The Project

PHASE 1: Mapping out the problem (0 months to 6 months)

Objectives: To derive estimates of the prevalence of MDD in patients with diabetes, rheumatologic disorders and COPD as examples of common LTCs and to identify genetic and psychosocial factors associated with resilience to MDD across several settings (routine care/AED) (Study 1A).

To standardize the RS-14, SOC-29 and RCOPE for use within the Greek population and to assess their potential in enhancing resilience during the current social and financial crisis.

Study 1A: The "resilience to MDD during social & financial crises prediction rule" study - Part I

At months 0-6 we will undertake a cohort study of 450 participants divided into three groups based on the assumed degree of illness-distress they experience: (a) patients with LTCs who seek care in the AED of the Ioannina General Teaching Hospital (high levels of illness-related distress); (b) patients with LTCs who attend routine care in follow-up specialty clinics (medium levels of illness-related distress), and (c) alleged healthy people derived from hospital's staff (low levels of illness-related distress), during a period of 6 months (cross-sectional study) (150 within each group), and we will proceed prospectively to follow-up this cohort for an additional 6-month period (Study 2A).
We will consecutively interview using the Greek version of the Mini International Neuropsychiatric Interview (MINI) (58) a minimum of 600 people (200 within each category) to ensure that we receive 450 satisfactory responses (150 within each category), given that our previous studies in the same hospital have shown a reasonable response rate both at interviews and self-report completion ranging from 65 to 85% [34,53]. The usual caseload of our A&E department is about 80 patients per day, and a review of the recent records of the department showed that patients with the diseases under study represent a percentage of approximately 10%. Given that our AED provides 24-hour service every other day, we expect a total number of approximately 700 (eligible/agreed and non-eligible/non-participant) patients during the 6-month period. On the other hand, our previous research with patients attending follow-up clinics demonstrates the feasibility of obtaining the required number of patients for each disease under study [34,53]. Recruitment of the alleged healthy individuals from our  hospital staff will take place following the previously described procedure in our study on the impact of the A/H1N1 influenza pandemic upon hospital's staff distress [46]. 
After the psychiatric interview we will administer the baseline questionnaires with full explanation of the project and we will take peripheral blood samples for the genetic analyses after specific for genetic analysis written informed consent was obtained. The baseline questionnaires will include measures of socio-demographic data, financial and employment status, reproduction rates, perceived impact of the current crisis, insurance status, immigration issues, family, social and state support, severity of illness (as appropriate), Illness perceptions (as appropriate), health care use, history of depression and other mental illnesses, and psychosocial background.The battery of the questionnaires will include instruments with known applicability in our research population based on our previous research in the field (see Scientific Supervisor's publications). More specifically, we plan to administer the standardized Greek versions of the following instruments: (a) PHQ-9, for the assessment of depressive symptoms [53], (b) the anxiety module of the SCL-90-R for the assessment of anxiety symptoms (59), (c) the Risk Assessment Suicidality Scale (RASS) for the assessment of suicidality risk (60), and (d) the World Health Organization Quality of Life instrument - Short-form (WHOQOL-BREF) for the assessment of HRQoL (61).The patients' illness perceptions will be assessed with the Brief Illness Perceptions Questionnaire (B-IPQ) 62), and we will administer instruments assessing factors relevant to resilience and coping capacities [i.e., RS-14 (63), SOC (64) and RCOPE (65)]. The latter 3 questionnaires will be standardized for the Greek population.
Genomic DNA will be extracted from peripheral blood samples using commercially available kit (QIAGEN, DNA BloodMidi kit).  Genotyping of the sequence variants of the genes of interest will be performed with established PCR-based methods that have been previously applied in the Department of Pharmacology [68,79]. Specifically, the amplification refractory mutation system (ARMS) [79] (66), or the restriction fragment polymorphism (RFLP) technique [68] (67) will be used.  The PCR reaction will contain human genomic DNA, MgCl2, dNTPs, primers, and Taq DNA polymerase, according to standard protocols [68,79]. Primers will be designed using the Primer3 express software (www-genome.wi.mit.edu/cgibin/primer/primer3_www.cgi). For the PCR-ARMS approach, 2 allele-specific primers and one common for each polymorphism will be designed.  When necessary, additional mismatches will be incorporated at the 3′ end in order to enhance the discrimination between the two alleles [79](66). The end products will be analyzed by electrophoresis and visualized under UV light. For the PCR-RFLP approach, PCR products will be digested with the appropriate restriction enzymes and digestion fragments will be analyzed by electrophoresis and visualized under UV light. We will study 3 categories of genes with their respective polymorphisms:

