about lithium use in elderly with mild cognitive impairement

1. CLINICAL AND BIOLOGICAL EFFECTS OF LONG-TERM
LITHIUM TREATMENT IN OLDER ADULTS WITH
AMENESTIC MILD COGNITIVE IMPAIREMENT :
RANDOMISED CLINICAL TRIAL
DR.ANUSUYA.M

2. AUTHORS
• 1) Orestes V. Forlenza : Professor and HOD in the Department of Psychiatry at
FMUSP(Geriatric Psychiatry and Neurosciences).
• 2) Márcia Radanovic : Neurologist, Department of Psychiatry, University of São
Paulo. (Cognitive Neurology and Psychiatry, Language disturbances in MCI
patients)
• 3 ) Leda L. Talib : Department and Institute of Psychiatry, University of São
Paulo School of Medicine Hospital , Brazil.
• 4) Wagner F. Gattaz : Laboratory of Neurosciences (LIM-27), Department and
Institute of Psychiatry, University of São Paulo and School of Medicine
Hospital, Brazil.

3. ABOUT THE JOURNAL
• Article was published in THE BRITISH JOURNAL OF PSYCHIATRY IN 2019.
• Peer-reviewed journal, covering all branches of psychiatry established in 1853.
• The journal is owned by the Royal College of Psychiatrists and published monthly
by Cambridge university.
• Impact factor : 10.6 (2023)
• Editor in chief : Professor Gin Malhi, University of Sydney, Australia.

4. INTRODUCTION
• Experimental studies provide evidence that lithium possess potential neurotrophic
and neuroprotective properties, and can be used for the treatment and prevention
of neurodegenerative disorders.
• Epidemiological and imaging studies in bipolar disorder show that long-term
lithium treatment is associated with lower rates of dementia and has beneficial
brain responses, such as increased grey matter density and better metabolic
integrity of the cerebral tissue.

5. INTRODUCTION
• A recent, nationwide, population-based study conducted in Denmark indicated a
negative association between trace lithium in ground water and the incidence of
dementia across different geographical regions.
• Lithium concentrations in drinking water in several counties in Texas, USA were
negatively associated with mortality rates as a result of Alzheimer's disease, this
fact supports consideration of lithium as a disease modifier in Alzheimer’s disease.
• A meta-analysis of data from three clinical trials which tested the effects of lithium
in patients with dementia showed that lithium treatment may indeed be associated
with a significant decrease in cognitive decline.

6. OBJECTIVE OF THE STUDY
To Determine the potential clinical and biological benefits
of low-dose, long-term lithium treatment for patients with
amnestic mild cognitive impairment (MCI).

7. MILD COGNITIVE IMPAIREMENT
• A clinical condition that comprises the transitional state between normal cognitive
ageing and incipient dementia.
• Associated with a high risk of Alzheimer's disease.
• In persons over age 65 years, AD is the most common etiology of MCI and mild
dementia.
• Amnestic impairment is most typical for AD in the stage of MCI or mild dementia .
• Low performance is seen in one or more cognitive domains.(1) Learning and
memory, 2) Language, 3) Visuo-spatial, 4) Executive and 5) Psychomotor activity)

9. Alzheimer’s disease (AD)
• Amyloid pathology -characteristic of Alzheimer’s disease (AD), results from altered
metabolism of the beta-amyloid (Aβ) peptide in terms of synthesis, clearance, or
aggregation.
• A decrease in cerebrospinal fluid (CSF) level Aβ1–42 is evident in AD.
• Neuropathological brain lesions consist of aggregates of hyper-phosphorylated tau
proteins (neurofibrillary tangles).
• Cerebrospinal fluid (CSF) phosphorylated tau (p‐tau) is one of the core biomarkers
for Alzheimer’s disease (AD).
• Tau phosphorylated at threonine 181 (p-tau181) measured in blood plasma has
recently been proposed as an accessible, scalable, and highly specific biomarker for
Alzheimer’s disease.

10. MATERAILS AND METHODOLOGY
• STUDY DESIGN AND DURATION:
• Double - Blinded Randomized controlled trial ( 2 years) and 1 year single- blinded
phase was done.
• Followed by a single-blinded extension phase for an additional 24 months.
• Trial registration was done at clinicaltrials.gov: NCT01055392.
• An interim analysis of this trial, addressing the modification of clinical and
biological outcome variables after 12 months in a smaller sample was published
prior in 2011.

