Original Paper Needs Analysis for a Parenting App to Prevent Unintentional Injury in Newborn Babies and Toddlers: Focus Group and Survey Study Among Chinese Caregivers

Peishan Ning
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

MD;Deyue Gao
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Peixia Cheng
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Ba ; David
C Schwebel
Department of Psychology
University of Alabama at Birmingham
BirminghamALUnited States

PhD; Xiang
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Wei
Ba ; Liheng Tan
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Ba ; Wangxin
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Xiao
Ba ; Jieyi He
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Ba ; Yanhong Fu
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Ba ; Bo Chen
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Ba ;
Yang Yang
Department of Biostatistics
College of Public Health and Health Professions
University of Florida
GainesvilleFLUnited States

Jing Deng
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

PhD;Yue Wu
Department of Occupational and Environmental Health
Xiangya School of Public Health
Central South University
ChangshaChina

PhD; Renhe
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

Yu
Shukun Ba;
Li
Information and Network Center
Department of Epidemiology and Health Statistics Xiangya School of Public Health Central
Central South University
ChangshaChina

South University
110 Xiangya Road Changsha410078China

Ma ;
Guoqing Hu
Department of Epidemiology and Health Statistics
Xiangya School of Public Health
Central South University
ChangshaChina

PhDGuoqing Hu
Original Paper Needs Analysis for a Parenting App to Prevent Unintentional Injury in Newborn Babies and Toddlers: Focus Group and Survey Study Among Chinese Caregivers
10.2196/11957* these authors contributed equally Corresponding Author:
Background: With the growing popularity of mobile health technology, app-based interventions delivered by smartphone have become an increasingly important strategy toward injury prevention.Objective: This study aimed to develop a framework supporting the design of an app-based intervention to prevent unintentional injury, targeted for caregivers of Chinese children aged 0 to 6 years.Methods:A theory-based mixed-method study, including focus groups and Web-based quantitative survey, was performed. Adult caregivers who care for children aged 0 to 6 years and own a smartphone were recruited into 2 sequential stages of research. First, focus groups were conducted among the caregivers at community health care centers and preschools from December 2015 to March 2016. Focus groups (8-10 participants per group) explored awareness, experiences, and opinions of caregivers toward using an app to prevent unintentional injury among children. Second, based on the focus groups findings, a Web-based quantitative survey was designed and distributed to caregivers in November 2016; it collected information on specific needs for the app-based intervention. Thematic analysis and quantitative descriptive analyses were performed.Results:In total, 12 focus groups were completed, involving 108 caregivers. Most participants expressed a strong desire to learn knowledge and skills about unintentional child injury prevention and held positive attitudes toward app-based interventions. Participants expressed multiple preferences concerning the app-based intervention, including their contents, functions, interactive styles, installation and registration logistics, and privacy protection and information security. Following the focus groups, 1505 caregivers completed a WeChat-based quantitative survey, which generated roughly similar results to those of focus groups and added numerical metrics concerning participants' preferences on what to learn, when to learn it, and how to learn it. A detailed framework was established involving 5 components: (1) content design, (2) functional design, (3) interactive style, (4) installation and registration logistics, and (5) privacy protection and information security, and 15 specific requirements.Conclusions:We developed a framework that can be used as a guide to design app-based interventions for parents and caregivers, specifically for unintentional injury prevention of children aged 0 to 6 years.

Introduction

Background Unintentional injury is a serious public health problem among children aged 0 to 6 years in China. In 2017, over 20,000 Chinese children younger than 5 years died of unintentional injury, and nearly 2.4 million children required emergency or outpatient care because of unintentional injury [1].

Parenting interventions can substantially reduce child unintentional injury risk by improving caregivers' safety knowledge and perceptions about the risk for injury to their children and through adoption of safety equipment and practices [2][3][4][5]. For a variety of reasons, implementation of such interventions has been inadequate in low-and middle-income countries such as China [6,7]. In fact, 2 domestic studies from China indicate that less than 20% of caregivers had ever received education concerning unintentional injury prevention for children aged 0 to 6 years or attended a professional prevention course on the topic [8,9].

