Nada Lukkahatai, PhD, RN

Co-authors: B. Majors, BS; S. Reddy, PhD; B. Walitt, MD; L. Saligan, PhD

Conflict of Interest

There is no conflict of interest to report.

Study Objective and Questions

Objective:

To examine differential expression of genes from fatigued fibromyalgia women with different levels of pain and catastrophizing.

Specific Research Questions:

Does the gene expression profile different in fatigued fibromyalgia women with high and low pain severities?

Does the gene expression profile different in fatigued fibromyalgia women with high and low catastrophizing?

Fibromyalgia

Fibromyalgia (FM) is characterized by prolonged widespread muscle pain, profound fatigue, and sleep disturbance.

An estimated 10 million Americans and 200-400 million adults worldwide suffer from FM.

Prevalence: 3 times higher among women than men.

5.5 million FM patients visit the ambulatory care per year.

The etiology of the fibromyalgia is unknown.

Diagnostic Criteria for FM

1990 American College of Rheumatology (ACR)

Criteria

1.History of chronic widespread pain

2.Pain in at least 11/18 tender point sites on digital palpation.

2010 ACR Criteria

1. Widespread Pain Index (WPI) > 7 and Symptom Severity (SS) Scale score > 5

or

2.WPI = 3-6 and SS > 9

3.Symptoms present at similar level for > 3 months

4.No other disorder explaining the pain.

Diagnostic Criteria for FM

Catastrophizing

Catastrophizing is an exaggerated negative attention to symptoms.

Pain catastrophizing significantly predicts pain severity, chronic illness-related disability & emotional distress.

High fatigue catastrophizing is associated with high fatigue severity in breast cancer patients and predicts post cancer treatment fatigue.

Cognitive behavioral intervention focus on addressing maladaptive thinking improve fatigue severity and depression in chronic fatigue syndrome.

Gaps in Knowledge

No study has explored the role of catastrophizing in influencing possible biologic correlates of pain and fatigue in FM.

Little is known about the mechanisms that can explain both pain and fatigue symptoms experienced by FM patients.

Few studies have looked at possible genomic correlates of FM.

Study

Actively recruiting MedStar Research Institute protocol.

Women diagnosed with FM based on the 1990 and 2010 diagnostic criteria are currently enrolled in this study.

Methods

Questionnaires:

Pain severity subscale of Brief Pain Inventory (BPI) 4-item subscale; the mean subscale score ranged from 0 to 10. The cutoff score that is clinically significant is 5.

General fatigue subscale - Multidimensional Fatigue Inventory (MFI) 4-item subscale; score range from 4-20. The score of ≥ 13 is significant fatigue.

Pain Catastrophizing Scale (PCS) = 13-item questionnaire; scores ranged from 0 - 52. Suggested clinical cut point is 16.

Gene Expression

Microarray technology using Affymetrix GeneChip® human genome U133 Plus 2.0 array was used on blood collected using Paxgene® tubes.

Differential gene expression was analyzed using Partek software.

Gene selection criteria: over 2-fold increase or decrease, p < 0.05.

Network analyses by Ingenuity® software.

Whole blood Collected using Paxgene tubes

Extracted RNA

Microarray

Biological sample collection and analysis:

U133 Plus 2.0

Sample Characteristics

9 Caucasian, female, diagnosed with FM, 26-64 years old

Min Max Mean (SD) Clinical Cut-off

General Fatigue 13 20 17.1 (2.7) >13

Pain severity 0.5 6.3 4.1 (1.9) >5

Catastrophizing 4.0 36.0 17.0 (9.8) >16

Categories of Pain and Catastrophizing

N = 9

Pain:

- Low pain (pain severity < 5) n = 6

- High pain (pain severity ≥ 5) n = 3

Catastrophizing:

- Low catastrophizing (PCS < 16) n = 4

- High catastrophizing (PCS ≥ 16) n = 5

High Pain Severity vs. Low Pain Severity (112 genes)

