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Trial registered on ANZCTR

Registration number

ACTRN12617000403336

Ethics application status

Approved

Date submitted

30/09/2015

Date registered

17/03/2017

Date last updated

20/01/2020

Date data sharing statement initially provided

23/01/2019

Type of registration

Prospectively registered

Titles & IDs

Public title

Effects of A1 versus A2 milk on cardiometabolic risk in overweight and obese Australians

Scientific title

Overweight and obese Australians and the effects of A1 beta-casein versus A2 beta-casein on human cardiometabolic risk factors.

Secondary ID [1]

None

Universal Trial Number (UTN)

U1111-1194-2629

Trial acronym

Linked study record

Health condition

Health condition(s) or problem(s) studied:

Overweight and obesity

Cardiometabolic risk factors related to A1 beta-casein

Gastrointestinal symptoms related to A1 beta-casein

Condition category

Condition code

Cardiovascular

Diseases of the vasculature and circulation including the lymphatic system

Diet and Nutrition

Obesity

Inflammatory and Immune System

Other inflammatory or immune system disorders

Intervention/exposure

Study type

Interventional

Description of intervention(s) / exposure

All participants will undergo a washout period of 2 weeks during which they will replace all dairy with rice milk. Other dairy containing product consumption will be restricted.

Intervention 1: 250ml three times a day (750 ml/day) A1A2 beta-casein Milk for 12 weeks
Intervention 2: 250ml three times a day (750 ml/day) A2A2 beta-casein Milk for 12 weeks

Adherence will be monitored via a milk intake calendar

Intervention code [1]

Treatment: Other

Comparator / control treatment

Control: 250ml three times a day (750 ml/day) Rice Milk for 12 weeks

Adherence will be monitored via a milk intake calendar

Control group

Active

Outcomes

Primary outcome [1]

Fasting serum cholesterol

Timepoint [1]

Pre-washout, baseline and 12 weeks.

Primary outcome [2]

Fasting serum triglyceride

Timepoint [2]

Pre-washout, baseline and 12 weeks.

Primary outcome [3]

Antibodies to oxidised LDL measured by ELISA

Timepoint [3]

Pre-washout, baseline and 12 weeks.

Secondary outcome [1]

Fasting serum inflammatory markers (Glutathione, MPO, IL-4, IL-6, IL-8, CRP, TNF-alpha, IFN-gamma and TGF-beta1

Timepoint [1]

Pre-washout, baseline and 12 weeks.

Secondary outcome [2]

Fasting serum insulin

Timepoint [2]

Pre-washout, baseline and 12 weeks.

Secondary outcome [3]

Blood pressure measured by automated, calibrated sphygmomanometer

Timepoint [3]

Pre-washout, baseline and 12 weeks

Secondary outcome [4]

Fasting serum leptin

Timepoint [4]

Pre-washout, baseline and 12 weeks.

Secondary outcome [5]

Body composition measured by dual-energy X-ray absorptiometry (DXA)

Timepoint [5]

Baseline and 12 weeks

Secondary outcome [6]

Gut permeability measured by dual sugar test (Lactulose/Mannitol)

Timepoint [6]

Baseline and 12 weeks

Secondary outcome [7]

Gut inflammation measured by assesing faecal calprotectin with a single step enzyme linked immunosorbant assay (ELISA)

Timepoint [7]

Baseline and 12 weeks

Secondary outcome [8]

A1A2, A2A2 and Rice milk tolerance measured by GI symptom report diary

Timepoint [8]

Baseline, 6 weeks and 12 weeks

Secondary outcome [9]

Body weight will be recorded in light clothing without shoes with a Body Composition Monitor (Omron Healthcare, Kyoto, Japan)

Timepoint [9]

Pre-washout, baseline and 12 weeks

Secondary outcome [10]

Fasting serum glucose

Timepoint [10]

Pre-washout, baseline and 12 weeks.

Secondary outcome [11]

Fasting whole blood Hba1C

Timepoint [11]

Pre-washout, baseline and 12 weeks.

