participants were recruited from two prospective studies : the prospective research in memory ( prime ) clinics study and the australian imaging , biomarkers and lifestyle ( aibl ) study of aging . only data and biochemical measurements pertaining to baseline visits were included in this analysis .
the prime study recruited 970 participants from 9 sites in australia , including 3 each in victoria and new south wales and 1 each in queensland , western australia , and south australia .
the aibl study of aging recruited 1,112 participants in victoria ( 60% ) and western australia ( 40% ) .
the study cohorts and methods of the prime and aibl studies are described elsewhere ( 26,27 ) . a further 862 participants who resided in the barwon region of southeastern australia between 2001 and 2011
also were recruited through the cognitive , dementia and memory services clinic at the mckellar centre , a rehabilitation and aged - care facility .
patients with ad who were seen at a geriatrician s private practice during the same period also were recruited ( n = 935 ) .
the mini - mental state examination ( mmse ) was used to assess cognitive ability .
subjects were assessed at scheduled visits during the prime and aibl studies or ad hoc during routine patient care .
all participants with serum vitamin b12 measurements taken within 6 months of cognitive assessment were included .
participants without serum measurements taken within 6 months of cognitive assessment ( n = 1,015 ) were excluded . the data from records pertaining to subjects who were recruited from more than one source ( n = 566 ) were merged . of the remaining participants ,
there were 121 with stroke and 566 with diagnoses other than ad , such as frontotemporal dementia , parkinson disease , dementia with lewy bodies , or mixed dementias .
a further 291 participants were excluded because they had incomplete medical histories ; information that was missing included dates of birth , medication use , and comorbid conditions .
a subgroup analysis was performed with participants who had diabetes ( n = 104 ) or impaired glucose tolerance ( n = 22 ) . patients with type 1 diabetes were not specifically excluded , but there were none with serum vitamin b12 measurements taken within 6 months of cognitive assessment .
reviewing committees included the barwon health human research ethics committee ( hrec ) ( victoria ) , austin health hrec ( victoria ) , st .
vincent s hospital governance review unit ( victoria ) , hunter new england hrec ( new south wales ) , northern hospital network hrec ( new south wales ) , northern sydney central coast hrec ( new south wales ) , metro north health service district hrec ( queensland ) , south metropolitan area health service hrec ( western australia ) , and the queen elizabeth hospital ethics of human research committee ( south australia ) .
an ordinal logistic regression model was formed with categories of cognitive performance as the response variable and diabetes as a predictor .
( mmse < 18 ; n = 137 ) ,  mildly impaired  ( mmse 1823 ;
n = 240 ) ,  minimally impaired  ( mmse 2427 ; n = 295 ) , and  not impaired  ( mmse 2830 ; n = 682 ) .
the model was adjusted for age , sex , level of education , and depression because these factors were previously reported to affect mmse testing ( 28,29 ) .
( mmse < 18 ; n = 39 ) ,  mildly impaired  ( mmse 1823 ; n = 40 ) ,  minimally impaired  ( mmse 2427 ; n = 32 ) , and  not impaired  ( mmse 2830 ; n = 15 )
. the effect of metformin on the cognitive performance of patients with diabetes was investigated in this model , which then was adjusted for serum vitamin b12 measurements and use of calcium supplements to investigate any possible interactions .
there was insufficient information on use of other antidiabetic drugs , the duration of metformin use , and markers for socioeconomic status , diet , or exercise to investigate these variables .
participants were recruited from two prospective studies : the prospective research in memory ( prime ) clinics study and the australian imaging , biomarkers and lifestyle ( aibl ) study of aging . only data and biochemical measurements pertaining to baseline visits were included in this analysis .
the prime study recruited 970 participants from 9 sites in australia , including 3 each in victoria and new south wales and 1 each in queensland , western australia , and south australia .
the aibl study of aging recruited 1,112 participants in victoria ( 60% ) and western australia ( 40% ) .
the study cohorts and methods of the prime and aibl studies are described elsewhere ( 26,27 ) . a further 862 participants who resided in the barwon region of southeastern australia between 2001 and 2011
also were recruited through the cognitive , dementia and memory services clinic at the mckellar centre , a rehabilitation and aged - care facility .
patients with ad who were seen at a geriatrician s private practice during the same period also were recruited ( n = 935 ) .
aibl participants volunteered after an advertisement on television in late 2006 . the mini - mental state examination ( mmse )
subjects were assessed at scheduled visits during the prime and aibl studies or ad hoc during routine patient care .
all participants with serum vitamin b12 measurements taken within 6 months of cognitive assessment were included .
participants without serum measurements taken within 6 months of cognitive assessment ( n = 1,015 ) were excluded . the data from records pertaining to subjects who were recruited from more than one source ( n = 566 ) were merged . of the remaining participants ,
there were 121 with stroke and 566 with diagnoses other than ad , such as frontotemporal dementia , parkinson disease , dementia with lewy bodies , or mixed dementias .
