Here's a Journal Club Presentation I made on the article 'Genetic Predisposition to Schizophrenia Associated with Increased Use of Cannabis' which appeared in the Sep 14 issue of Molecular Psychiatry. It examined and found a correlation between shortened telomere length in women and a risk for depression in their daughters.

1. Telomere Length And Cortisol Reactivity In
Children Of Depressed Mothers
IH Gotlib
J LeMoult
NL Colich
LC Foland-Ross
J Hallmayer
J Joormann
J Lin
OM Wolkowitz
Molecular Psychiatry advance online publication, 30
September 2014; doi:10.1038/mp.2014.119 1

2. Overview
• Introduction
• Telomeres
• HPA-Axis derangement in Major Depressive Disorder (MDD)
• Related studies
• Article
Aim
Material and Methods
Results
Discussion
Limitations 2

3. Molecular Psychiatry
• “Molecular Psychiatry publishes work aimed at elucidating biological
mechanisms underlying psychiatric disorders and their treatment. The
emphasis is on studies at the interface of pre-clinical and clinical
research, including studies at the cellular, molecular, integrative,
clinical, imaging and psychopharmacology levels”
• Impact Factor: 15.147*
• Rank:
1/135 Psychiatry
5/251 Neuroscience
4/291 Biochemistry & Molecular Biology
• *2013 Journal Citation Report (Thomson Reuters, 2014)
3

4. Introduction - Telomeres
• Regions of repetitive nucleotide sequences at each end of a
chromatid
• Vertebrates – TTAGGG
1. Prevent chromosomes from losing base-pair sequences at their ends
2. Prevent chromosomes from fusing / oxidative stress
• Disposable buffers at the ends of chromatids
– Truncated during cell division
– Protect the genes before them on the chromatids from being truncated
instead
The 2009 Nobel Prize in Physiology or Medicine - Press Release
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9. Introduction – HPA Axis Dysregulation in MDD
9

10. Introduction – HPA Axis Dysregulation in MDD
• CRH & corticosteroid levels
remain elevated
• NE, 5-HT, DA and GABA
levels decrease
• HYPERCORTISOLEMIA
• ↓ inhibitory 5-HT tone
• ↑ drive from NE, ACh or CRH
• ↓ feedback inhibition from
Hippocampus
• Comprehensive Textbook of Psychiatry, 9th
Ed Kaplan & Sadock
10

11. Related studies - 1
Leukocyte Telomere Length in Major Depression:
Correlations with Chronicity, Inflammation and Oxidative
Stress - Preliminary Findings
AIM : To determine if
1. Leucocyte telomeres are shortened in Major Depressive Disorder
(MDD)
2. Whether this is a function of lifetime depression exposure
3. Whether this is related to putative mediators, oxidation &
inflammation
• METHOD
– Leucocyte telomere length compared in 18 unmedicated MDD
subjects & 17 controls
– Correlated with Lifetime depression chronicity, peripheral markers
of oxidation & inflammation
• Wolkowitz OM, et al. (2011) Leukocyte Telomere Length in Major Depression: Correlations with Chronicity,
Inflammation and Oxidative Stress - Preliminary Findings. PLoS ONE 6(3): e17837. doi:10.1371/journal.pone.0017837
11

12. Related studies - 1 (contd…)
• Results
– Depressed group (as a whole) did not differ from controls in
telomere length
– Telomere length was significantly inversely correlated with lifetime
depression exposure (even after age control) p<0.05
• Avg telomere length in depressed subjects above median of lifetime
depression exposure was 281 BPs shorter than controls ≈ 07yrs of
accelerated cell aging
• Conclusion
– Indication that accelerated aging at cellular level is proportional to
lifetime exposure to MDD
– Suggests that telomere shortening does not antedate depression
• Telomere shortening may progress in proportion to lifetime
depression exposure
12

13. Related studies - 2
Major depressive disorder and accelerated cellular aging :
results from a large psychiatric cohort study
• AIM : To determine whether
1. MDD is associated with accelerated biological aging
2. Depression characteristics (severity, duration, medication) further
impact biological aging
• METHOD
– TL assessed using Telomere sequence copy number (T) compared
to single-copy gene copy number (S) using quantitative PCR
• 1095 current MDD patients
• 802 remitted MDD patients
• 510 Control subjects
• JE Verhoeven Molecular Psychiatry advance online publication, 12 November 2013; doi:10.1038/mp.2013.151
13