Encouragingly, the mobile health (mHealth) movement to deliver health interventions through technology has potential to overcome this barrier to parent education for child injury prevention. The number of smartphone users in China has grown quickly over the past decade, with recent estimates suggesting that about 403 million adults aged 20 to 39 years (95% of the Chinese population in that age group) accessed the internet through mobile phones in December 2017 [10,11]. Compared with traditional health education methods delivered in person by professionals, mHealth interventions are low-cost and easy-to-implement, allow interactions between users and providers, and can be accessed anonymously with flexibility at any time and any place [12][13][14].

Recently, app-based interventions have been developed to prevent or reduce unintentional injury risk in specific injury domains, including sports injuries [15], fire injuries [16], road traffic injuries [16,17], falls [18], and burns [19]. Some app-based interventions have been critiqued, however, for failing to meet basic principles for effective prevention programs such as being based in theory or being tailored to risks in the target population [20].

Multiple health behavior change theories can be used in the development of an app to reduce injuries. The Theory of Planned Behavior (TPB) offers a social psychological theory that fits nicely, as it is used to interpret and predict why individuals perform specific behaviors [21]. A recent systematic review concluded that TPB is an effective framework to identify and understand child and adolescent nutrition-related behaviors. It found, for example, that attitude was strongly related to dietary behavioral intention (mean r=0.52) and intention was the most common predictor of behavior performance (mean r=0. 38) [22]. Interventions grounded in TPB must attend to topics such as the individual's attitudes toward using an app-based intervention and changing their behavior as a result of that app, the individual's subjective norms about behavior that should be conducted, and the individual's perception of how much they can control their own behavior surrounding the health area of interest.

Computer science theories are also critical to the development of an app-based intervention. The Framework for the Rational Analysis of Mobile Education (FRAME) model offers a theoretical grounding in computer science to guide the development of mHealth learning programs. The FRAME model considers 3 aspects of mobile learning: usability of the device and app, capacities of the learners, and social interaction between users. Regarded as a comprehensive model to develop and implement mobile learning, attention to the components of the FRAME model allows app development to proceed with attention to all relevant components of user learning [23].

Objectives

We, therefore, conducted a mixed-method study to establish a theory-and need-based framework that would support and lead to the design of an app-based intervention for child injury prevention to be used by Chinese caregivers of children aged 0 to 6 years. All assessment protocols were grounded in TPB and the FRAME model and were designed to gather information that would be valuable for the design of app-based health interventions focused on improving parenting among Chinese caregivers to reduce child injury risk.

Methods

Study Design

Grounded in TPB and the FRAME model [21,23], we designed a 2-step sequential mixed-method study. The TPB suggests that an effective app parenting intervention must offer strategies to alter the attitudes, subjective norms, and perceived control to engage in actions that will improve the safety of children. We targeted several aspects of these strategies in our inquiries to caregivers concerning their preferences in the design of an app-based intervention. The FRAME model addresses users' preferences for app usability, the capacity of the app users, and the social interactions between app users; we addressed these aspects of app functioning also in the inquiries to caregivers.

To gather qualitative data first, focus groups were conducted to explore the experiences and preferences of adult caregivers on using an app-based intervention to prevent unintentional injury among children younger than 7 years. Next, focus group findings were used to guide development and implementation of a Web-based survey to quantify key needs for the app-based interventions (eg, frequency and length of content, variety and types of forms of learning, and duration of app-based learning).

Focus Group

Participants

Participants were recruited for focus groups using purposive sampling. Eligible participants included primary caregivers of 1 or more children younger than 7 years and who used a smartphone regularly. No exclusion criteria applied beyond the age of children that caregivers looked after.

Caregivers of children aged 0 to 3 years were recruited primarily from community health centers. Caregivers of children aged 4 to 6 years were recruited from preschools. To maximize sampling variation, we sampled from a range of preschool types (including both public and private) and geographic locations (including varied socioeconomic status and living areas). Focus group members represented both sexes and a range of household incomes, ages, and levels of education.

Setting

Focus groups were completed between December 2015 and March 2016 in Changsha, China. A semistructured discussion guide was developed and refined through pilot testing first among the research group and then with an independent focus group of caregivers of children aged 0 to 3 years (Multimedia Appendix 1). Informed consent was obtained from all participants before each focus group began.