A. Genes of the serotonin system and their respective polymorphisms:
1. The serotonin transporter (5HTT) with a 44bp linked polymorphic region (5HTTLPR) in the proximal 5’ regulatory region of the gene.
2. Monoamine oxidase A (MAOA) with a variable number-tandem-repeat (VNTR) polymorphism consisting of a 30bp sequence present in 2, 3, 3.5, 4 or 5 repeats (R).
3. Catechol-O-methyltransferase (COMT) with a single nucleotide polymorphism (rs46805) and
4. HTR1A, 5-HT2A polymorphisms (more than one according to the current literature status)

B. Genes of the HPA axis and their respective polymorphisms:
1. The corticotropin releasing hormone (CRH) receptor 1 (CRHR1)  and the glucocorticoid receptor (GR)  with more than one single nucleotide polymorphisms according to the current literature.

C. Neurotrophins and housekeeping genes and their respective polymorphisms:
1. Brain derived neurotrophic  factor (BDNF) with the Val66Met substitution (rs6265).
2. Housekeeping HSP70 (A8) and HSP90 genes, with more than one single nucleotide polymorphisms according to the current literature status.

Hardy–Weinberg equilibrium, minor allele frequency and pair-wise linkage disequilibrium (LD) between SNPs will be computed using Haploview 4.2 software. In this context, Haplotypes analyses will be performed according to the results of Linkage Disequilibrium. Given that statistical analyses will be conducted using several SNPs and non genetic variables the possibility of false-positive results has to be addressed and it will be taken into account.  It should be mentioned that possibly  some  polymorphisms will not have actual allelic / genotypic variation. This means that less statistical analyses will be finally performed, a fact that leads to minimization of false-positive results.

The genotyping and the psychosocial measures will be taken place over a 6 month period and then the main outcomes (PHQ-9, SCL-90-R Anxiety scale, RASS, RS-14, and WHOQOL-BREF) will be followed up over the subsequent 6 months to assess changes between baseline and follow-up.

Study 1B (months 0-6): Standardization of the RS-14, SOC-29 and RCOPE
We will test the psychometric properties (dimensionality, internal consistency, convergent and concurrent validity) of the RS-14, SOC-29 and RCOPE using the methods developed by our research team for the assessment of the psychometric properties of the Defense Style Questionnaire [41], the Life Style Index [48] and the Zuckerman-Khulman Personality Questionnaire [76].

Output
WP1. Estimation of the prevalence of MDD and the level of resilience in Greek patients with diabetes, rheumatologic disorders and chronic obstructive pulmonary disease (COPD) attending a routine clinic or AED and comparison with alleged healthy individuals.
WP2. Identification of the genetic profile and the psychosocial factors associated with MDD and resilience in patients with these 3 long term conditions compared to alleged healthy individuals.
WP3. Standardized versions of the screening for resilience (RS-14), sense of coherence (SOC-29) and religious coping (RCOPE) inventories.

PHASE 2: Identifying which people to target (months 6-12)

Objectives: To fully develop and test a clinical prediction rule that will identify predictors of MDD and factors predicting resilience to MDD and social and financial crisis in patients with LTCs.