a further 291 participants were excluded because they had incomplete medical histories ; information that was missing included dates of birth , medication use , and comorbid conditions .
a subgroup analysis was performed with participants who had diabetes ( n = 104 ) or impaired glucose tolerance ( n = 22 ) . patients with type 1 diabetes were not specifically excluded , but there were none with serum vitamin b12 measurements taken within 6 months of cognitive assessment .
institutional review was performed at each study host site . reviewing committees included the barwon health human research ethics committee ( hrec ) ( victoria ) , austin health hrec ( victoria ) , st .
vincent s hospital governance review unit ( victoria ) , hunter new england hrec ( new south wales ) , northern hospital network hrec ( new south wales ) , northern sydney central coast hrec ( new south wales ) , metro north health service district hrec ( queensland ) , south metropolitan area health service hrec ( western australia ) , and the queen elizabeth hospital ethics of human research committee ( south australia ) .
an ordinal logistic regression model was formed with categories of cognitive performance as the response variable and diabetes as a predictor .
( mmse < 18 ; n = 137 ) ,  mildly impaired  ( mmse 1823 ;
n = 240 ) ,  minimally impaired  ( mmse 2427 ; n = 295 ) , and  not impaired  ( mmse 2830 ; n = 682 ) . the model was adjusted for age , sex , level of education , and depression because these factors were previously reported to affect mmse testing ( 28,29 ) .
categories were  most impaired  ( mmse < 18 ; n = 39 ) ,  mildly impaired  ( mmse 1823 ; n = 40 ) ,  minimally impaired  ( mmse 2427 ; n = 32 ) , and  not impaired  ( mmse 2830 ; n = 15 )
. the effect of metformin on the cognitive performance of patients with diabetes was investigated in this model , which then was adjusted for serum vitamin b12 measurements and use of calcium supplements to investigate any possible interactions .
there was insufficient information on use of other antidiabetic drugs , the duration of metformin use , and markers for socioeconomic status , diet , or exercise to investigate these variables .
participants were 5199 years old ( mean age  sd 73.8  8.3 years ) ; females outnumbered males ( 59.5 vs. 40.5% ) .
just more than half of the participants ( 50.4% ) scored 2830 on the mmse and so were considered not impaired ; 21.8% were minimally impaired ( mmse 2427 ) , 17.7% were mildly impaired ( mmse 1823 ) , and 10.1% scored less than 18 on the mmse ( most impaired ) .
participants with diabetes were marginally older than participants without diabetes ( 75.5 vs. 73.6 years ; p = 0.013 ) .
the number of males was proportionally larger among participants with diabetes ( 46.8 vs. 39.9% ) , but this difference was not statistically significant .
prevalence of depression was similar between participants with diabetes and those without diabetes ( 31.7 vs. 27.3% ; p = 1.000 ) .
the proportion of participants with a tertiary level of education was higher among participants without diabetes than participants with diabetes ( 39.3 vs. 22.2% ; p < 0.001 ) .
the number of participants who scored below 28 on the mmse was proportionally higher among participants with diabetes than those without diabetes ( 69.0 vs. 47.6% ; p = 1.000 ) . after adjusting for age , sex , education , and depression ,
participants with type 2 diabetes had worse cognitive performance than participants without diabetes ( adjusted or 1.51 [ 95% ci 1.032.21 ] ) .
cognitive performance was better in younger participants , those without depression , and those with a higher level of education .
the adjusted ors for each predictor are shown in table 1 . cognitive performance in 1,354 participants who had ad or mci or were cognitively intact among participants with diabetes , cognitive performance was worse in patients who were taking metformin ( adjusted or 2.23 [ 95% ci 1.054.75 ] ) .
mmse scores were lower in participants with diabetes who used metformin ( mean score  sd 22.8  5.5 ) than in those who did not use metformin ( 24.7  4.4 ) .
participants with diabetes who had vitamin b12 levels < 250 mol / l also had worse cognitive performance ( 2.29 [ 1.124.66 ] ) .
< 250 mol / l ( 22.9  4.7 ) than those with higher levels ( 25.0  4.7 ) . each 1-year increase in age was associated with an 8% increased risk of impaired cognitive performance ( or 1.08 [ 95% ci 1.031.13 ] ) .
a secondary or tertiary level of education was the strongest predictor of better cognitive performance in patients with diabetes .
the adjusted ors for each variable are shown in table 2 . cognitive performance in 126 patients with either type 2 diabetes or impaired glucose tolerance
in our series , patients with diabetes who were taking metformin had worse cognitive performance than participants who were not taking metformin .
our observations agree with those previously reported by imfeld and colleagues ( 22 ) , in particular that patients who are taking metformin may be at an increased risk for cognitive impairment .
this association was weakened after adjusting for serum vitamin b12 levels ; thus any effect metformin has on cognitive performance may be at least partially mediated by altering serum vitamin b12 levels .