Broadly, discussions were organized to concentrate on perceptions and opinions about unintentional child injury prevention and app-based injury interventions as well as on issues of designing app-based intervention (eg, content, function, interfaces, and data security). To support discussion about intervention interfaces, the facilitators prepared a series of slides that illustrated different styles of interface designs.

When each focus group discussion concluded, participants completed short paper-based questionnaires that collected information on sociodemographic characteristics for both caregivers and their children and unintentional injury history for their children.

Following scholarly recommendations to organize focus groups [24,25], each focus group consisted of 8 to 10 participants and lasted for 60 to 90 min. A total of 3 trained facilitators led the discussion of all groups; 1 served as moderator and the other 2 took extensive discussion notes and supported the moderator as needed to organize the discussion. All focus group discussions were audio-taped, transcribed, and then reviewed before the next focus group. This allowed facilitators to include new concepts or opinions and exclude old but less relevant subtopics iteratively. Focus groups were concluded when no new concepts and opinions emerged.

Data Analysis

Focus group audiotapes were transcribed verbatim and checked for accuracy. All participant names were removed to permit anonymized analysis. The transcripts were then analyzed using thematic analysis strategies [26]. First, 2 researchers spent several days familiarizing themselves with all transcripts and generating initial codes independently. Second, the 2 coders combined initial codes and organized them into themes based on the discussion guideline, concordance between codes, and the underlying theory. Third, the 2 researchers' proposed themes were combined and re-evaluated to ensure no major themes from the discussions were omitted. Finally, group discussions of the research group (including 2 coders) were held to achieve consensus on any themes where the 2 researchers held differing views. Transcript analysis and coding were completed using MAXQDA 12.0 (qualitative data analysis software by VERBI GmbH).

Web-Based Survey

Participants

Study participants for the Web-based survey component of the study were recruited through snowball sampling. Inclusion criterion matched those of the focus groups: caregivers of children younger than 7 years who owned a smartphone. No exclusion criteria applied beyond the age of children that caregivers looked after.

To maximize reach to the target population, we used WeChat, the most popular social media communication platform in China, with an average of 889 million active users each month [27], to conduct the survey. Specifically, the research team sent a study recruitment message through an official WeChat account known as La Ma Xue Yuan (School for Young Mothers), which includes 9290 users throughout China (mostly caregivers of young children). This message included an invitation letter and the questionnaire. Participants consented to participate online and then completed and submitted the questionnaire. Participants were also encouraged to share the message with their social network, many of whom met the inclusion criterion, allowing us to "snowball" to a larger sample size. The Web-based survey remained open until the number of completed questionnaires plateaued.

Setting

The Web-based survey was completed in November 2016. It was developed based on the results of the focus groups, grounded in TPB and especially the FRAME model, and finalized through pilot testing with 20 caregivers. The survey included items assessing demographic characteristics, history of child unintentional injury, prior learning experiences about child unintentional injury prevention, and preferred learning contents (Multimedia Appendix 2). To avoid repeated questionnaire completion, we restricted the survey to a single response from each smartphone. To encourage participation in the Web-based survey portion of the study, raffle prizes with cash incentives were distributed at a probability of 1/3 after participants completed the survey.

Data Analysis

The collected questionnaires were screened to exclude those completed by caregivers who did not meet the inclusion criteria. Proportions were then calculated to describe preferred contents of the app-based intervention, forms of app-based learning, preferred learning time, and frequency and duration for using app-based learning. Chi-square test examined group differences across groups. Data analysis was performed using the Statistical Product and Service Solutions (SPSS 18.0; IBM Corporation). P<.05 was considered to be statistically significant.

Ethical Considerations

The protocol was approved by the ethics committee of the Institute of Clinical Pharmacology of Central South University. This study was conducted, analyzed, and reported according to the consolidated criteria for reporting qualitative research (COREQ) [28] and the checklist for reporting results of internet E-survey (CHERRIES [29]; Multimedia Appendix 3). All participants were informed about the study and provided informed consent before participating in the research. All data were analyzed anonymously.