14. Related studies - 2 (contd…)
• Results
– Compared with control subjects (mean bp=5541)
• TL was shorter in remitted MDD patients (mean bp=5459, p=0.014)
• TL was shorter in current MDD patients (mean bp=5461, p=0.012)
– Within current MDD patients, shorter TL was associated with both
• Higher depression severity (p<0.01)
• Longer symptom duration in the past 04 years (p=0.01)
• Conclusion
– Depressed patients show accelerated cellular aging according to a
dose-dependent gradient
– Exposure to depression leaves an imprint
• Remitted MDD patients had shorter TL than control
14

15. Related studies - 3
The Association of Major Depression, Conduct Disorder, and Maternal
Overcontrol with a Failure to Show a Cortisol Buffered Response in 4-
Month-Old Infants of Teenage Mothers
• AIM : To explore whether MD (lifetime, during pregnancy,
postpartum), CD, and maternal over-control are associated with
increased cortisol reactivity in 4-month-old infants of teenage
mothers
• METHOD
– Between 2 and 4 months, a normal shift occurs in the
adrenocortical system
• marked decrease in infant cortisol response when facing mild stressors
– Using arm restraint as a stressor, morning salivary cortisol was
taken pre-stressor and post-stressor in 212 infants
• R. Azar et al.The Association of Major Depression, Conduct Disorder, and Maternal Overcontrol with a Failure to Show a
Cortisol Buffered Response in 4-Month-Old Infants of Teenage Mothers Biol Psychiatry 2007;62:573–579
15

16. Related studies - 3 (contd…)
• Results
– Independent of the predictors, there was no cortisol response to arm
restraint in 4-month-old infants
– Cortisol of infants whose mother had a lifetime history of MD
significantly increased from t1 to t2 [from .20 to .29 μg/dL, p< .01]
– Infant cortisol was NOT related to
• MD Severity/ Depression during pregnancy / Post-partum depression
• Conclusion
– Lower baseline levels of cortisol (T1) in infants of mothers with history of
MD may partly explain their increased post-stressor cortisol measure
– Finding on lifetime MD may indicate a biological risk for stress reactivity
– Possible that it is not history of MD that is linked to stress reactivity but
rather other associated factors like prenatal stress or anxiety
16

17. Related studies - 4
Reduced telomerase activity in human T lymphocytes
exposed to cortisol
• AIM : To determine whether cortisol might affect telomerase activity
in immune cells
• METHOD
– Cellular telomerase activity measured using the Telomeric repeat
amplification protocol (TRAP) assay
• Results expressed as “Total Product Generated”, TPG, which reflects the
telomerase activity in the sample
• Calculated according to a computer program which integrates signal
intensity over the telomere length distribution on the gel as a function of
molecular weight
• Choi J et al. Reduced telomerase activity in human T lymphocytes exposed to cortisol. Brain Behav Immun 2008; 22:
600–605. 17

18. Related studies - 4 (contd…)
• Results
– Exposure of human T lymphocytes to cortisol associated with a
significant reduction in telomerase activity both during primary
stimulation of resting cells and secondary stimulation of previously
activated cells.
• The effect is observe in both CD4 and CD8 T
– Hydrocortisone in physiological ranges of cortisol had no effect on the
telomerase activity
– Higher concentrations of hydrocortisone, comparable to those that might
be reached in vivo during stress, reduced telomerase activity by as much
as 50%.
• Conclusion
– Findings provide a potential mechanism for stress-associated
telomere length attrition
18

19. 1. Telomere shortening α dose dependent lifetime
depression exposure
2. Persons at risk for depression manifest HPA-axis
dysregulation (exaggerated activation to stress)
3. Telomere shortening is assoc with low Telomerase
activity, which may be seen on exposure to high
levels of cortisol
19

20. Telomere Length And Cortisol Reactivity In
Children Of Depressed Mothers
• IH Gotlib
• J LeMoult
• NL Colich
• LC Foland-Ross
• J Hallmayer1
• J Joormann
• J Lin
• OM Wolkowitz
• Molecular Psychiatry advance online publication,
30 September 2014; doi:10.1038/mp.2014.119 20