alternatively , patients who are prescribed metformin may have worse glycemic control or diabetes - related complications than patients with diabetes who are not prescribed metformin , so there remains the potential for confounding , despite restricting the analysis to only patients with diabetes .
however , because metformin is a first - line pharmacotherapy for the treatment of type 2 diabetes , this would seem unlikely ( 30 )
. there was insufficient information regarding the duration of metformin use , the severity of diabetes ( e.g. , hba1c levels ) , duration of diabetes , or use of other antidiabetic drugs to enable us to investigate these effects in our study , particularly because these findings were based on a small sample .
we recommend a larger study to examine the effect of dose and duration of metformin use , and the effects of other antidiabetic agents using a battery of cognitive assessments and following participants over a number of years .
calcium supplements have previously been reported to reverse vitamin b12 deficiency induced by metformin . in this study , patients with diabetes who used calcium supplements were less likely to be cognitively impaired .
however , calcium supplements have been reported to be associated with an increased risk for myocardial infarction in postmenopausal women and in patients with chronic kidney disease ( 3134 ) .
in contrast , a recent meta - analysis indicates that supplementation with both vitamin d and calcium is associated with a reduction in mortality compared with vitamin d supplementation alone ( 35 ) .
because this population already has increased cardiovascular risk ( 36 ) , the safety of calcium supplementation in patients with diabetes treated with metformin would need to be established before such interventions could be recommended .
patients with diabetes are at an increased risk for ad ( 10 ) . in diabetes ,
amylin aggregation destroys the -cells of the pancreas ( 37 ) . by the same mechanism
metformin is a widely prescribed first - line monotherapy for type 2 diabetes but is associated with vitamin b12 deficiency and peripheral neuropathy
. a case - control study of more than 14,000 patients reported that long - term metformin use was associated with an increased risk for ad in those 65 years old ( 22 ) .
metformin at pharmacological doses was reported to increase the expression of -secretase in cell culture ; this may be a possible disease mechanism ( 21 ) .
alternatively , metformin also impairs absorption of vitamin b12 via a drug interaction that occurs at the distal ileum .
low serum vitamin b12 levels are associated with ad and other neurodegenerative diseases ( 38 ) .
the mmse may be inadequate for detecting differences between higher functioning adults ( 39 ) .
the mmse is sensitive to age , depression , and level of education , so all models were adjusted for these factors .
most of our subjects were assessed during routine clinical care by clinicians who use the mmse as part of their standard assessment . in australia ,
documentation of cognitive impairment using an mmse score and improvement while receiving therapy is required to obtain subsidized antidementia therapies ( 40 ) .
more comprehensive assessment tools may be preferable for use in future investigations of cognitive impairment in at - risk populations such as those we have studied .
increased monitoring of cognitive ability in patients with diabetes who use metformin is warranted , particularly among older adults ( aged older than 50 years ) .
vitamin b12 supplements are inexpensive and may improve the cognitive outcomes of patients with diabetes .
adequately powered , prospective , controlled trials are warranted to investigate further the association between diabetes , cognitive decline , and the effect of metformin therapy , as well as the possible amelioration using vitamin b12 and/or calcium supplementation .
 objectiveto investigate the associations of metformin , serum vitamin b12 , calcium supplements , and cognitive impairment in patients with diabetes.research design and methodsparticipants were recruited from the primary research in memory ( prime ) clinics study , the australian imaging , biomarkers and lifestyle ( aibl ) study of aging , and the barwon region of southeastern australia . 
 patients with alzheimer disease ( ad ) ( n = 480 ) or mild cognitive impairment ( n = 187 ) and those who were cognitively intact ( n = 687 ) were included ; patients with stroke or with neurodegenerative diseases other than ad were excluded . 
 subgroup analyses were performed for participants who had either type 2 diabetes ( n = 104 ) or impaired glucose tolerance ( n = 22).resultsparticipants with diabetes ( n = 126 ) had worse cognitive performance than participants who did not have diabetes ( n = 1,228 ; adjusted odds ratio 1.51 [ 95% ci 1.032.21 ] ) . among participants with diabetes , worse cognitive performance 
 was associated with metformin use ( 2.23 [ 1.054.75 ] ) . after adjusting for age , sex , level of education , history of depression , serum vitamin b12 , and metformin use , participants with diabetes who were taking calcium supplements had better cognitive performance ( 0.41 
 [ 0.190.92]).conclusionsmetformin use was associated with impaired cognitive performance . 
 vitamin b12 and calcium supplements may alleviate metformin - induced vitamin b12 deficiency and were associated with better cognitive outcomes . 
 prospective trials are warranted to assess the beneficial effects of vitamin b12 and calcium use on cognition in older people with diabetes who are taking metformin . 
RESEARCH DESIGN AND METHODS
Participants and settings
Study design
Ethical approval
Statistical analyses
RESULTS
CONCLUSIONS