Return to Affinity EATING DISORDER RESEARCH Sponsored by the Kartini Foundation PROJECT ONE: Gene expression analysis before and after novel approach which remits anorexia nervosa (AN) symptom complex with a brief sequence of treatment. Determination of epigenetic modification following novel, symptom remitting treatment of anorexia nervosa. RESEARCH PROCESS: Research subjects under the age of 22 years with a DSMIV diagnosis of AN will be recruited from the active treatment population at the Kartini Clinic for Disordered Eating, Portland Oregon (www.kartiniclinic.com). Healthy anorexia free control subjects will be recruited from a local high school and community college. All AN subjects receive the standard clinic treatment. All subjects will contribute an initial saliva sample. AN experimenters receiving a novel, symptom remitting treatment and AN controls not receiving the novel treatment (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf Walsh, 2010) will contribute saliva samples one hour and twenty four hours after AN experimenters receive the novel treatment. AN experimenters and AN controls will provide saliva samples at approximately the same chronological sequence of standard clinic treatment. mRNA is extracted from saliva samples to observe any changes in gene expression before and after the novel treatment, as well as to compare these affected genes to those of the AN-free control group. The R.E.D.S.C test (DeSosio et al, 2011; Goldner, 2000; O’Toole, 2000) will be administered to all AN subjects prior to administration of the novel treatment and again following the novel treatment to those receiving this treatment to determine severity of AN symptoms. IMPLICATIONS: If gene expression is altered as a result of this novel treatment, then the treatment field is privy to a specific application of behavioral epigenetics treatment and also able to identify genes likely involved in AN expression. The results of the novel treatment have been consistently observed over a number of years, so it is time to scientifically validate what seems to likely be happening at a treatment level. This will provide evidence to support the propagation of a more effective treatment which empowers the affected individual to fully manage the disorder. More details DESCRIPTION: This project aims to find unique gene signature of anorexia nervosa (AN) and assess gene expression changes (epigenetic modification) occurring after AN symptoms remit following the application of a unique treatment intervention (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf ; Walsh, 2010). Documentation of eating disorder treatment using non-conscious or unconscious communication, ostensibly under the guise of hypnosis, is largely anecdotal without supporting empirical research. The novel treatment applied here requires no hypnotic induction. Verifying epigenetic modification following the remission of AN symptoms not only invites a teachable form of innovative behavioral treatment into the eating disorder treatment arena but also identifies specific genes contributing to AN. Saliva samples will be collected from patients undergoing the same intensive treatment at a multidisciplinary eating disorder treatment clinic serving eating disordered youth (Kartini Clinic www.kartiniclinic.com). In addition to the prescribed course of clinic treatment, a subgroup will receive the unique intervention involving communication with the unconscious through established finger movements for “yes” and “no” known as ideomotor signaling (Cheek & LeCron, 1968; Rossi & Cheek, 1988; Cheek, 1994 ). This intervention, applied to at least 50 AN patients, consistently results in remission of symptoms during a brief treatment sequence (1-4 sessions). Saliva samples will be taken from the intervention group before the unique intervention and one hour and 24 hours following the intervention. AN patients not receiving the unique intervention will provide saliva samples on their clinic visit day and the following day. Saliva samples from healthy, non-anorexic youth will be collected to serve as a healthy control group. Saliva will be collected using the Qiagen RNeasy Protect Saliva Mini Kit which has been shown to effectively preserve RNA for gene expression analysis. Then, mRNA will be separated from saliva samples and hybridized with Affymetrix Human Gene 1.0 ST Arrays for gene expression profiling. Any epigenetic modifications in both anorexia groups will be assessed by comparing gene expression profiles. Modified genes will be identified and searched for in the healthy control group to determine their status (silent/expressed), if they are present. The R.E.D.S.C test (DeSosio et al, 2011; Goldner, 2000; O’Toole, 2000) will be administered to all AN subjects prior to administration of the novel treatment and again following the novel treatment to those receiving this treatment to determine severity of AN symptoms. IMPLICATIONS: This information has the potential of 1) identifying at least some of the genes likely involved in AN symptom expression, 2) confirming gene expression as a