+ 35 0 30 * 0 20 0 10 0 0 *$ Saline LPS Crh+/+ Saline LPS Crh-/- Corticosterone ( g/dl) ACTH (pg/ml) HPA axis response 24 hr after LPS or saline * 60 * 50 40 30 20 10 0 Saline LPS Crh+/+ *: P<0. 05 between treatments in the same genotype $: P<0. 05 between genotypes following the same treatment Saline LPS Crh-/-

375 Corticosterone ( g/dl) ACTH (pg/ml) Effect of pre-treatment with antalarmin on the HPA 375 Corticosterone ( g/dl) ACTH (pg/ml) Effect of pre-treatment with antalarmin on the HPA axis response 24 hr after LPS 40 300 32 225 n. s. 24 * 150 75 0 Vehicle + LPS Antalarmin + LPS 16 8 0 Vehicle + LPS Antalarmin + LPS

Plasma glucose levels 1 and 24 hr after LPS or saline 250 Glucose ( Plasma glucose levels 1 and 24 hr after LPS or saline 250 Glucose ( g/dl) 200 Crh+/+ 150 100 Saline LPS 50 200 Crh-/- 150 100 50 1 24 Time (hr) following the injection

P g/ ml Plasma epinephrine at basal levels, and 24 hours post-LPS or –saline P g/ ml Plasma epinephrine at basal levels, and 24 hours post-LPS or –saline injection P<0. 01 6000 4000 2000 0 basal saline Crh+/+ LPS basal saline Crh-/- LPS

Systemic Bacterial Inflammation ? ? ? CRH CYTOKINES ACTH ? ? GC LEPTIN ? Systemic Bacterial Inflammation ? ? ? CRH CYTOKINES ACTH ? ? GC LEPTIN ? Decreased food intake / Body weight loss-cachexia

Plasma TNF 11/2, 4 hours and 24 hours post-LPS injection 30000 P<0. 01 * Plasma TNF 11/2, 4 hours and 24 hours post-LPS injection 30000 P<0. 01 * 25000 Pg/ml 20000 15000 10000 * 5000 0 Crh+/+ Crh-/- 11/2 hour Crh+/+ Crh-/- 4 hours Crh+/+ Crh-/- 24 hours * : statistical significance of p<0. 01 from TNF levels 4 and 24 hours post-LPS injection between animals of the same genotype

Plasma IL-6 4 hours and 24 hours post-LPS injection 5000 Pg/ml 4000 3000 2000 Plasma IL-6 4 hours and 24 hours post-LPS injection 5000 Pg/ml 4000 3000 2000 1000 0 Crh+/+ Crh-/- 4 hour 24 hour

Systemic Bacterial Inflammation TLRs ? ? ? CRH ACTH CYTOKINES ? ? GC LEPTIN Systemic Bacterial Inflammation TLRs ? ? ? CRH ACTH CYTOKINES ? ? GC LEPTIN ? Decreased food intake / Body weight loss-cachexia

Increased leptin levels have been reported to exert protective effects during inflammation Increased leptin levels have been reported to exert protective effects during inflammation

CRH and inflammationassociated processes • Angiogenesis CRH receptors are found in endothelial cells CRH CRH and inflammationassociated processes • Angiogenesis CRH receptors are found in endothelial cells CRH causes chemotaxis of endothelial cells in vitro CRH induces NF-k. B DNA binding activity in human endothelial cells • Wound healing Crh-/- mice show altered wound healing

HPA axis, LEPTIN, and LPS administration results in * activation of the immune system HPA axis, LEPTIN, and LPS administration results in * activation of the immune system manifested by increased plasma levels of proinflammatory cytokines, including TNF , IL-1 and IL-6, *activation of the HPA axis, and * induction of the ob gene, most likely by the action of cytokines on adipocytes.

Leptin (ng/ml) Plasma leptin 24 hr after LPS or saline 35 30 25 20 Leptin (ng/ml) Plasma leptin 24 hr after LPS or saline 35 30 25 20 15 10 5 0 * *$ Saline LPS Crh+/+ Saline LPS Crh-/- *: P<0. 05 between treatments in the same genotype $: P<0. 05 between genotypes following the same treatment

Leptin (ng/ml) Effect of pre-treatment with antalarmin on leptin secretion 24 hrs after LPS Leptin (ng/ml) Effect of pre-treatment with antalarmin on leptin secretion 24 hrs after LPS 25 20 15 * 10 5 0 Vehicle + LPS * P=0. 05 Antalarmin + LPS

Responses of Crh-/- splenocytes to LPS Responses of Crh-/- splenocytes to LPS

TNF- (pg/ml/million of plated cells) A B 250 * $ 200 150 TNF- * TNF- (pg/ml/million of plated cells) A B 250 * $ 200 150 TNF- * 100 50 0 actin control LPS Crh+/+ control LPS Crh-/- Crh+/+ Crh-/-

IL-1 (pg/ml/million of plated cells) A 10. 0 B * $ 7. 5 IL-1 IL-1 (pg/ml/million of plated cells) A 10. 0 B * $ 7. 5 IL-1 5. 0 -actin 2. 5 0. 0 control LPS Crh+/+ control LPS Crh-/- Crh+/+ Crh-/-

WT WT SPL +LPS HT CRHKO Crh WT WT SPL +LPS HT CRHKO Crh

Regulation of systemic inflammation by CRH and glucocortioid What are the target organs / Regulation of systemic inflammation by CRH and glucocortioid What are the target organs / cells? Is there a role for CRH and/or glucocorticoid on the regulation of innate immunity? What is the contribution of the above hormones in the metabolic changes triggered by inflammation?