Results

Focus Group Results

Participants

In total, 12 focus groups were organized; together they included 108 caregivers (90 parents, 6 grandparents, and 12 preschool teachers; Table 1). The largest portions of participants were female (96/108, 88.9%) and aged 20 to 39 years (94/108, 87.0%). Among the parent and grandparent participants, the caregivers supervised 96 children who ranged in age from 0 to 6 years, with the largest portions of children aged 4 to 6 years (35/96, 37%) and 0 to 1 year (34/96, 35%). The ratio of boys to girls supervised was 1.04 (Table 1).

Attitude and Behavioral Intention Toward Child Unintentional Injury Prevention

Most caregivers reported a strong desire to learn unintentional injury prevention strategies for children aged 0 to 6 years. Many participants mentioned that they felt unintentional injuries were largely preventable and that they had a strong intention to learn knowledge and skills that would help them prevent unintentional injuries to children under their care. A typical opinion was as follows:

We definitely want to learn it[knowledge about unintentional injury prevention for children ages 0-6 years]. From my point of view, it would be very useful and valuable for us [to learn that information].

[Participant #DWT-A-01] Beyond wanting to learn knowledge about child injury prevention, the majority of focus group participants expressed the opinion that they could use a smartphone app to learn that knowledge. The participants believed that an app developed by a professional team to address child injury prevention would be credible and could be created to include sufficient content for their learning. They also expressed the opinion that an app may be a more convenient educational method than alternatives, such as written brochures or social networks like WeChat, to search for injury prevention content when they wanted it: A few participants expressed some concerns about the utility of a child injury prevention app. They worried about the possible difficulty in the installation and registration process and the possibility that it would use too much memory space on their smartphones, but they said they would accept the app if these problems were addressed.
Compared

Content of App-Based Intervention

Participants mentioned with some frequency 10 major causes of child unintentional injury that would be worth including in the app: exposure to animate mechanical forces (including animal bites and being trampled or bumped by other people); exposure to inanimate mechanical forces (including being pinched between 2 surfaces, such as in the doors of elevators, or cut or punctured by sharp objects); falls; contact with heat and hot substances; exposure to smoke, fire, and flames; transport crashes; unintentional threats to breathing that create suffocation risk; unintentional poisoning by and exposure to noxious substances; unintentional drowning and submersion; and exposure to electrical currents. Example statements appear below: Participants also recommended 3 facets of content design that could potentially increase the use of app-based intervention by caregivers: (1) providing professional and believable content to gain the trust of app users, (2) using plain language to make the contents easy to learn, and (3) providing easy-to-implement interventions to improve the practicability of applying lessons from the app-based intervention.
I

Forms of Learning Delivered by an App-Based Intervention

Several forms of learning from an app were proposed by the participants. Most frequently mentioned were short written alarms or warnings with pictures, cartoon vignettes, video testimonials, and interactive games. The participants believed these 4 forms would make learning easier and more engaging. Participants also mentioned the desire to learn the knowledge interactively with their children, a strategy they felt might maximize the effectiveness of the injury prevention program:

Pictures and video are easy to understand for both adults and children and might be the best way to disseminate the prevention knowledge. I also think both caregivers and children need to learn unintentional injury prevention. The easier the learning form, the better the learning outcome. In addition, it is better to include short warning words in the pictures to more effectively stop the children from adopting dangerous behaviors. [Participant #DWT-B-01]

Timing for App-Based Learning

There was some variation across caregivers, but the general consensus was that they preferred the opportunity to use the app twice a week, for between 2 to 5 min at each session. As an example, a participant made the following statement:

The duration of the app-based learning should not be too long. I think "no more than 5 minutes" is fine for me because over five minutes learning would make me dizzy. Some participants felt the length of time they used the app would depend greatly on how much they enjoyed using it and that tangible cases and visual attraction would improve the authority and authenticity of the training, increasing their desire to use the app.

Functional Design of the App-Based Intervention

Almost all focus group participants mentioned interactive features as an indispensable function of the app-based intervention. They explained that interactive portions would maintain the attention of app users and increase the effectiveness of the intervention. Web-based chats, forums, and message boards were suggested as ways to implement interactive processes:

Web-based chatting is the best way to solve the problems that we encounter in our lives. [Participant #HHY-A-01]

As we know, our kids may encounter various unintentional injuries in daily life. If the app intervention can set up a module listing possible ways to prevent common injury causes, it would be great. In addition, it would be attractive to me if the app had a forum in which I could discuss these topics with other caregivers who confront the same questions. [Participant #YL-C-06]

In addition, a customer service agent who was accessible through Web-based chatting and a frequently asked questions module was suggested by the caregivers participating in the focus groups.