factor in AN symptoms and 3) verifying a new, teachable approach to behavioral epigenetics treatment. Comments about significance of this research from NIH reviewers: •Anorexia is a lethal psychiatric disorder with a substantial mortality rate and with few treatment options – those that exist are complex interventions requiring the entire family to participate. Demonstration of some symptom improvement and identification of genes responding to an intervention and possibly associated with symptom improvement would be welcome. Project proposes multiple innovative strategies and methods to collect preliminary data: a longitudinal usual care and intervention treatment response paradigm; collection of saliva for gene expression analysis; and use of a recently developed microarray with multiple probe sequence that interrogates genes along the transcript. If the aims are achieved, this may stimulate the collection and analysis of saliva as a biospecimen for analysis of treatment response. •The study design includes a novel clinical intervention for the treatment of eating disorders (unconscious communication ), saliva biospecimen collection and carefully outlined analyses of salivary gene expression. •Analysis of salivary gene expression is novel with a few studies published in cross-sectional and in longitudinal designs. •Using high-throughput expression analysis to identify differentially expressed genes between different treatment groups is a better choice over measuring a few candidate genes. OPERATIONAL PREMISE OF NOVEL TREATMENT: The premise guiding this work assumes that ego-states (Watkins & Watlins, 1997; Watkins, 1993) or intra-psychic parts are functioning at the expressive level of anorexia nervosa (AN) and that the functional force directing these ego-states can be muted once all AN ego-states assume a different function. Extracting ego-states from the AN role and muting the AN force via ideomotor questioning (Cheek, 1994; Rossi & Cheek, 1988) of the unconscious then results in remission of the AN symptom complex. The treatment intervention targets the entire symptom complex, as opposed to focusing on single symptoms or presumed underlying causal factors. HYPOTHESIS: IF genes are involved in AN expression as science indicates (Bergen et al., 2003; Becanu et al., 2005; Bulik, 2006; Bulik, 2007; Bulik, Sullivan & Kendler, 1998; Grice et al., 2002; Mercader et al., 2007; Ribases et al., 2003; Ribases et al., 2004; Ribases et al, 2005; Devlin, et al., 2002; Gunstad et al., 2006; Frieling et al., 2007; Frieling et al., 2008; Steiger et al., 2006; Javaras, Laird, Reichborn-Kjennerud, Bulik & Pope, 2008; Klump, Miller, Keel, McGue and Iacono, 2001; Kendler et al., 1991) and IF experience-dependent gene expression can be influenced by behavioral or environmental conditions in animal and human models as science indicates (Weaver, Cervoni,, Champagne, Alessio, Sharma, Seckl, Dymov, Szyf & Meaney, 2004; Darnaudery & Maccari, 2008; Schanberg,Ingledue,Lee,Hannun, and Bartolome, 2003) and IF evidence exists supporting experience-dependent genes being involved in some aspects of eating disorder (ED) symptoms (Ribases et al., 2005; Kaplan et al., 2008) and IF AN-like symptoms can be expressed and remitted in animal studies through epigenetic modification (activating or silencing genes) (Owen, Treasure & Collier, 2001; Kas, Van Elburg, Van Engeland & Adan, 2003; Siegfried, Berry, Hao & Avraham, 2003; Schanberg, Ingledue, Lee, Hannun, & Bartolome, 2003; Weaver, Cervoni, Champagne, et al., 2004) and IF consistent rapid remission of the AN symptom complex occurs with this novel intervention (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf ), THEN there is reason to consider gene expression being influenced with this intervention. Comparing gene expression profiles of AN subjects pre and post novel treatment to AN subjects undergoing conventional treatment and AN-free healthy controls will refute or support this premise. This gene expression profile analysis could also validate the efficacy of perhaps the first documented behavioral epigenetics treatment and identify genes contributing to AN expression, opening the door for further research. REFERENCES: Becanu, S-A, Bulik, C.M., Klump, K.L., Fichter, M.M., Halmi, K.A., Keel, P.K., et al. (2005). Linkage analysis of anorexia and bulimia nervosa cohorts using selected behavioral phenotypes as quantitative traits or covariates. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics, 139, 61-68. Barbarasz, M. (2000). Hypnosis in the treatment of eating disorders. In L.M. Hornyak & J.P. Green, Healing from within: The use of hypnosis in women’s health. Washington, D.C.: American Psychological Association. Bergen, A., Van Den Bree, M., Yeager, M., Welch, R., Ganjei, K., Haque, K., Bacanu, S., Berrettini, W., Grice, D., Goldman, D., Bulik, C., Klump, K., Fichter, M., Halmi, K., Kaplan, A., Strober, M., Treasure, J., Woodside, B., Kaye, W.. (2003). Candidate genes for anorexia nervosa in the 1p33-36 linkage region: serotonin 1D and delta opioid receptors display significant association to anorexia nervosa. Molecular Psychiatry, 8 (4): 397-406. Bulik, C.M., Sullivan, P.F., Tozzi, F., Furberg, H., Lichtenstein, P., Pederden, N.L. (2006), Prevalence, heritability, and prospective risk factors for anorexia nervosa. Archives of General. Psychiatry, 63, 305-312. Bulik, C.M., Hebebrand, J., Keski-Rahkonen, A., Klump, K.L., Reichborn-Kjennerud, T., Mazzeo, S.E., Wade, T.D. (2007). Genetic epidemiology, endophenotypes and eating disorder classification. International Journal of Eating Disorders. 