Secondary outcome [12]

Arterial stiffness assessed by pulse contour analysis using the Pulse Trace PCA 2 (Carefusion, NSW, Australia)

Timepoint [12]

Pre-washout, baseline and 12 weeks

Secondary outcome [13]

Satiety will be assessed by 100 mm Visual Analogue Scale (VAS) questionnaires

Timepoint [13]

Baseline and 12 weeks

Secondary outcome [14]

Waist circumference will be measured in the standing position at the narrowest area between the lateral lower rib and the iliac crest with a tape measure.

Timepoint [14]

Pre-washout, baseline and 12 weeks

Secondary outcome [15]

Hip circumference will be taken at the largest circumference of the lower abdomen with a tape measure.

Timepoint [15]

Pre-washout, baseline and 12 weeks

Secondary outcome [16]

Plasma BCM-7 will be assessed by ELISA and cross-checked with HPLC MS

Timepoint [16]

Pre-washout, baseline and 12 weeks.

Secondary outcome [17]

Fasting serum Non-Esterified Fatty Acids (NEFA)

Timepoint [17]

Pre-washout, baseline and 12 weeks.

Secondary outcome [18]

Fasting serum Dipeptidyl Peptidase-IV (DPPIV) activity

Timepoint [18]

Pre-washout, baseline and 12 weeks.

Secondary outcome [19]

Psychological health (quality of life, psychological well-being, self-rated health, and anxiety and depression) will be measured using previously-validated self-report psychometric inventories.

Timepoint [19]

Baseline and 12 weeks

Secondary outcome [20]

Executive Functioning will be assessed by the Subtle Cognitive Impairment Test (SCIT)

Timepoint [20]

Baseline and 12 weeks

Secondary outcome [21]

Short Chain Fatty Acids (SCFA) in faecal samples will be analysed

Timepoint [21]

Baseline and 12 weeks

Secondary outcome [22]

Gut microbiome will be analysed from faecal samples

Timepoint [22]

Baseline and 12 weeks

Eligibility

Key inclusion criteria

overweight or obese (BMI: 25-40 kg/m2)
willing to consume 750 ml of milk/day

Minimum age

18 Years

Maximum age

70 Years

Sex

Both males and females

Can healthy volunteers participate?

Yes

Key exclusion criteria

Milk allergy, pregnancy and lactation, diabetes, current habitual opioid consumption and smokers

Study design

Purpose of the study

Treatment

Allocation to intervention

Randomised controlled trial

Procedure for enrolling a subject and allocating the treatment (allocation concealment procedures)

Eligible participants will be assigned an ID as they are recruited. Allocation will involve contacting the holder of the allocation schedule who will be off-site.

Methods used to generate the sequence in which subjects will be randomised (sequence generation)

The randomisation scheme will be created using www.randomization.com with 3 treatments.

Masking / blinding

Blinded (masking used)

Who is / are masked / blinded?

The people receiving the treatment/s
The people administering the treatment/s
The people assessing the outcomes
The people analysing the results/data

Intervention assignment

Parallel

Other design features

Phase

Not Applicable

Type of endpoint/s

Efficacy

Statistical methods / analysis

A sample size of 32 participants/group is predicted to provide sufficient power (80%) to detect a large effect size (7% difference) between groups at the p < 0.05 level. Recruiting 46 participants/group will accommodate for 30% dropout rate. A total of 140 participants will be recruited for the 3 groups. Calculations are based on standard deviation of 10% within a group observed in our previous studies examining changes in cardiometabolic risk factors in overweight and obese participants. Statistical analysis will be undertaken using SPSS 22 for Windows (SPSS Inc., Chicago, IL). Data will be expressed as mean (+/-SEM) and assessed for normality to ensure that the assumptions of the analysis are met. The data will be analysed using General Linear Models with baseline value covariates. If significant between groups effects are present, post hoc comparisons between the treatment groups will be made using the Least Significant Difference (LSD) method. Statistical significance will be considered at p<0.05.