Study 2A: The "resilience to MDD during social & financial crises prediction rule" study – Part II
After six months (months 6-12) we will administer a further brief questionnaire to assess health services use, resilience, depressive and anxiety symptom severity, suicidal risk and HRQoL. Data will be obtained by re-interviewing the participants involved in the cross-sectional study either at the hospital or by telephone. This will establish the variables that predict depression and well-being.

Statistical analysis plan to identify predictors of resilience to/risk for MDD.
A clinical predictor rule for MDD and resilience will be developed using logistic and linear regression employing a variety of selection algorithms in the search for effective explanatory variables. Internal validation of the models will also be performed using an appropriate statistical method (i.e., boot-strapping analysis).  Any moderating or mediating effects of variables of interest (i.e. gene polymorphism, sex, financial deprivation, unemployment, etc.) upon outcome (MDD) and the additive moderating effects of resilience and SOC will also be tested using Sobel tests based also on the methods proposed by Baron & Kenny (68), and Miles & Davis (69).

Sample Size and Power
Our main outcome is the proportion of participants diagnosed with MDD. Based on evidence indicating that the prevalence of the putative risk factor (MDD) in patients with LTCs is between 20% and 30% [11,70], we considered the maximum prevalence of the putative risk factor at baseline at 25%-30%. Given error margins of 5% and 95% confidence intervals, the sample size required to powerfully detect MDD prevalence between 25% and 30% (±5%) (i.e. 20%-35%) was a-priori estimated at approximately 300 participants. Concerning intended multiple regression analyses with up to 12 predictors for each subsample, a desired significance level (alpha) of 0.05, a medium anticipated effect size (f2=0.15) and a desired power of 0.8, the required sample sizes for each subsample was a-priori estimated at approximately 130 participants.

Output
WP4. A clinical prediction rule for identifying resilience factors for MDD, focusing on those participants on whom the impact of the current social and financial crisis was estimated as highly important.

PHASE 3: Developing a pilot low-intensity intervention (months 9-12)

Objectives: To develop a pilot intervention that will guide future research focusing on enhancing resilience to depression and reducing the impact of the social and financial crises on mental health and well-being. Due to the very limited available time, our intention is to provide guidance to future research to develop a preventative intervention.

Study 3. An intervention to prevent MDD and enhance resilience (months 9-12)
The purpose of this part of the programme is to provide an evidence-based guidance for developing a credible, practical, low intensity intervention to be used in conjunction with the clinical prediction rule. In other words, what should happen when a patient is identified by the clinical prediction rule, as being potentially depressed.

Patient sample. Using the criteria and procedures of Studies 1A & 2A, we shall purposively sample from each disease group about 10 at risk for MDD individuals for interview, using the definition of "vulnerable to depression" established by the study 2A. We will also select 5 individuals with low MDD risk in each disease group for interview about how they manage the kinds of factors that drive to psychological distress, focusing on factors relevant to the current social and financial crisis. 

Patient interviews and intervention. For individuals at risk, interviews will identify their own reasons for their condition by prompting them to describe their illness-perceptions and their experience of health care in the era of current recession. To avoid generalised or idealised accounts, interviews will focus on specific instances where a coping strategy was sought, considered or avoided. We cannot be specific about the nature of the intervention at this stage, but it is likely to involve a whole systems approach. We will introduce problem solving and cognitive strategies to enhance self management techniques, lifestyle changes including diet and exercise and other forms of intervention that will be determined as the programme develops, using also several formats including for example a book, CDROM, or via a website.

Output:
WP3. A detailed and practical understanding of who is resilient to or at risk for MDD in relation to the current social and financial crisis, what they experience, and what are the major therapeutic factors for enhancing resilient and reducing depressive symptomatology. Our programme should demonstrate the feasibility of introducing the clinical prediction rule care intervention into the medical setting.
WR4. Dissimination (months 0-12). We plan that there will be a continuous process of dissemination of the findings of the research as the programme develops. We have allocated specific monies for dissemination throughout the programme and we plan to hold a major conference on ‘Depression in chronic illness' at the end of the programme to highlight our findings. We will also produce relevant papers for publication in open access scientific journals.