40(sup): s52-60. Cheek, David B. (1994). Hypnosis: The Application of Ideomotor Techniques. Needham Heights, Massachusetts: Allyn & Bacon. Bulik, C.M., Sullivan, P.F., Kendler, K.S. (1998). Heritability of binge-eating and broadly-defined bulimia nervosa. Biological Psychiatry, 44, 1210-1218. Cheek, D. & LeCron, L. (1968). Clinical Hypnotherapy. New York: Grune and Stratton. Cheek, D. (1994). Hypnosis: The Application of Ideomotor Techniques. Boston: Allyn and Bacon. Darnaudéry, Muriel and Maccari, Stefania (2008). Epigenetic programming of the stress response in male and female rats by prenatal restraint stress. Brain Research Reviews Vol 57(2), 571-585. DeSocio, J.E., O’Toole, J.K., He, H., Koeller, P., Baird, S.A., Lukach, M. (2008). The REDS-C Rating of Eating Disordered Severity Interview for Children and Adolescents: Clinical Usefulness and Comparison with the Modified EDI-2. Manuscript submitted for publication. Devlin, B., Bacanu, S-A., Klump, K.L., Berrettini, W., Bergen, A., Goldman, D., et al. (2002). Linkage analysis of anorexia nervosa incorporating behavioral covariates. Human Molecular Genetics, 11, 689-696. Frieling, H., Bleich, S., Otten, J., Romer, K.D., Kormhuver, J., de Zwaan, M., Jacoby, G.E., Wihelm, J., Hillemacher, T., (2008). Epigenetic downregulation of atrial natriuretic peptide but not vasopressin mRNA expression in females with eating disorders is related to impusivity. Neuropharmacology, 2, January: advance online: doi:10.1038/sj.npp.1301662. Frieling, H., Gozner, A., Romer, K.D., Lenz, B., Bonsch, D., Wihelm, J., Hillemacher, T., de Zwaan, M., Kornhuber, J., Bleich, S. (2007). Global DNA hypomethylation and DNA hypermethylation of the alpha synuclein promoter in females with anorexia nervosa. Molecular Psychiatry, 12:229-230. Goldner, E.M. (2000). The REDS rating of eating disorder severity. Unpublished instrument and administration guidelines. Vancouver, British Columbia, Canada: Simon Fraser University. Grice, D.E., Halmi, K.A., Fichter, M.M., Strober, M., Woodside, D.B., Treasure, J.T., Kaplan, A.S., Magistretti, P.J., Goldman, D., Bulik, C.M., Kaye, W.H., Berrettini, W.H. (2002), Evidence for a susceptibility gene for anorexia nervosa on chromosome 1. American Journal of Human Genetics, 70 (3), 787-792. Gunstad,J., Schofield, P., Paul, R.H., Spitznagel, M.B., Cohen, R.A., Williams, L.M., Kohn, M., Gordon, E. (2006). BDNF Val66Met polymorphism is associated with body mass index in healthy adults. Neuropsychobiology, 53 (3), 153-156. Javaras, K.N., Laird, N.M., Reichborn-Kjennerud, T., Bulik, C.M., Pope, H.G.J., Hudson, J.I. (2008). Familiarity and heritability of binge eating disorder: Results of a case-control family study and a twin study. International Journal of Eating Disorders, 41, 174-179. Kaplan, A. (2004). Exploring the gene-environment nexus in anorexia, bulimia. Psychiatric Times, 21 (9). Kaplan, A.S., Levitan, R.D., Yilmaz, Z., Davis, C., Tharmalingam, S., James L. Kennedy, J.S. (2008). A DRD4/BDNF gene-gene interaction associated with maximum BMI in women with bulimia nervosa. International Journal of Eating Disorders, 41(1), 22-28. Kartini Clinic for Disordered Eating: Portland, Oregon. www.kartiniclinic.com Kas, M.J., VanElburg, A.A., VanEngeland, H. & Adan, R.H. (2003). Refinement of behavioral traits in animals for the genetic dissection of eating disorders. European Journal of Pharmacology, 480, 13-20. Kendler, K.S., MacLean, C. Heale, M.C., Kessler, R.C., Heath, A.C., Eaves, L.J. (1991). The genetic epidemiology of bulimia nervosa. American Journal of Psychiatry, 148, 1627-1635. Klump, K.L., Bulik, C.M., Kaye, W.H., Treasure, J., Tyson, E. (2009). Academy for eating disorders position paper: Eating disorders are serious mental illnesses. International Journal of Eating Disorders, 42(2), 97-103. Klump, K.L., Culbert, K.M. (2007). Molecular genetic studies of eating disorders: Current status and future directions. Current Directions Psychological Science, 16, 37-41. Klump, K.L., Miller, K.B., Keel, P.K., McGue, M., Iacono, W.G. (2001). Genetic and environmental influences on anorexia nervosa syndromes in a population-based twin sample. Psychological Medicine, 31, 737-740. Lynn, S.J., Rhue, J.W., Kvarl, S., Mare, C. (1993). The treatment of anorexia nervosa: A suggestive framework. Contemporary Hypnosis, 10(2), 73-80. Majdan, M., Shatz, C.J. (2006). Effects of visual experience on activity-dependent gene regulation in cortex. Nature Neuroscience 9, 650:650-659. Martinowich, K., Hattori, D., Wu, H., Fouse, S., He, F., Hu,Y., Fan, G., Sun, Y.E., (2003). DNA methylation-related chromatin