Beyond interactive portions of the app, most caregivers in the focus groups felt that a survey with feedback would be an important function of the app-based intervention. Such a component would allow app users to be aware of children's unintentional injury risk in their homes and to obtain tailored professional recommendations to prevent injuries to their children. Both Web-based and printed questionnaire surveys were recommended as appropriate approaches to obtain feedback from professionals.

In addition, some participants suggested surveys and feedback should be scheduled to be brief (eg, 5 min to complete each survey) and to be repeated regularly (eg, 3-4 times per year). Parents felt their responses might change as children develop new skills with older age, and therefore, tailored information that coincides with their children's development would be valuable. Participants recommended that questionnaires be based on items with categorical response options (eg, true or false) that could be responded to quickly. A few caregivers suggested providing survey participants with small gifts or bonuses to maintain their adherence to the intervention. Furthermore, to increase compliance in completing Web-based surveys, participants suggested the use of short message service text message reminders:

I'd like to use Web-based questionnaires to send feedback. I think survey questions with single options are much better than open-ended survey questions because I really do not know what should be filled in in many cases. [Participant #HHY-B-03]

Both Web-based and printed questionnaire surveys are acceptable for me, but a Web-based questionnaire survey is preferred since it is more convenient and takes less time than a printed questionnaire survey. I probably would agree to complete a paper questionnaire survey if I received small gifts after I completed it. [Participant #HHY-B-04] Some caregivers extended the survey idea further, suggesting that it would be valuable to have the app customized to their preferences and priorities in terms of content, form, frequency, and interface of the app. As an example, preventive content concerning riding a bicycle may not be of interest or relevant to caregivers whose children are younger than 3 years, as most Chinese children do not learn to ride bicycles until they are at least 4 years old. Thus, parents suggested the app be tailored so that such segments would be omitted in their version of the program.

Design of the Interface

Participants stated that they would like to choose the app interface (eg, color and style) based on their preferences. During the group discussion, the moderator demonstrated various interfaces, including the layout of 3 existing apps, to obtain participants' opinions on the app design. Of the 3 options offered by the research team, most caregivers chose a cartoon style. A few caregivers preferred a simple interface with 3 replaceable pages, and fewer still selected a simple interface with only 1 page.

Installation and Registration

Caregivers strongly suggested the app should take up only a small amount of memory space on their smartphone and that it should require a simple registration procedure. They believed a user manual or help module might assist them with app use, and they requested an informational module that described the purpose, details, and benefits of the project before starting to use the app-based intervention.

Privacy and Data Security

Many caregivers expressed privacy and security concerns about using the app, especially if they were requested to provide sensitive information about their children (eg, home address and activities and locations where children engage in those activities). They did state that they would trust apps downloaded from officially certified app shops or promoted by official agencies such as preschools.

Web-Based Survey Results

Participants

In total, 1505 valid questionnaires were collected through the Web-based survey, including 687 from men (45.65%) and 818 from women (54.35%). The respondents supervised 807 children aged 0 to 3 years (53.62%) and 698 children aged 4 to 6 years (45.97%). The proportion of male to female children supervised was 1.33 ( Table 1).

Content of App-Based Intervention

Of the 1505 participants who completed the Web-based survey, 1313 (87.24%) expected the app would teach them relevant knowledge concerning unintentional injury prevention for children aged 0 to 6 years. Participants felt it would be valuable to learn knowledge about preventing the 10 major causes of child unintentional injury at different rates: contact with heat and hot substances (534/1313, 40.67% felt it would be valuable to learn); inanimate mechanical forces (including being pinched between 2 surfaces, such as in the doors of elevators, or cut or punctured by sharp objects; 520/1313, 39.60%); falls (449/1313, 34.20%); transport crashes (362/1313, 27.57%); unintentional threats to breathing (361/1313, 27.49%); exposure to animate mechanical forces (317/1313, 24.14%);exposure to smoke, fire, and flames (210/1313, 15.99%); unintentional poisoning by and exposure to noxious substances (146/1313, 11.12%); unintentional drowning and submersion (113/1313, 8.61%); and exposure to electrical currents (108/1313, 8.23%). The differences were statistically significant (χ 2 9 =989.7; P<.05; Table 2).