11. RCT
Prospective experimental design that randomly assigns participants to an
experimental or control group.
RCTs are the gold standard for establishing causal relationships and ruling out
confounding variables and selection bias.
Permuted-block randomization or blocked randomization: a "block size" and
"allocation ratio" (number of subjects in one group versus the other group) are
specified, and subjects are allocated randomly within each block.
Stratified randomization - Method of sampling which first stratifies the whole
study population into subgroups with same characteristics (strata), then followed
by simple random sampling from the stratified groups, where each element within
the same subgroup are selected unbiasedly during any stage of the sampling process,
randomly and entirely by chance..

12. Double-blinded randomized controlled
trial
• A double-blinded randomized controlled trial - Type of clinical trial where
neither the participants nor the researchers know which treatment group
each participant belongs to.
• Prevent any bias in the study and ensures that the results are as objective as
possible

13. MATERAILS AND METHODOLOGY
• Study was conducted in an academically oriented psychogeriatric
service.
• At a tertiary public hospital in Sao Paulo, Brazil.
• The study protocol was approved by the local Ethics Committee
(CAPPesq-HCFMUSP).
• Conducted within the principles of the Helsinki Declaration and Good
Clinical Practice recommendations.

14. Participants
COMMUNITY DWELLERS AROUND THE HOSPITAL CATCHMENT AREA
WERE RECRUITED
OLDER PATIENTS ≥60YRS WITH NORMAL TO MILD COGNITION
IMPAIREMENT / DEMENTIA
SINGLE PHYSICIAN NOT TAKING PART
TOTAL 106 INVITED
ELIGIBLE- 76
TOTAL ENROLLED - 61
15 DECLINED PARTICIPATION
NO H/O MAJOR PSYCHIATRIC
ILLNESS/UNCONTROLLED MEDICAL ILLNESS.

16. INCLUSION AND EXCLUSION CRITERIA
The inclusion criteria :
• Age ≥60 years
• Diagnosis of amnestic MILD COGNITIVE IMPAIREMENT.
Exclusion criteria :
 Clinical history of major psychiatric disorders.
 Evidence of relevant/uncontrolled medical diseases.

17. PROCEDURE
• Participants were enrolled after obtaining informed consent.
• A single physician who did not take part in the assessment of baseline or
outcome variables performed the recruitment, diagnostic screening and
allocation of eligible participants into study groups, in addition to
prescribing medications throughout the trial.
• Ssystematic examination was done by a comprehensive clinical and
neuropsychological protocol.
• Group allocation, was done using permuted blocks and stratified
randomization method with account of age and education levels.

18. PROCEDURE
• Participants were randomized into experimental (lithium) or comparison
(placebo) groups (1:1 ratio).
• Participants were longitudinally reassessed at 3-month intervals by
raters who were unaware of group allocation.
• An independent geriatrician systematically performed physical
examination and administered the 56-item UKU Side Effect Scale.
• Prior to each visit for safety monitoring peripheral blood samples were
obtained and a panel of laboratory tests were performed.
• Additional tests as per clinician judgement was also done.

19. PROCEDURE
• Lumbar puncture was done by an experienced neurologist at baseline and after 12
and 36 months for the collection of cerebrospinal fluid (CSF) samples.
• The third lumbar puncture was postponed for 12 months, instead of being
performed at the end of the double-blind phase, for the sake of tolerability and
adherence, warranting a longer interval between subsequent procedures, and to
better account for long-term changes.
• Alzheimer's disease-related CSF biomarkers were determined by commercially
available kits (INNo-BiaAlzBio3 assay, Innogenetics, Ghent, Belgium) using a
multiplexed method (xMAP-Luminex).

20. PROCEDURE
• Identical tablets holding 150 mg, 300 mg, 450 mg or 600 mg of lithium carbonate or
placebo packaged in identical coded blisters, were prescribed by a dedicated
pharmacist.
• Treatment was started with a single daily tablet of lithium carbonate 150 mg or
placebo taken orally in the evenings for a week.
• Participants in the comparison group were instructed to take a second placebo
tablet in the morning from the second week onwards.
• For participants in the lithium group, daily doses were titrated by adding a second
lithium (150 mg) or placebo tablet in the morning, and then adjusted to target
serum levels at weekly visits, using distinct combinations of lithium/placebo tablets
(one in the morning and one at night) to fulfil total doses of 150 mg, 300 mg,
450 mg or 600 mg per day

21. PROCEDURE
• Target lithium levels were defined at a subtherapeutic window (0.25 –0.5 mEq/L).
• Serum lithium levels were determined weekly in the titration phase and at 3-
month intervals throughout the study.
• Patients were instructed to report any symptoms suggestive of adverse events .
• In cases of occurrence of relevant side-effects, the lithium dose was tapered down
to the highest previously tolerated dose within the target window.
• Once stable lithium levels were achieved, the prescription was maintained for the
following 3 months.