remodeling in activity-dependent bdnf gene regulation. Science, 302 (5646), 890 – 893. Mercader, J. M., Ribasés, M., Gratacós, M., González, J.R., Bayés, M., de Cid, R., Badía, A., Fernández-Aranda,F., Estivill, X. (2007). Altered BDNF blood levels and gene variability are associated to anorexia and bulimia. Genes, Brain and Behavior ,6(8): 706-716. Oberlander T.F. , Weinberg J., Papsdorf M., Grunau R, Misri S., Devlin A.M. (2008). Prenatal exposure to maternal depression, neonatal methylation of human glucocorticoid receptor gene (NR3C1) and infant cortisol stress responses. Epigenetics, 3 (2); 1-9. O’Toole, J.K. (2000). The REDS-C rating of eating disorder severity interview for children. Unpublished instrument and administration guidelines. Portland, Oregon: The Kartini Clinic. Owen,J.B., Treasure, J., Collier, D.A. (2001). Animal Models- Disorders of Eating Behavior and Body Composition. New York: Springer. Polan HJ, Ward MJ. (1994). Role of the mother's touch in failure to thrive: a preliminary investigation. Journal American Academy Child Adolesc Psychiatry, 33(8):1098-105. Reichborn-Kjennerud, T., Bulik, C.M., Tambs, K., Harris, J.R. (2004). Genetic and environmental influences on Binge eating in the absence of compensatory behaviors: A population-based twin study. International Journal of Eating Disorders, 36, 307-314. Ribasés, M., Gratacòs, M., Fernández-Aranda, F., Bellodi, L., Boni, C., Anderluh, M., Cavallini, M.C., Cellini, E., Di Bella, D., Erzegovesi, S., Foulon, C., Gabrovsek, M., Gorwood, P., Hebebrand, J., Hinney, A., Holliday, J., Hu, X., Karwautz, A., Kipman, A., Komel, R., Nacmias, B., Remschmidt, H., Ricca, V., Sorbi, S., Wagner, G., Treasure, J., Collier, D.A. and Estivill, X. (2004), Association of BDNF with anorexia, bulimia and age of onset of weight loss in six European populations. Human Molecular Genetics, 13 (12), 1205-1212. Ribasés, M., Gratacòs, M., Fernández-Aranda, F., Bellodi, L., Boni, C., Anderluh, M., Cavallini, M.C., Cellini, E., Di Bella, D., Erzegovesi, S., Foulon, C., Gabrovsek, M., Gorwood, P., Hebebrand, J., Hinney, A., Holliday, J., Hu, X., Karwautz, A., Kipman, A., Komel, R., Nacmias, B., Remschmidt, H., Ricca, V., Sorbi, S., Tomori, M., Wagner, G., Treasure, J., Collier, D.A., and Estivill, X. (2005). Association of BDNF with restricting anorexia nervosa and minimum body mass index: a family-based association study of eight European populations. European Journal of Human Genetics, 13, 428–434. Ribases, M.; Gratacos, M.; Armengol, L.; de Cid, R.; Badia, A.; Jimenez, L.; Solano, R.; Vallejo, J.; Fernandez, F.; Estivill, X. (2003), Met66 in the brain-derived neurotrophic factor (BDNF) precursor is associated with anorexia nervosa restrictive type. Molecular Psychiatry, 8, 745-751. Rossi, E.L. & Cheek, D.B. (1988). Mind-body therapy: Ideodynamic healing in hypnosis. New York: W.W. Norton & Co. Rossi, E.L. (2002). The psychobiology of gene expression. New York, N.Y.: W.W. Norton & Company, Inc. Rotenberg, K.J., Taylor, D. & Davis, R. (2004). Selective mood-induced body image disparagement and enhancement effects: Are they due to cognitive priming or subjective mood? International Journal of Eating Disorders, 35 (3), 317 - 332. Schanberg, S. M., Ingledue, V. F., Lee, J. Y., Hannun, Y. A., and Bartolome, J. V. (2003). PKC Mediates Maternal Touch Regulation of Growth-Related Gene Expression in Infant Rats. Neuropsychopharmacology. 28, 1026-1030. Segerstrile, U., Molner, P. (1997) Eds. Nonverbal Communication: Where Nature Meets Culture. Mahwah, New Jersey: Lawrence Erlbaum Associates, Publishers. Siegfried, Z., Berry, E.M., Hao, S. & Avraham, Y. (2003). Animal models in the investigation of anorexia. Physiology and Behavior, 29, 39-45. Strahl, DB., Allis, CD. (2000). The language of covalent histone modifications. Nature 403:41-45 Steiger, H., Guavin, L., Joober, R., Israel, M., YingKin, M.K.Ng., Bruce, K.R., Richardson, J., Young, S.N., Hakim, J. (2006). Intrafamilial correspondences on platelet[H-] paroxetine-binding indices in bulimic probands and their unaffected first-degree relatives. Neurophychopharmacology, 31:1785-1792. Wade, T.D., Bulik, C.M., Neale, M., Kendler, K.S. (2000). Anorexia nervosa and major depression: Shared genetic and environmental risk factors. American Journal of Walsh, B.J. (2005). Utilization Sobriety: A Substance Abuse Solution. Portland, Oregon: Peaceful Pilgrim Press: 125-144. Walsh, B.J. (2008). Hypnotic alteration of body image in the eating disordered. American Journal of Clinical Hypnosis 50(4): 301-310. Walsh, B.J. (2010). Rapid remission of anorexia nervosa and unconscious communication. American journal of Clinical Hypnosis, 52(4): 315-328. Watkins, H.H. (1993). Ego-state therapy: An overview. American Journal of Clinical Hypnosis, 35(4): 232-240. Watkins, J. G. & Watkins, H.H. (1997). Ego states: Theory and therapy. New York: W.W. W.W. Norton & Co.. Weaver, I.C.G., Cervoni, N.,, Champagne, F.A.,1, 2, Alessio, A.C.D., Sharma, S., Seckl, J.R., Dymov, S., Szyf, M., & Meaney, M.J. (2004). Epigenetic programming by maternal behavior. Nature Neuroscience, 7, 847