Forms of App-Based Learning

The 5 most frequent learning forms that participants stated they would like to use were as follows: (1) short written alarms or warnings with pictures, (2) video testimonials, (3) cartoon vignettes, (4) pictures, and (5) interactive games, which were, respectively, preferred by 62.30% (818/1313), 54.53% (716/1313), 40.67% (534/1313), 38.31% (503/1313), and 28.03% (368/1313) of respondents (note that respondents were permitted to select multiple preferred learning forms). Differences were significant (χ 2 4 =393.0; P<.05).

Preferred Times, Frequency, and Duration for Using App-Based Learning

Caregivers varied in their preferred frequency for using an app to learn about unintentional child injury prevention.

Suggested Framework to Design the App Based on Focus Group and Web-Based Survey

On the basis of responses from the focus groups and the Web-based survey, as well as the 3 principles of TPB (attitudes toward the behavior, subjective norms, and perceived behavioral control) and the 3 aspects of the FRAME model (learner, devices, and social aspects), we developed a framework to guide design of an app to teach caregivers knowledge about preventing child unintentional injury. The design included 5 primary components: (1) content design, (2) functional design, (3) interface design, (4) installation and registration, and (5) privacy and data security, plus 15 subcomponents. Details, including the theoretical basis for each recommendation of the participants, appear in Table 3. Table 3. Framework for developing an app to teach knowledge about preventing child unintentional injury to Chinese caregivers of children aged 0 to 6 years.

Theoretical basis Description Requirements

Content, form, time, frequency, and duration of knowledge dissemination

Learner aspect (FRAME a model) Ten major unintentional injury causes: contact with heat and hot substances; inanimate mechanical forces; falls; transport crashes; unintentional threats to breathing; animate mechanical forces; exposure to smoke, fire, and flames; unintentional poisoning by and exposure to noxious substances; unintentional drowning and submersion; and exposure to electrical currents, with the suggested proportions for knowledge disseminations of these 10 injury causes at the suggested proportions Tied to theory-based goals to improve attitudes, alter subjective norms, and increased perceived control to perform child unintentional injury prevention behaviors

Learner aspects (FRAME model) Short written alarms or warnings with pictures, cartoon vignettes, video testimonials, and interactive games, with the suggested proportions of 2:2:1:1 across the 4 forms

Form

Learner aspects (FRAME model) Preferred time to learn (evening, afternoon, or noon) Time

Learner aspects (FRAME model) Twice a week Frequency

Learner aspects (FRAME model) No more than 5 min per time Duration

Learner aspects (FRAME model) Professionally disseminated contents Other attributes

Learner aspects (FRAME model) Using plain language Learner aspects (FRAME model) Easy to implement

Functions

Social aspects (FRAME model); attitude and subjective norms (TPB) Regular communication between users and experts through online chat, forums, and message boards; frequently asked questions module; and Web-based customer service agents and ask-and-answer service Interactive style c Use Web-based and printed questionnaires and short message service text message reminders for surveys and to collect feedback, no more than once every 2 months Survey and feedback -After each survey, motivate users through virtual rewards Learner aspect (FRAME model) Allow personalized customization of contents, forms, frequency, and interface of app intervention Personalized customization

Design of the interface

Device aspect (FRAME model) Offer several choices of app interfaces: cartoon interface preferred for default Interface design

Installation and registration

Device aspect (FRAME model) Simple app registration procedure Registration

Device aspect (FRAME model) Minimize the size of app so smartphone storage is not used excessively Memory space

Device aspect (FRAME model) Provide informational background to users' informed consent process, so they understand before downloading and using the app intervention

Informed background

Device aspect (FRAME model) Provide a user manual and help module Manual and help module

Privacy protection and data security

Device aspect (FRAME model) Do not collect sensitive individual or family information such as name, home address, or family income Personal privacy and data security Device aspect (FRAME model) Ensure safe sharing and storage of data, for example, by using an individualized password a FRAME: Framework for the Rational Analysis of Mobile Education.