21. Introduction
AIM
To assess telomere length in individuals who are at
high risk of developing MDD but who have not yet
experienced a depressive episode
• Shortened telomere length is a pre-existing
condition or risk factor for developing MDD
vs
• Shortened telomere length is a response to or
concomitant of major depressive episodes
21

22. Materials & methods – Subjects & Clinical Assessment
• 97 girls aged 10-14yrs with no current or past Axis 1 dx
– 47 = mothers with no current or past Axis 1 dx (CTL)
– 50 = mothers with h/o recurrent depressive episodes during
daughter’s lifetime (RSK)
• Daughters administered Kiddie Schedule for Affective
Disorders and Schizophrenia (KSADS-PL)
– assesses current and past episodes of psychopathology in children
and adolescents according to DSM-III-R and DSM-IV criteria
– Interviews of parent(s) & child
– Final summary ratings which include all sources of information
(parent, child, school, chart, and other)
22

23. Materials & methods – Subjects & Clinical Assessment
• Mothers administered SCID-IV
– Recurrent episodes of MDD incl in RSK
– Absence of current or past Axis 1 dx in CTL grp
• Random evaluation of 10% of SCID-IV & KSADS interviews
by independent rater blind to group membership
• In all cases diagnoses of the following matched original
interviewer’s diagnosis (k=1)
– Recurrent Depressive Episodes in RSK mothers
– Absence of Axis 1 disorder in CTL mothers
– Absence of Axis 1 disorder in daughters
• Eligible daughters also completed
– 10-item version of Children’s Depression Inventory
– Pubertal status via Tanner Staging 23

24. Materials & methods – Telomere Assessment
• Genomic DNA purified from 500μl of saliva
• Telomere length commonly assessed in leucocytes
– Significant positive correlation (0.72, p=0.002) between leucocyte telomere
length and telomere length measured in saliva
For each sample of DNA
determine the
factor by which
the sample
differs from
a reference DNA sample
in its
Ratio of Telomere copy number
to Single copy gene copy
number (T:S)

25. Telomere Repeats in each
sample
The level of dilution of a
reference DNA sample
that would
make the experimental and
reference samples equivalent
WRT
the number of PCR cycles
needed to generate
a given amount of
telomere PCR product
during
exponential phase of PCR
amplification
Single copy gene in each
sample
The level of dilution of a
reference DNA sample
that would
make the experimental and
reference samples equivalent
WRT
the number of PCR cycles
needed to generate
a given amount of
single copy gene PCR product
during
exponential phase of PCR
amplification
25

26. Materials & methods – Stress Task
26
• 30mins Rest period
• Part 1 – 3mins serial subtraction task (reverse serial 7 starting @ 400)
• Part 2 – 12mins Ewart Social Competence Interview
– 6 cards detailing stressor categories (school, work, family, friends, money,
neighborhood) and common subjects given by peers
– Determined why stressful
– Reconstruct a particular situation & probe
– Imagine hypothetical solution
• Neutral video
• Saliva samples imdtly before and at 15, 30 and 45mins after onset
– Peak cortisol response 21-40mins after onset of stressor
– Cortisol recovery to baseline 41-60mins after onset of stressor

27. Results
• Participant Characteristics
– RSK and CTL groups did not differ wrt age, ethnicity or
Tanner stage (all p>0.05)
– RSK group had slightly but significantly higher scores on
CDI than CTL group BUT scores of both groups were well
below clinical cutoff of 8
– Both within groups and across groups, Telomere length
was not significantly correlated with age, Tanner Stage
or CDI scores
27

28. Results - Telomere Length
• Girls in RSK group had
significantly shorter
telomeres than girls in the
CTL group (M=1.524T/S vs
1.754T/S, p=0.001)
• Results did not significantly
change when age, Tanner
stage and CDI were
included as covariates
28

29. Results – Cortisol Response to Stress
• Baseline cortisol levels did
not differ significantly as a
function of telomere length,
risk group or their
interaction
• Indls with shorter telomeres
exhibited significantly
greater cortisol reactivity to
stress
• No main effect of risk group
on reactivity
• Cortisol recovery was not
affected by TL, risk group,
their interaction 29