EATING DISORDER RESEARCH

Sponsored by the Kartini Foundation

PROJECT ONE: Gene expression analysis before and after novel approach which remits anorexia nervosa (AN) symptom complex with a brief sequence of treatment.

Determination of epigenetic modification following novel, symptom remitting treatment of anorexia nervosa.

RESEARCH PROCESS: Research subjects under the age of 22 years with a DSMIV diagnosis of AN will be recruited from the active treatment population at the Kartini Clinic for Disordered Eating, Portland Oregon (www.kartiniclinic.com). Healthy anorexia free control subjects will be recruited from a local high school and community college. All AN subjects receive the standard clinic treatment. All subjects will contribute an initial saliva sample. AN experimenters receiving a novel, symptom remitting treatment and AN controls not receiving the novel treatment (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf Walsh, 2010) will contribute saliva samples one hour and twenty four hours after AN experimenters receive the novel treatment. AN experimenters and AN controls will provide saliva samples at approximately the same chronological sequence of standard clinic treatment. mRNA is extracted from saliva samples to observe any changes in gene expression before and after the novel treatment, as well as to compare these affected genes to those of the AN-free control group. The R.E.D.S.C test (DeSosio et al, 2011; Goldner, 2000; O’Toole, 2000) will be administered to all AN subjects prior to administration of the novel treatment and again following the novel treatment to those receiving this treatment to determine severity of AN symptoms.

IMPLICATIONS: If gene expression is altered as a result of this novel treatment, then the treatment field is privy to a specific application of behavioral epigenetics treatment and also able to identify genes likely involved in AN expression. The results of the novel treatment have been consistently observed over a number of years, so it is time to scientifically validate what seems to likely be happening at a treatment level. This will provide evidence to support the propagation of a more effective treatment which empowers the affected individual to fully manage the disorder.

More details

DESCRIPTION: This project aims to find unique gene signature of anorexia nervosa (AN) and assess gene expression changes (epigenetic modification) occurring after AN symptoms remit following the application of a unique treatment intervention (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf ; Walsh, 2010). Documentation of eating disorder treatment using non-conscious or unconscious communication, ostensibly under the guise of hypnosis, is largely anecdotal without supporting empirical research. The novel treatment applied here requires no hypnotic induction. Verifying epigenetic modification following the remission of AN symptoms not only invites a teachable form of innovative behavioral treatment into the eating disorder treatment arena but also identifies specific genes contributing to AN. Saliva samples will be collected from patients undergoing the same intensive treatment at a multidisciplinary eating disorder treatment clinic serving eating disordered youth (Kartini Clinic www.kartiniclinic.com). In addition to the prescribed course of clinic treatment, a subgroup will receive the unique intervention involving communication with the unconscious through established finger movements for “yes” and “no” known as ideomotor signaling (Cheek & LeCron, 1968; Rossi & Cheek, 1988; Cheek, 1994 ). This intervention, applied to at least 50 AN patients, consistently results in remission of symptoms during a brief treatment sequence (1-4 sessions). Saliva samples will be taken from the intervention group before the unique intervention and one hour and 24 hours following the intervention. AN patients not receiving the unique intervention will provide saliva samples on their clinic visit day and the following day. Saliva samples from healthy, non-anorexic youth will be collected to serve as a healthy control group. Saliva will be collected using the Qiagen RNeasy Protect Saliva Mini Kit which has been shown to effectively preserve RNA for gene expression analysis. Then, mRNA will be separated from saliva samples and hybridized with Affymetrix Human Gene 1.0 ST Arrays for gene expression profiling. Any epigenetic modifications in both anorexia groups will be assessed by comparing gene expression profiles. Modified genes will be identified and searched for in the healthy control group to determine their status (silent/expressed), if they are present. The R.E.D.S.C test (DeSosio et al, 2011; Goldner, 2000; O’Toole, 2000) will be administered to all AN subjects prior to administration of the novel treatment and again following the novel treatment to those receiving this treatment to determine severity of AN symptoms.

IMPLICATIONS: This information has the potential of 1) identifying at least some of the genes likely involved in AN symptom expression, 2) confirming gene expression as a factor in AN symptoms and 3) verifying a new, teachable approach to behavioral epigenetics treatment.

Comments about significance of this research from NIH reviewers:

•Anorexia is a lethal psychiatric disorder with a substantial mortality rate and with few treatment options – those that exist are complex interventions requiring the entire family to participate.

•Demonstration of some symptom improvement and identification of genes responding to an intervention and possibly associated with symptom improvement would be welcome.
•Project proposes multiple innovative strategies and methods to collect preliminary data: a longitudinal usual care and intervention treatment response paradigm; collection of saliva for gene expression analysis; and use of a recently developed microarray with multiple probe sequence that interrogates genes along the transcript.
•If the aims are achieved, this may stimulate the collection and analysis of saliva as a biospecimen for analysis of treatment response.

•The study design includes a novel clinical intervention for the treatment of eating disorders (unconscious communication ), saliva biospecimen collection and carefully outlined analyses of salivary gene expression.

•Analysis of salivary gene expression is novel with a few studies published in cross-sectional and in longitudinal designs.

•Using high-throughput expression analysis to identify differentially expressed genes between different treatment groups is a better choice over measuring a few candidate genes.