22. OUTCOME
• Primary outcome:
• Changes in cognitive and functional parameters during the double-blind
phase of the study (end-point 2 years).
1. Alzheimer's Disease Assessment Scale (ADAS-Cog) – Global cognitive
state.
2. Clinical Dementia Rating (CDR) Sum of Boxes (SoB) – Functional
Performance.

23. Secondary outcomes
• The changes in neuropsychological test scores with emphasis on
memory, attention and executive functions.
• Changes in CSF concentrations of the amyloid-beta peptide (Aβ1−42),
total tau and phosphorylated tau at threonine 181;
• Conversion from MCI to dementia;
• Safety/tolerability data (published elsewhere)
• Changes in peripheral biomarkers. ( Planned to be presented in a future
publication).

24. SCALES
Alzheimer’s Disease Assessment Scale-Cognitive Subscale (ADAS-Cog):
• Brief neuropsychological assessment scale used to assess the severity of cognitive
symptoms of dementia.
• Most widely used cognitive scales in clinical trials.
• Considered to be the “gold standard” for assessing antidementia treatments .
• ADAS-Cog consists of 11 parts and takes approximately 30 minutes to administer.

25. ADAS-Cog consists of 11 questions:
• Word Recall Task
• Naming Objects and Fingers
• Following Commands
• Constructional Praxis
• Ideational Praxis
• Orientation
• Word Recognition Task
• Remembering Test Directions
• Spoken Language
• Comprehension
• Word-Finding Difficulty

26. The Clinical Dementia Rating (CDR) Sum of Boxes
(SoB)
• Used to stage the severity of dementia.
• It is a more detailed and provides more information than the global CDR score in
patients with mild dementia.
• The CDR-SOB is calculated by summing the scores of six domains: memory,
orientation, judgment and problem-solving, community affairs, home and hobbies,
and personal care.
• The scores range from 0 to 18, with higher scores indicating greater severity of
dementia.
• CDR-SOB is a copyrighted instrument and its use requires a license.

27. STATISTICS
• Differences in the distribution of demographic, clinical and biological variables
between lithium and placebo groups at baseline were statistically examined using
chi-squared (χ2), Fisher's and independent sample Student's t-tests.
• Interim analyses of primary outcome variables were performed with data collected
annually.
• The analysis of cognitive and functional outcome variables was limited to the
double-blind phase and therefore included data collected at baseline and at 12
months and 24 months of follow-up.

28. STATISTICS
• Analysis of Alzheimer's disease-related biomarkers was based on CSF samples
collected at baseline and after 12 and 36 months.
• Linear mixed effects model was used to determine the longitudinal changes in
cognitive, functional and biological outcome variables.
• Group allocation (treatment), duration of the intervention (time) and
treatment/time interactions at distinct time points were taken in to consideration.
• Paired sample t-tests were additionally used to address differences in continuous
variables between baseline and end-point within treatment groups.

29. STATISTICS
• Non-parametric tests (Mann-Whitney U-test and Wilcoxon rank test) done for
analysis of biological data in sub analyses with limiting sample sizes.
• Normative data generated were further used to classify subgroups of patients
according to baseline amyloid burden (‘low-’ or ‘high-’ CSF Aβ1−42) using
416 pg./mL as a threshold.
• Analysis of conversion from MCI to Dementia was done after full period of the trial
(48 months), considering the possibility that this outcome might take longer to
occur.

30. STATISTICS
• Kaplan–Meier curves were built to compare the conversion rates in
lithium versus placebo groups, censoring the changes in functional state
according to modifications in CDR scores (from CDR <1 to CDR ≥1).
• Statistical analyses were performed using the Statistical Package for
Social Sciences (SPSS) version 18.