Gene Symbol

Gene Title Functionp-

valueFold-

Change

BATF2

basic leucine zipper

transcription factor, ATF-

like 2

Protein binding/ protein

dimerization activity

0.0002 2.1

CASP5

caspase 5, apoptosis-

related cysteine

peptidase

Cyteine-type endopeptidase

activity0.0003 2.7

CCR1chemokine (C-C motif) receptor 1

C-C chemokine binding,

chemokine (C-C motif) activity

0.0004 2.2

CEACAM1

carcinoembryonic antigen-related cell adhesion

molecule 1

Molecular function, protein

binding0.001 2.4

COMMD6COMM domain

containing 6

NF-kappa-B binding/ protein

binding0.001 4.1

Gene Symbol

Gene Title Functionp-

valueFold-

Change

RPL7ribosomal protein L7

protein dimerization

activity0.02 -2.9

SH2D1BSH2 domain containing

1B

Immune responses

0.04 -2.8

SIGLEC1

sialic acid binding Ig-

like lectin 1, sialoadhesin

Carbonydrate binding

0.04 -2.7

UQCRB

ubiquinol-cytochrome c

reductase binding protein

ubiquinol-cytochrome-c

reductase activity

0.05 -2.6

ZCCHC2

zinc finger, CCHC domain

containing 2

Nucleic acid binding

0.05 -2.6

Up- regulated genes Down- regulated genes

Network Analysis for high pain vs. low pain

High Catastrophizing vs. Low Catastrophizing (63 genes)

Gene Symbol

Gene Title function p-valueFold-

Change

USP46ubiquitin specific

peptidase 46

Ubiquitin-specific protease activity

0.001 -2.2

SEPT10 septin 10 GTP binding 0.002 -2.0

CLEC4DC-type lectin

domain family 4, member D

Carbonydrate binding

0.005 -2.3

ZDHHC2zinc finger, DHHC-type containing 2

Zinc ion binding

0.005 -2.2

FAS

Fas (TNF receptor

superfamily, member 6)

Identical protein binding/ signal

transducer activity

0.010 -2.0

Gene Symbol

Gene Title functionp-

valueFold-

Change

SPP1secreted

phosphoprotein 1Cytokine activity

0.01 2.1

TMTC1

transmembrane and

tetratricopeptide repeat containing

1

Integral to membrane

0.02 2.5

SLC6A8

solute carrier family 6

(neurotransmitter transporter, creatine), member 8

Creatine transporter

activity/ molecular function

0.03 2.4

NPRL3

nitrogen permease

regulator-like 3 (S. cerevisiae)

Molecular function

0.04 2.1

HEMGN hemogenProtein binding

0.04 2.2

Up- regulated genes Down- regulated genes

Network Analysis high vs. low catastrophizing

Differentially Expressed Genes of Interest

Basic leucine zipper protein (BATF2) is regulated by interferon and serves as a suppressor of activating protein-1 (AP-1).

Caspase 5, apoptosis-related cysteine peptidase (CASP5) and chemokine (C-C motif) receptor 1 (CCR1) are associated with immune response and inflammation in musculoskeletal disorders.

Secreted phosphoprotein 1 (SPP1 or Osteopontin) is up-regulated during inflammation and associated with muscular dystrophies.

Ubiquitin specific peptidase 46 (USP46) is a GABA regulation gene. In animal study, deletion of USP46 is associated with depression-like behaviors in mice and rats.

High vs Low Pain High vs Low Catastrophizing

Discussion

FibromyalgiaFibromyalgia

Physiological responses: Pain and fatigue

Physiological responses: Pain and fatigue

Behavioral responses: Catastrophizing,

depression

Behavioral responses: Catastrophizing,

depression

STRESSOR

USP46

Pituitary adrenocorticotropin (ACTH)

InflammationInflammation

Inflammatory Cells: Leukocytes, Lymphocytes, Platelets, Mast

Cells, Macrophages

Inflammatory Cells: Leukocytes, Lymphocytes, Platelets, Mast

Cells, Macrophages

Cytokines, Nerve Growth factor, prostaglandins,

Thromboxanes, Leukotrienes, Serotonin

Cytokines, Nerve Growth factor, prostaglandins,

Thromboxanes, Leukotrienes, Serotonin

BATF2CASP5CCR1

SPP1

Conclusion of Preliminary Study

Differentially expressed genes may delineate mechanisms between pain and fatigue.

Catastrophizing may serve as a behavioral correlate in FM.

Differentially expressed genes may serve as a biological correlate for catastrophizing.

Leorey N. Saligan, PhD, CRNPTenure-Track Investigator

National Institute of Nursing Research, Intramural Research Program

Brian Walitt, MD, MPH, FACRAssociate Professor of Medicine

Georgetown University Medical Center

Benjamin Majors, BSNational Institute of Nursing Research, Intramural Research Program

Swarnalatha Reddy, PhDNational Institute of Nursing Research, Intramural Research Program

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