OPERATIONAL PREMISE OF NOVEL TREATMENT: The premise guiding this work assumes that ego-states (Watkins & Watlins, 1997; Watkins, 1993) or intra-psychic parts are functioning at the expressive level of anorexia nervosa (AN) and that the functional force directing these ego-states can be muted once all AN ego-states assume a different function. Extracting ego-states from the AN role and muting the AN force via ideomotor questioning (Cheek, 1994; Rossi & Cheek, 1988) of the unconscious then results in remission of the AN symptom complex. The treatment intervention targets the entire symptom complex, as opposed to focusing on single symptoms or presumed underlying causal factors.

HYPOTHESIS: IF genes are involved in AN expression as science indicates (Bergen et al., 2003; Becanu et al., 2005; Bulik, 2006; Bulik, 2007; Bulik, Sullivan & Kendler, 1998; Grice et al., 2002; Mercader et al., 2007; Ribases et al., 2003; Ribases et al., 2004; Ribases et al, 2005; Devlin, et al., 2002; Gunstad et al., 2006; Frieling et al., 2007; Frieling et al., 2008; Steiger et al., 2006; Javaras, Laird, Reichborn-Kjennerud, Bulik & Pope, 2008; Klump, Miller, Keel, McGue and Iacono, 2001; Kendler et al., 1991) and IF experience-dependent gene expression can be influenced by behavioral or environmental conditions in animal and human models as science indicates (Weaver, Cervoni,, Champagne, Alessio, Sharma, Seckl, Dymov, Szyf & Meaney, 2004; Darnaudery & Maccari, 2008; Schanberg,Ingledue,Lee,Hannun, and Bartolome, 2003) and IF evidence exists supporting experience-dependent genes being involved in some aspects of eating disorder (ED) symptoms (Ribases et al., 2005; Kaplan et al., 2008) and IF AN-like symptoms can be expressed and remitted in animal studies through epigenetic modification (activating or silencing genes) (Owen, Treasure & Collier, 2001; Kas, Van Elburg, Van Engeland & Adan, 2003; Siegfried, Berry, Hao & Avraham, 2003; Schanberg, Ingledue, Lee, Hannun, & Bartolome, 2003; Weaver, Cervoni, Champagne, et al., 2004) and IF consistent rapid remission of the AN symptom complex occurs with this novel intervention (http://www.asch.net/portals/0/journallibrary/articles/ajch-52/52-4/walsh.pdf ), THEN there is reason to consider gene expression being influenced with this intervention.

Comparing gene expression profiles of AN subjects pre and post novel treatment to AN subjects undergoing conventional treatment and AN-free healthy controls will refute or support this premise.

This gene expression profile analysis could also validate the efficacy of perhaps the first documented behavioral epigenetics treatment and identify genes contributing to AN expression, opening the door for further research.

REFERENCES:

Becanu, S-A, Bulik, C.M., Klump, K.L., Fichter, M.M., Halmi, K.A., Keel, P.K., et al. (2005). Linkage analysis of anorexia and bulimia nervosa cohorts using selected behavioral phenotypes as quantitative traits or covariates. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics, 139, 61-68.

Barbarasz, M. (2000). Hypnosis in the treatment of eating disorders. In L.M. Hornyak & J.P. Green, Healing from within: The use of hypnosis in women’s health. Washington, D.C.: American Psychological Association.

Bergen, A., Van Den Bree, M., Yeager, M., Welch, R., Ganjei, K., Haque, K., Bacanu, S., Berrettini, W., Grice, D., Goldman, D., Bulik, C., Klump, K., Fichter, M., Halmi, K., Kaplan, A., Strober, M., Treasure, J., Woodside, B., Kaye, W.. (2003). Candidate genes for anorexia nervosa in the 1p33-36 linkage region: serotonin 1D and delta opioid receptors display significant association to anorexia nervosa. Molecular Psychiatry, 8 (4): 397-406.

Bulik, C.M., Sullivan, P.F., Tozzi, F., Furberg, H., Lichtenstein, P., Pederden, N.L. (2006), Prevalence, heritability, and prospective risk factors for anorexia nervosa. Archives of General. Psychiatry, 63, 305-312.

Bulik, C.M., Hebebrand, J., Keski-Rahkonen, A., Klump, K.L., Reichborn-Kjennerud, T., Mazzeo, S.E., Wade, T.D. (2007). Genetic epidemiology, endophenotypes and eating disorder classification. International Journal of Eating Disorders. 40(sup): s52-60.

Cheek, David B. (1994). Hypnosis: The Application of Ideomotor Techniques. Needham Heights, Massachusetts: Allyn & Bacon.

Bulik, C.M., Sullivan, P.F., Kendler, K.S. (1998). Heritability of binge-eating and broadly-defined bulimia nervosa. Biological Psychiatry, 44, 1210-1218.

Cheek, D. & LeCron, L. (1968). Clinical Hypnotherapy. New York: Grune and Stratton.

Cheek, D. (1994). Hypnosis: The Application of Ideomotor Techniques. Boston: Allyn and Bacon.

Darnaudéry, Muriel and Maccari, Stefania (2008). Epigenetic programming of the stress response in male and female rats by prenatal restraint stress. Brain Research Reviews Vol 57(2), 571-585.

DeSocio, J.E., O’Toole, J.K., He, H., Koeller, P., Baird, S.A., Lukach, M. (2008). The REDS-C Rating of Eating Disordered Severity Interview for Children and Adolescents: Clinical Usefulness and Comparison with the Modified EDI-2. Manuscript submitted for publication.