31. Chi-squared - (Pearson's chi-square) test is a statistical method used to
evaluate data by comparing categorical data from a sample to expected or
"true" results.
It is used to analyze differences in categorical variables, especially those that
are nominal in nature.
Fisher’s and independent sample Student’s t-tests are statistical tests used to
compare the means of two groups.
Student’s t-test is used when the data is normally distributed and the sample
size is small.
Fisher’s test is used when the data is not normally distributed and the sample
size is small.
Used to determine whether the difference between the means of two groups
is statistically significant or due to chance.

32. Mann-Whitney U test : Used to compare the differences between two
independent samples when the sample distributions are not normally
distributed and the sample sizes are small .
The Wilcoxon signed rank test (also called the Wilcoxon signed rank sum
test) - Non-parametric test to compare data, used if the differences
between pairs of data are non-normally distributed.

33. • Linear regression is a linear approach for modelling the relationship between a
scalar response and one or more explanatory variables.
• Linear mixed-effects models are extensions of linear regression models for data
that are collected and summarized in groups.
• These models describe the relationship between a response variable and
independent variables.
• The Kaplan–Meier estimator is a non-parametric statistic used to estimate
the survival function from lifetime data.
• It has a series of declining horizontal steps which, with a large enough sample
size, approaches the true survival function for that population.

34. RESULTS
• 61 participants were randomized to receive lithium (50.8%, n = 31) or placebo
(49.2%, n = 30).
• Both groups at baseline had similar distributions of sociodemographic, clinical and
biological variables.
• The mean age of the total sample was 72.6 years, patients in the lithium group
were slightly younger compared with those in the placebo group.
• No statistically significant differences were observed between the two groups in
other sociodemographic, clinical and biological variables at baseline
• No differences found in frequency of common comorbidities such as systemic
hypertension, osteoarthritis, dyslipidemia, hypothyroidism, diabetes mellitus and
minor depression (data not shown).

35. Lithium (n=31) Placebo (n=30)
71.2 (5.4) * 74.4 (6.1) *
9.47 (5.29) 9.04 (4.74)
74% 63%
26 / 1 / 4 24 / 3 / 3
Socio-demographic
variables
Age [a]
years, mean (SD)
Education [a]
years, mean (SD)
Gender [b]
Female, %
Ethnicity [c]
Caucasian/Negroid/Asian
(n)
a] Student’s t test (t); [b] Chi-squared test; [c] Fisher’s exact test. * t (59), p=0.03; N.S. otherwise.

36. Cognitive and functional variables
CDR=0.5 [b]
number of cases (%)
22 (71.0%) 21 (70.0%)
CDR Sum of Boxes[a]
mean score (SD); test range 0-18 § 1.5 (1.3) 1.7 (1.4)
ADAS-Cog[a]
mean total score (SD); test range 0-
79 §
10.9 (6.1) 10.7 (5.4)
Delayed recall[a]
mean score (SD); test range 0-10 # 4.6 (2.0) 4.2 (2.4)
Figure recall[a]
mean score (SD); test range 0-4 # 2.3 (1.3) 2.0 (1.2)
Trail Making Test-A[a]
seconds (SD)§ 76.5 (49.1) 85.6 (50.0)
Trail Making Test-B[a]
seconds (SD)§ 176.6 (80.1) 187.0 (79.1)
Sequence of Letters and Numbers[a]
6.5 (2.2) 6.4 (1.9)

37. Cerebrospinal fluid biomarkers (pg/ml)
Amyloid-β1-42
[a]
concentration, mean
(SD)
444.5 (156.4) 429.2 (163.8)
Total Tau [a]
concentration, mean
(SD)
92.8 (56.1) 88.5 (49.9)
Phosphorylated Tau at
threonine181
concentration, mean
(SD)
63.8 (34.4) 59.9 (29.0)

38. RESULTS
• In total, 52 patients (85.2%, n = 52) completed the double-blind phase of the study.
• (87.1%, n = 27 lithium; 83%, n = 25 placebo).
• In the 1st year of the trial, four patients dropped out (two from each group):
• Lithium group : 1 patient had an ischemic stroke, and another patient discontinued
due to personal reasons.
• In the placebo group 1 patient died from sepsis secondary to pneumonia, and 1
patient discontinued the trial for personal reasons.
• In the 2nd year 3 patients in the placebo group were advised to withdraw as they
were diagnosed either with prostate cancer, severe hypertension or ventricular
arrhythmia

39. RESULTS
• 1 patient in the lithium group required hospital admission because of a delirious
state (unrelated to lithium toxicity) and another had clinical symptoms of lithium
intolerance (tremor and nausea), also therefore being excluded from the trial.
• In the extension phase of the study the attrition rate was considerably higher
(nine individuals dropped out from each group), resulting in 55.7% (n = 40) overall
completion rate (58%, n = 18 lithium; 53%, n = 16 placebo).
• All discontinuations in this phase occurred in the third year of follow-up, largely
from participant’s decisions.