30. Results
1. Telomere length was not significantly correlated
with age, Tanner stage or CDI scores
2. Girls in the RSK group had significantly shorter
telomeres than girls in the CTL group
3. Girls with shorter telomeres exhibited significantly
greater cortisol reactivity to stress in both RSK and
CTL
4. Cortisol recovery from stress was not affected by
telomere length, risk group or their interaction
30

31. Discussion
• Healthy children at familial risk for depression have shorter
telomeres than their non-risk peers
– Hence telomere shortening seems to be antecedent to and
potentially a risk factor for onset of depression
• Genetic v/s Environmental – Why has shortening taken
place?
– Genetic
• Operationalized risk based on h/o recurrent depressive episodes in
mother
– Environmental
• All girls stayed with mothers
• Maternal depression is assoc with a stressful early environment for
children
Lovejoy MC, Graczyk PA, O’Hare E, Neuman G. Maternal depression and parenting behavior: a meta-analytic review.
Clin Psychol Rev 2000; 20: 561–592. 31

32. Discussion
• Nature of the assocn betn TL and cortisol
secretion still unclear
– Cortisol inhibits telomerase in vitro
– Telomere shortening is a precursor to depression ,
condition char by HPA Axis deregulation
• ? Possible a 3rd variable may be responsible for
both
32

33. Discussion
• Telomere shortening is not only a marker of stress, but also
a mechanism of biological aging
• Medical illnesses observed most frequently in indls with
psychiatric conditions are those most commonly seen with
advanced age (e.g. Cardiovascular Dx, stroke, dementia)
• Blackburn EH. Telomere states and cell fates. Nature 2000; 408: 53–56.
• Evans DL et al. Mood disorders in the medically ill: scientific review and recommendations. Biol Psychiatry 2005; 58:
175–189.
• Possibility that psychiatric illnesses are associated with
accelerated aging at cellular/organismic level
33

34. Limitations
1. Narrow age range of participants (10-14yrs)
– Cannot examine whether findings extend to a broader
age range of indls at risk for depression
2. Girls in RSK grp had slightly but significantly higher
CDI scores
3. Superficial nature of Tanner staging as a measure
of pubertal status
34

35. Limitations
• Numerous confounders are completely ignored
– Life events, Socio-economic status
• Repetitive
35

36. Checklist for assessing the quality of
quantitative studies
Criteria Yes(2) Partial(1) No(0) N/A
1 Question/ objective
sufficiently described? √
2 Study design evident &
appropriate? √
3 Method of
subject/comparision
group selection or
source of information/
input variable describe
and appropriate?
√
4 Subject ( and
comparison group, if
applicable)
characteristics
sufficiently described?
√
36

37. Yes (2) Partial (1) No (0) NA
5 If interventional and random allocation
was possible, was it described? √
6 If interventional and blinding of
investigators was possible, was it
reported?
√
7 If interventional and blinding of subjects
was possible, was it reported? √
8 Outcome and (if applicable) exposure
measures well-defined and robust to
measurement/misclassification bias?
Means of assessment reported
√
37

38. Yes (2) Partial (1) No (0) NA
9 Sample size appropriate?
√
10 Analytic methods
described/justified and
appropriate?
√
11 Controlled for confounding?
√
12 Results reported in sufficient
detail? √
13 Conclusions supported by the
results? √
38

39. References
• Comprehensive Textbook of Psychiatry, 9th Ed Kaplan & Sadock
• Choi J et al. Reduced telomerase activity in human T lymphocytes
exposed to cortisol. Brain Behav Immun 2008; 22: 600–605.
• JE Verhoeven Molecular Psychiatry advance online publication, 12
November 2013; doi:10.1038/mp.2013.151
• Wolkowitz OM, et al. (2011) Leukocyte Telomere Length in Major
Depression: Correlations with Chronicity, Inflammation and Oxidative
Stress - Preliminary Findings. PLoS ONE 6(3): e17837.
doi:10.1371/journal.pone.0017837
• Cawthon RM Telomere measurement by quantitative PCR Nucleic
Acids Research 2002, Vol 30 No 10 39

40. Thank you
40