Devlin, B., Bacanu, S-A., Klump, K.L., Berrettini, W., Bergen, A., Goldman, D., et al. (2002). Linkage analysis of anorexia nervosa incorporating behavioral covariates. Human Molecular Genetics, 11, 689-696.

Frieling, H., Bleich, S., Otten, J., Romer, K.D., Kormhuver, J., de Zwaan, M., Jacoby, G.E., Wihelm, J., Hillemacher, T., (2008). Epigenetic downregulation of atrial natriuretic peptide but not vasopressin mRNA expression in females with eating disorders is related to impusivity. Neuropharmacology, 2, January: advance online: doi:10.1038/sj.npp.1301662.

Frieling, H., Gozner, A., Romer, K.D., Lenz, B., Bonsch, D., Wihelm, J., Hillemacher, T., de Zwaan, M., Kornhuber, J., Bleich, S. (2007). Global DNA hypomethylation and DNA hypermethylation of the alpha synuclein promoter in females with anorexia nervosa. Molecular Psychiatry, 12:229-230.

Goldner, E.M. (2000). The REDS rating of eating disorder severity. Unpublished instrument and administration guidelines. Vancouver, British Columbia, Canada: Simon Fraser University.

Grice, D.E., Halmi, K.A., Fichter, M.M., Strober, M., Woodside, D.B., Treasure, J.T., Kaplan, A.S., Magistretti, P.J., Goldman, D., Bulik, C.M., Kaye, W.H., Berrettini, W.H. (2002), Evidence for a susceptibility gene for anorexia nervosa on chromosome 1. American Journal of Human Genetics, 70 (3), 787-792.

Gunstad,J., Schofield, P., Paul, R.H., Spitznagel, M.B., Cohen, R.A., Williams, L.M., Kohn, M., Gordon, E. (2006). BDNF Val66Met polymorphism is associated with body mass index in healthy adults. Neuropsychobiology, 53 (3), 153-156.

Javaras, K.N., Laird, N.M., Reichborn-Kjennerud, T., Bulik, C.M., Pope, H.G.J., Hudson, J.I. (2008). Familiarity and heritability of binge eating disorder: Results of a case-control family study and a twin study. International Journal of Eating Disorders, 41, 174-179.

Kaplan, A. (2004). Exploring the gene-environment nexus in anorexia, bulimia. Psychiatric Times, 21 (9).

Kaplan, A.S., Levitan, R.D., Yilmaz, Z., Davis, C., Tharmalingam, S., James L. Kennedy, J.S. (2008). A DRD4/BDNF gene-gene interaction associated with maximum BMI in women with bulimia nervosa. International Journal of Eating Disorders, 41(1), 22-28.

Kartini Clinic for Disordered Eating: Portland, Oregon. www.kartiniclinic.com

Kas, M.J., VanElburg, A.A., VanEngeland, H. & Adan, R.H. (2003). Refinement of behavioral traits in animals for the genetic dissection of eating disorders. European Journal of Pharmacology, 480, 13-20.

Kendler, K.S., MacLean, C. Heale, M.C., Kessler, R.C., Heath, A.C., Eaves, L.J. (1991). The genetic epidemiology of bulimia nervosa. American Journal of Psychiatry, 148, 1627-1635.

Klump, K.L., Bulik, C.M., Kaye, W.H., Treasure, J., Tyson, E. (2009). Academy for eating disorders position paper: Eating disorders are serious mental illnesses. International Journal of Eating Disorders, 42(2), 97-103.

Klump, K.L., Culbert, K.M. (2007). Molecular genetic studies of eating disorders: Current status and future directions. Current Directions Psychological Science, 16, 37-41.

Klump, K.L., Miller, K.B., Keel, P.K., McGue, M., Iacono, W.G. (2001). Genetic and environmental influences on anorexia nervosa syndromes in a population-based twin sample. Psychological Medicine, 31, 737-740.

Lynn, S.J., Rhue, J.W., Kvarl, S., Mare, C. (1993). The treatment of anorexia nervosa: A suggestive framework. Contemporary Hypnosis, 10(2), 73-80.

Majdan, M., Shatz, C.J. (2006). Effects of visual experience on activity-dependent gene regulation in cortex. Nature Neuroscience 9, 650:650-659.

Martinowich, K., Hattori, D., Wu, H., Fouse, S., He, F., Hu,Y., Fan, G., Sun, Y.E., (2003). DNA methylation-related chromatin remodeling in activity-dependent bdnf gene regulation. Science, 302 (5646), 890 – 893.

Mercader, J. M., Ribasés, M., Gratacós, M., González, J.R., Bayés, M., de Cid, R., Badía, A., Fernández-Aranda,F., Estivill, X. (2007). Altered BDNF blood levels and gene variability are associated to anorexia and bulimia. Genes, Brain and Behavior ,6(8): 706-716.

Oberlander T.F. , Weinberg J., Papsdorf M., Grunau R, Misri S., Devlin A.M. (2008). Prenatal exposure to maternal depression, neonatal methylation of human glucocorticoid receptor gene (NR3C1) and infant cortisol stress responses. Epigenetics, 3 (2); 1-9.

O’Toole, J.K. (2000). The REDS-C rating of eating disorder severity interview for children. Unpublished instrument and administration guidelines. Portland, Oregon: The Kartini Clinic.