40. RESULTS
• Patients in the lithium group had mean serum levels of 0.39 mEq/L (s.d.
0.08) during the double-blind phase, and of 0.41 mEq/L (s.d. = 0.10)
during the extension phase of the trial.

41. NS, non-significant differences.
a. Cognitive and functional changes
calculated as end-point (2 years) minus
baseline scores.
b. Independent sample t-test (d.f.) = 59.
c. Negative values indicate improvement
on test performance.
d. Negative values indicate decline on test
performance.
Patients in the lithium group were cognitively and
functionally stable over 24 months, whereas
patients in the placebo group displayed mild
statistically significant cognitive and functional
decline lithium treatment was beneficial,
although the magnitude of the differences
between the two groups was small.

42. Cognitive and functional changes according to group
allocation (lithium versus placebo) in 2 years of follow-up.
The statistical analysis was made by a linear
mixed model (type III test of fixed effects)
addressing the interactions between treatment
group, time of intervention and group × time.
In (a) and (b) increments in test scores indicate
cognitive or functional worsening; in (c) and (d)
increments indicate improvement. Significant
changes favouring the lithium group were
observed in the CDR-SoB (F = 22.66, P<0.001),
ADAS-Cog (F = 3.68, P = 0.05), delayed recall
(F = 16.48, P<0.001) and figure recall
(F = 6.16, P = 0.02); in the latter test there was
also a significant effect of time on the observed
changes (F = 5.22, P = 0.01).

43. Kaplan–Meier curves of lithium-treated and placebo groups,
indicating the rate of conversion from mild cognitive impairment
(MCI) to dementia,
Five patients in the lithium group (16%) and nine in the
placebo group (30%) converted from MCI to dementia
during follow-up, but the statistical significance of this
difference (P = 0.06) was marginally above the pre-
defined threshold of 0.05.All patients with MCI had
baseline scores on the Clinical Dementia Rating Scale
(CDR) <1.

44. Longitudinal changes in cerebrospinal fluid (CSF) concentrations
of amyloid-β peptide (Aβ1−42) in lithium-treated and control
groups
(a) Indicates a 30% increase
in CSF Aβ1−42 after 3
years in the lithium
group
(F = 3.11, P = 0.003).
(b) Displays changes in CSF
concentrations of
Aβ1−42 in lithium and
placebo groups
subdivided at baseline
according to the
magnitude of
Aβ1−42 burden, i.e., high-
and low-CSF Aβ1−42. NS,
non-significant.
.

45. DISCUSSION
• Main findings:
• Lithium is believed to modulate important intracellular signaling systems
implicated in neurotrophic responses and in mechanisms related to
neurodegeneration.
• The overall results were in line with our preliminary findings from our interim
analysis after 12 months.
• Chronic lithium use may be associated with a lesser deterioration of
cognitive abilities and with a relative preservation of functional capacity.
• The magnitude of the differences between treated and untreated groups was
small, and both groups displayed a mild deterioration over time.

46. DISCUSSION
• The present findings suggest that chronic lithium use was beneficial to patients
with amnestic MCI.
• The analysis of primary outcome variables indicate that participants in the lithium
group performed better than those in the placebo group after 2 years of treatment.
• The differences in global cognitive and functional performance observed at the end
of the trial were already detectable after 12 months.
• Differences in memory and attention scores favouring the lithium group after 2
years were not statistically significant in the interim analysis.
• The number of conversions from MCI to dementia was smaller among lithium
users

47. Methodological considerations
• It was difficult to maintain the double-blind procedures as the study period was
long (4 years).
• The study was subdivided into an initial, 2 year, double-blind phase followed by
an extension phase of 2 additional years.
• Participants (but not raters) were made aware of group allocation and entitled
to decide whether or not to continue in the trial.
• The assessment of primary outcome variables (global cognitive and functional
status) and a subset of secondary outcome variables (neuropsychological test
scores) was restricted to the double-blind phase of the study.