Immunomodulatory neuropeptides (*acting through GPCRs) Appetite control Body weight regulation Cachexia of chronic diseases Immunomodulatory neuropeptides (*acting through GPCRs) Appetite control Body weight regulation Cachexia of chronic diseases Obesity Diabetes Atherosclerosis CNS disease Cytokines and Related peptides INFLAMMATION INFECTION

Jie Zhao Lilian van. Vlerken Christina Chandras Maria Venihaki Jerome Gay Joseph Majzoub Children’s Jie Zhao Lilian van. Vlerken Christina Chandras Maria Venihaki Jerome Gay Joseph Majzoub Children’s Hospital Endocrine Division, Boston Harris Pothoulakis Efi Kokkotou Beth Israel Hospital, Boston Yassemi Koutmani Despina Xanthaki Christina Chandras Maria Venihaki Developmental Biology Section, Foundation for Biomedical Research, Academy of Athens (IIBEAA)

Reduced toxin A-associated histologic damage and inflammation in CRH deficient mice Epithelial damage Congestion Reduced toxin A-associated histologic damage and inflammation in CRH deficient mice Epithelial damage Congestion & edema PMN Histologic scores 2. 0 Crh +/+ 1. 5 1. 0 Crh - / + ++ 0. 5 0. 0 +, ++ significantly different (p<0. 05, 0. 01) from WT toxin A treated mice

plasma corticosterone ( g/dl) Toxin A modulation of plasma corticosterone in CRH deficient mice plasma corticosterone ( g/dl) Toxin A modulation of plasma corticosterone in CRH deficient mice buffer toxin A 50 40 30 20 10 * * 0 Crh +/+ Time 0 Crh -/- Crh +/+ Crh -/- 4 hours * significantly different (p<0. 05, p<0. 001) from their respective Crh +/+ counterparts (4 hours)

Intestinal fluid secretion (mg/cm) CRH receptor antagonists reduce toxin A-induced ileal fluid secretion 140 Intestinal fluid secretion (mg/cm) CRH receptor antagonists reduce toxin A-induced ileal fluid secretion 140 120 * 100 80 60 40 20 0 ++ ++ Control Tx. A + antalarmin Tx. A + -helical CRH * significantly different (p<0. 05) from controls ; ++ significantly different (p<0. 01) from Tx A

LEPTIN and HPA axis Leptin, the product of the ob gene, interacts reciprocally with LEPTIN and HPA axis Leptin, the product of the ob gene, interacts reciprocally with the hypothalamic-pituitary adrenal (HPA) axis: Glucocorticoid, stimulates the expression of the ob gene in adipocytes. Administration of leptin increases CRH and ACTH secretion, while it decreases glucocorticoid release in rodents.

TLR 4 expression in Crh+/+ and Crh-/- tissues following LPS or saline administration Crh+/+ TLR 4 expression in Crh+/+ and Crh-/- tissues following LPS or saline administration Crh+/+ saline 3. 5 p<0. 05 Crh-/- saline 3 m. RNA expression (densiometric units) Crh+/+ LPS Crh-/- LPS 2. 5 p<0. 05 2 p<0. 01 1. 5 p<0. 05 1 0. 5 0 lung thymus peritoneal leukocytes pituitary

Weight variation (% of d-1) A 110 Crh+/+ controls Crh-/- controls 105 100 95 Weight variation (% of d-1) A 110 Crh+/+ controls Crh-/- controls 105 100 95 90 Crh-/- TNBS ** *** +/+ ** 85 Crh 80 d-1 B d 0 24 h Food intake (g) 5 Crh-/- controls Crh+/+ controls 4 3 2 Crh+/+ TNBS 1 Crh-/- TNBS 0 24 h 48 h TNBS time

Phospho-p 38 Total p 38 l ) /µl ng α( 10 TN F ng Phospho-p 38 Total p 38 l ) /µl ng α( 10 TN F ng 10 Fα ( TN H( 10 -6 M) CR ol tro ntr Co Co n

CRH CRH CRH Inflammatory stimulus intestine ? CRH IBD CRH 2 inflammation Inflammatory stimulus CRH CRH CRH Inflammatory stimulus intestine ? CRH IBD CRH 2 inflammation Inflammatory stimulus CRH deficient