Owen,J.B., Treasure, J., Collier, D.A. (2001). Animal Models- Disorders of Eating Behavior and Body Composition. New York: Springer.

Polan HJ, Ward MJ. (1994). Role of the mother's touch in failure to thrive: a preliminary investigation. Journal American Academy Child Adolesc Psychiatry, 33(8):1098-105.

Reichborn-Kjennerud, T., Bulik, C.M., Tambs, K., Harris, J.R. (2004). Genetic and environmental influences on Binge eating in the absence of compensatory behaviors: A population-based twin study. International Journal of Eating Disorders, 36, 307-314.

Ribasés, M., Gratacòs, M., Fernández-Aranda, F., Bellodi, L., Boni, C., Anderluh, M., Cavallini, M.C., Cellini, E., Di Bella, D., Erzegovesi, S., Foulon, C., Gabrovsek, M., Gorwood, P., Hebebrand, J., Hinney, A., Holliday, J., Hu, X., Karwautz, A., Kipman, A., Komel, R., Nacmias, B., Remschmidt, H., Ricca, V., Sorbi, S., Wagner, G., Treasure, J., Collier, D.A. and Estivill, X. (2004), Association of BDNF with anorexia, bulimia and age of onset of weight loss in six European populations. Human Molecular Genetics, 13 (12), 1205-1212.

Ribasés, M., Gratacòs, M., Fernández-Aranda, F., Bellodi, L., Boni, C., Anderluh, M., Cavallini, M.C., Cellini, E., Di Bella, D., Erzegovesi, S., Foulon, C., Gabrovsek, M., Gorwood, P., Hebebrand, J., Hinney, A., Holliday, J., Hu, X., Karwautz, A., Kipman, A., Komel, R., Nacmias, B., Remschmidt, H., Ricca, V., Sorbi, S., Tomori, M., Wagner, G., Treasure, J., Collier, D.A., and Estivill, X. (2005). Association of BDNF with restricting anorexia nervosa and minimum body mass index: a family-based association study of eight European populations. European Journal of Human Genetics, 13, 428–434.

Ribases, M.; Gratacos, M.; Armengol, L.; de Cid, R.; Badia, A.; Jimenez, L.; Solano, R.; Vallejo, J.; Fernandez, F.; Estivill, X. (2003), Met66 in the brain-derived neurotrophic factor (BDNF) precursor is associated with anorexia nervosa restrictive type. Molecular Psychiatry, 8, 745-751.

Rossi, E.L. & Cheek, D.B. (1988). Mind-body therapy: Ideodynamic healing in hypnosis. New York: W.W. Norton & Co.

Rossi, E.L. (2002). The psychobiology of gene expression. New York, N.Y.: W.W. Norton & Company, Inc.

Rotenberg, K.J., Taylor, D. & Davis, R. (2004). Selective mood-induced body image disparagement and enhancement effects: Are they due to cognitive priming or subjective mood? International Journal of Eating Disorders, 35 (3), 317 - 332.

Schanberg, S. M., Ingledue, V. F., Lee, J. Y., Hannun, Y. A., and Bartolome, J. V. (2003). PKC Mediates Maternal Touch Regulation of Growth-Related Gene Expression in Infant Rats. Neuropsychopharmacology. 28, 1026-1030.

Segerstrile, U., Molner, P. (1997) Eds. Nonverbal Communication: Where Nature Meets Culture. Mahwah, New Jersey: Lawrence Erlbaum Associates, Publishers.

Siegfried, Z., Berry, E.M., Hao, S. & Avraham, Y. (2003). Animal models in the investigation of anorexia. Physiology and Behavior, 29, 39-45.

Strahl, DB., Allis, CD. (2000). The language of covalent histone modifications. Nature 403:41-45

Steiger, H., Guavin, L., Joober, R., Israel, M., YingKin, M.K.Ng., Bruce, K.R., Richardson, J., Young, S.N., Hakim, J. (2006). Intrafamilial correspondences on platelet[H-] paroxetine-binding indices in bulimic probands and their unaffected first-degree relatives. Neurophychopharmacology, 31:1785-1792.

Wade, T.D., Bulik, C.M., Neale, M., Kendler, K.S. (2000). Anorexia nervosa and major depression: Shared genetic and environmental risk factors. American Journal of

Walsh, B.J. (2005). Utilization Sobriety: A Substance Abuse Solution. Portland, Oregon: Peaceful Pilgrim Press: 125-144.

Walsh, B.J. (2008). Hypnotic alteration of body image in the eating disordered. American Journal of Clinical Hypnosis 50(4): 301-310.

Walsh, B.J. (2010). Rapid remission of anorexia nervosa and unconscious communication. American journal of Clinical Hypnosis, 52(4): 315-328.

Watkins, H.H. (1993). Ego-state therapy: An overview. American Journal of Clinical Hypnosis, 35(4): 232-240.

Watkins, J. G. & Watkins, H.H. (1997). Ego states: Theory and therapy. New York: W.W. W.W. Norton & Co..

Weaver, I.C.G., Cervoni, N.,, Champagne, F.A.,1, 2, Alessio, A.C.D., Sharma, S., Seckl, J.R., Dymov, S., Szyf, M., & Meaney, M.J. (2004). Epigenetic programming by maternal behavior. Nature Neuroscience, 7, 847 - 854.