Discussion

Principal Findings

Using a mixed-method study, we explored the preferences of caregivers of Chinese children aged 0 to 6 years for an app offering knowledge about child unintentional injury prevention. Study results helped us establish a framework for the app that includes 5 primary components (content design, functional design, interface design, installation and registration, and privacy and security) and 15 subcomponents focused on educating Chinese caregivers on child unintentional injury prevention strategies. The framework in Table 3 lists details to design the app, including the theoretical basis for those details.

Comparison With Prior Work

In addition to replicating previous work underlying the urgency to implement unintentional injury prevention education strategies for caregivers of young children in China [9], this study explored caregivers' attitudes and preferences for app-based learning, offering comprehensive and valuable guidance for the design of an app-based intervention. A few previous studies, generally based on small-sample qualitative designs, proposed some fragmented recommendations on the design of health-related apps that parallel some of our findings [30][31][32][33][34]. For example, Gkatzidou et al [30] suggested that privacy and security, credibility, user journey support, and the task-technology-context fit from the patient's perspective should be considered in the design of health care apps to maximize their acceptability. Similarly, Curtis et al [34] reported preferences for a healthy eating app targeting parents about child weight management; they identified 4 main themes for app design: app features, time saving and convenience, aesthetics, and gamification.

Our findings agree to a large extent with previous work in the design of app interventions, as we uncovered user preferences to emphasize a user-centered design [35,36], including particular emphasis on the importance of inclusion of games [35] and on the protection of user privacy [36].

Implications

This study has important implications. First, the research will lead to the development of an app designed to help caregivers learn how to reduce injury risk among their children. Although mobile device interventions are most effective when they are based upon theory [37], many existing health-related mobile apps do not incorporate features derived from evidence-based theoretical frameworks, health behavior change theories, and clinical guidelines [38,39]. We propose research grounded in theory that will lead to the development of an evidence-based intervention program. More broadly, the framework we present may prove valuable to guide the design of app-based interventions to target other diseases and injuries in China, as users' preferences are likely to be similar across content areas.

With cultural tailoring, it might also be useful in other countries, and ultimately, the recommendations could facilitate using smartphone technology to improve public health in China and beyond. Furthermore, our methodological strategies proved useful and could be replicated. The 2-step mixed-method approach could be extended to guide the development of theory-driven frameworks or interventions for various health issues in various cultural contexts.

Strengths

Our study has 2 strengths. First, we adopted a mixed-method design with large sample sizes. We conducted a rigorous mixed-method study and analyzed data and reported results in accordance with COREQ [28] and CHERRIES [29] guidelines. In particular, we adopted the principles of an effective intervention program (eg, implementing various learning methods, sufficient dosage, theory driven, and appropriately timed) [20] and used the TPB [21] and the FRAME model [23] to design the semistructured discussion guide for focus groups. Use of the TPB and the FRAME model ensured that the framework we proposed both addresses the needs of users (FRAME) and creates a situation that will optimally encourage appropriate behavior change among Chinese caregivers and yield reduced risk of unintentional injury among their young children (TPB).

Second, we implemented rigorous science to develop the focus group guides and the questionnaire for the Web-based survey. Methodological strategies included 2-round discussions within the research member group and pilot tests to improve the experimental stimuli and ensure they were grounded in theory and constructed to yield the information we desired.

Our results also extend fragmented recommendations from previous publications [30,32,33] and provide systematic qualitative and quantitative guidance to design an effective app-based intervention. We expanded previous recommendations for privacy and information security by providing detailed suggestions concerning handling of sensitive information.

Limitations

This study has several limitations. First, focus groups were conducted using standard procedures and moderated by skilled individuals, but like all focus groups, the conversation was susceptible to bias and to swaying of individual opinions by dominant participants [40]. Second, the app framework was generated under the context of Chinese culture and the primary factors for child injury in China. The framework might need tailoring and adjustment if it were applied to other age groups, other diseases, or other countries.

Conclusions

A theory-driven and evidence-based framework was established to guide the design of an app-based unintentional injury intervention program for the caregivers of Chinese children aged 0 to 6 years.