Recruitment

Recruitment status

Recruiting

Date of first participant enrolment

Anticipated

4/02/2019

Actual

25/02/2019

Date of last participant enrolment

Anticipated

7/09/2020

Actual

Date of last data collection

Anticipated

14/12/2020

Actual

Sample size

Target

140

Accrual to date

127

Final

Recruitment in Australia

Recruitment state(s)

Funding & Sponsors

Funding source category [1]

Commercial sector/Industry

Name [1]

The a2 milk Company

Address [1]

PO Box 1564 North Sydney
NSW 2059

Country [1]

Australia

Primary sponsor type

University

Name

Curtin University

Address

GPO Box U1987 Perth
WA 6845

Country

Australia

Secondary sponsor category [1]

None

Name [1]

Address [1]

Country [1]

Ethics approval

Ethics application status

Approved

Ethics committee name [1]

Curtin Human Research Ethics Committee

Ethics committee address [1]

GPO Box U1987, Perth
Western Australia, 6845

Ethics committee country [1]

Australia

Date submitted for ethics approval [1]

24/03/2017

Approval date [1]

26/07/2017

Ethics approval number [1]

HRE2017-0490

Summary

Brief summary

Obesity significantly increases the risk of developing metabolic syndrome (MS), which is associated with increased cardiovascular morbidity/mortality. Presently, there is fierce debate surrounding the effects of the A1 beta-casein protein variant (A1) in bovine milk compared to the progenitor A2 variant (A2) with respect to their relative impact on MS risk factors. Most milk and milk products available commercially in Australia contain the A1 protein from dairy cows which carry the A1 gene. The A1 variant in Australian dairy cattle is the result of a mutation from the A2 gene in European herds ~5,000 years ago. Alternatively, milk produced by cows specially selected and then bred to produce only the original A2 beta-casein type is also available in Australia, but it is a specialty milk brand. Ample evidence now suggests that A1 milk beta-casein can affect features of MS and so has important public health implications. Given the Australian Dietary Guidelines recommend 2.5-4 dairy serves/day, the effect of milk variety on MS warrants investigation. A1 and A2 protein have differences in bioactivity on digestion, due to the release of the 7-amino acid opioid peptide beta-casomorphin-7 (BCM-7), from the digestion of A1 but not A2. In vitro and animal studies have demonstrated that BCM-7 oxidises low density lipoprotein (LDL)-cholesterol and increases levels of the inflammatory marker myeloperoxidase, both of which are implicated in the development of heart disease. A1 beta-casein and BCM-7 also stimulate inflammatory markers and so may contribute to the overall inflammatory milieu in MS. We have also shown in our recent randomised cross-over study that A1 milk stimulates differences in gastrointestinal inflammatory responses compared to A2 milk. 

The aim of the current proposed project is to investigate the difference between A1 and A2 beta-casein protein containing milk on cardiovascular and metabolic risk factors in overweight/obese individuals. Simple and affordable nutrition interventions that can positively affect modifiable risk factors for heart disease and T2D in Australian adults are needed.  If the predicted differences between A1 and A2 beta-casein containing milk on cardiometabolic risk outcomes are found, it will have important ramification for public health and the prevention of chronic illness like cardiovascular disease. Dairy farmers would be advised to choose A2 semen for their dairy breeding programs to change to the A2 milk type for Australian consumers.

We hypothesise that A1, but not A2 beta-casein containing milk, will correspond to shifts in cardiometabolic risk factors mediated via BCM-7 in blood. In contrast, we propose that A2 beta-casein containing milk will have a neutral, rather than beneficial, effect on metabolic syndrome factors. Thus, the hypothesis implies that A2 will do no harm, while the mutated A1 variant may increase the risk of chronic disease at a population health level.

Trial website

Trial related presentations / publications

Public notes

Contacts

Principal investigator

Name

A/Prof Sebely Pal

Address

Curtin University
School of Public Health
Kent Street, Bentley, Perth
Western Australia 6102

Country

Australia

Phone

+61 8 9266 4755

Fax

[email protected]

Contact person for public queries

Name

Suleen Ho

Address

Curtin University
School of Public Health
Kent Street, Bentley, Perth
Western Australia 6102

Country

Australia

Phone

+61 422238198

Fax

[email protected]

Contact person for scientific queries

Name

Suleen Ho

Address

Curtin University
School of Public Health
Kent Street, Bentley, Perth
Western Australia 6102

Country

Australia

Phone

+61 422238198

Fax

[email protected]

Data sharing statement

Will individual participant data (IPD) for this trial be available (including data dictionaries)?

No/undecided IPD sharing reason/comment

IPD availability was not discussed during planning for this study.

What supporting documents are/will be available?

No Supporting Document Provided

Results publications and other study-related documents

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No documents have been uploaded by study researchers.

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No additional documents have been identified.

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