48. Methodological considerations
• The assessment of the conversion status and the analysis of longitudinal
changes in Alzheimer's disease-related CSF biomarkers were based on
variables obtained in both phases of trial.
• Authors believe that these variables may not have been critically
affected by the awareness of group allocation by participants in the
extension phase of the study.

49. Methodological considerations
• Lithium treatment was administered at doses sufficient to yield serum levels of
0.25–0.5 mM/L, considering safety and tolerability.
• Preparatory studies conducted in the laboratory indicated that lithium u within
subtherapeutic range was associated with good availability of lithium in the brain.
•
• The inhibition of GSK3β is the possible mechanism associated with the
neuroprotective effects of lithium in Alzheimer's disease.
• It is likely that the effects of lithium on other molecular targets, or the combination
of its pharmacodynamics properties, may account for the clinical and biological
changes observed in this trial.

50. Methodological considerations
• After 3 years of treatment: patients in the lithium group had a statistically
significant,. 30% increase in the CSF concentrations of Aβ1−42.
• AD related biomarker was found to be increased on longer exposure to lithium as
no such effects were noticed at 12 months.
• Long-term lithium treatment may promote mechanisms related to the clearance of
the Aβ peptide from the brain.
• Those who had higher CSF concentrations of Aβ1−42prior to receiving lithium
treatment were more prone to respond with increment in CSF levels of Aβ1−42 after
3 years.

51. Methodological considerations
• Lithium-induced decrement in CSF concentrations of phosphorylated tau observed
after 12 months of treatment did not withstand late end-point (was not observed
after 36 months).
• The present evidence of changes in CSF concentrations of Aβ1−42 upon long-term
lithium treatment, if confirmed by other experimental and clinical models, may
warrant the chronic use of lithium at low doses as an approach to enhance
amyloid clearance.
• Depending on the biological target and on the stage of the disease process, some
patients may have a good response to lithium treatment, whereas others may not
respond at all.

52. STRENGTHS AND LIMITATIONS
• Relatively small sample size, compared with multicentre trials.
• As a single-centre study, the final sample enrolled to the trial is substantial,
particularly in the light of the long-term follow-up and the frequent monitoring
based on clinical and biological parameters.
• In addition to other recent clinical observations and evidence drawn from
experimental models, large-scale trials involving experts from different groups and
including a wider array of outcome variables for this set of data can be more
justifiable.
• Intervention groups would have been defined according to biological variables at
baseline (for example Alzheimer's disease-related CSF biomarkers and/or amyloid
and tau imaging with positron–emission- tomography)

53. STRENGTHS AND LIMITATIONS
• Patients with Prodromal Alzheimer's disease, clinical Alzheimer's disease, cognitive
impairment unrelated to Alzheimer's disease pathology upon enrolment could have
been excluded, which could have been a better approach to identify the clinical
conditions that might benefit most from lithium.
• Authors are still uncertain about the best therapeutic range of working lithium
concentrations to target biological effects within good safety–tolerability limits.
• Controlled data from the present trial indicates the long-term lithium treatment at
subtherapeutic doses may be safe and well tolerated by older adults.

54. CONCLUSION
• Long-term lithium attenuates cognitive and functional decline in
amnestic MCI, and modifies Alzheimer's disease-related CSF
biomarkers.
• The present data reinforces the disease-modifying properties of
lithium in the MCI–Alzheimer's disease continuum.

55. Declaration of Conflicting Interests:
The authors declared no potential conflicts of interest with respect to the research,
authorship and/or publication of this article.
Funding:
• The present work was supported by the National Council for Scientific Research
(CNPq, Project 554535/2005-0), Alzheimer's Association (NIRG-08-90688 São
Paulo Research Foundation (FAPESP, Project 2009/52825-8).
•
• The Laboratory of Neuroscience (LIM-27) receives financial support from Alzira
Denise Hertzog da Silva Charitable Association (ABADHS).

56. CRITICAL APPRAISAL
STRENGTHS
• Published in freely accessible
journal with good impact factor.
• Registered study.
• Interventional study
• Limitations are mentioned
• Ethical committee clearance
• No conflicts of interest
• Ways to improve the study in
future is discussed.
LIMITATIONS
• Small sample restricted to one
catchment area.
• Selection was based on invitations.
• Consent for extended period of study.
• Too many drop outs at the end of the
study.
• Decision to extend the study made after
the starting of the study,
• Data of the study was split up and
published in different studies.

57. .