[Participant #MWD-B-10] Participants also recommended evening as the best time of the day to interact with the app: For me, it is particularly suitable to learn injury prevention knowledge at 8 or 9 o'clock in the evening. At that time, I have finished housework and my child hasfallen asleep. [Participant #MWD-B-09]

Table 1 .
1Demographic characteristics of caregiver participants.Web-based survey participants, n (%) Focus group participants, n (%) Variable a Information was not collected. b Children of the participant supervisors (90 parents and 6 grandparents) in the focus groups, excluding participants who were preschool teachers.Caregivers

1505 (100.00)
108 (100.0)
Total

Sex

687 (45.65)
12 (11.1)
Male

818 (54.35)
96 (88.9)
Female

Age group (years)

33 (2.19)
2 (1.9)
≤19

805 (53.49)
48 (44.4)
20-29

604 (40.13)
46 (42.6)
30-39

62 (4.12)
8 (7.4)
40-49

1 (0.07)
4 (3.7)
≥50

Relationship with children

-a
90 (83.3)
Parent

-a
6 (5.6)
Grandparent

-a
12 (11.1)
Preschool teacher

Children b

Sex

859 (57.08)
49 (51)
Male

646 (42.92)
47 (49)
Female

Age group (years)

205 (13.62)
34 (35)
≤1

602 (40.00)
27 (28)
2-3

698 (46.38)
35 (37)
4-6

to other options, an app-based intervention developed by a professional team would provide rich content. Thus, it will be convenient to search for the knowledge we need. For example, knowledge we can obtain in WeChat is hard to retrieve when we need it again. [Participant #YL-B-06]

really worry about injury from falls because children always like jumping from high places to lower places, which highly increases the risk of getting injured. In addition, road crashes are another important injury that I want to learn about, because many children run across or play in the street even when they see a car nearby. [Participant #DWT-B-03] Falls and burns are common types of unintentional injury for children ages 0-6 years at home. Relatively speaking, their harm to children is not as great as other types, such as electricalcurrents, which is one of the most dangerous injury causes for children. [Participant #HHY-B-04]

Table 2 .
2Number of participants who expressed a desire to learn knowledge about preventing major types of child unintentional injury causes.n (%)

b TPB: Theory of Planned Behavior. c Not applicable. JMIR Mhealth Uhealth 2019 | vol. 7 | iss. 4 | e11957 | p. 9 http://mhealth.jmir.org/2019/4/e11957/ (page number not for citation purposes)
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AcknowledgmentsThis work was funded by the National Natural Science Foundation of China (No. 81573260). The funding body had no role in the design, collection, analysis, or interpretation of this study.Authors' ContributionsGH and PN contributed to the design of the study. PN performed the data analysis. PN, DG, and GH drafted the manuscript. All authors finalized and approved the final version of the manuscript.Conflicts of InterestNone declared.Multimedia Appendix 1Procedure and discussion guide for focus groups.[DOCX File, 21KB-Multimedia Appendix 1]Multimedia Appendix 2Web-based survey questionnaire.  Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work, first published in JMIR mhealth and uhealth, is properly cited. The complete bibliographic information, a link to the original publication on http://mhealth.jmir.org/, as well as this copyright and license information must be included.
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Predictive Value of Biomarkers for Normal and Pathological Ageing in a Rat Model
2015. 2015

A ; Stramek
W ; Wassif
K ; Evans
J Barkans
H A Macqueen
Article Research
A Stramek
Department of Life, Health and Chemical Sciences
The Open University
Walton HallMilton KeynesUK

Department of Clinical Biochemistry and Metabolic Medicine
Department of Life, Health and Chemical Sciences
Bedford Hospital NHS Trust
Kempston RoadBedford, UK

The Open University
Walton Hall, Milton KeynesMK7 6AAUK

Wassif W
Evans K
J Barkans
Department of Life, Health and Chemical Sciences
The Open University
Walton HallMilton KeynesUK

Macqueen Ha email@example.com
Department of Life, Health and Chemical Sciences
The Open University
Walton HallMilton KeynesUK

Professor,H Macqueen
Department of Life, Health and Chemical Sciences
The Open University
Walton HallMilton KeynesUK

Stramek A
Wassif W, Evans KBarkans J, MacQueen HA

Predictive Value of Biomarkers for Normal and Pathological Ageing in a Rat Model

International Journal of Food Science
3S32015. 201510.19070/2326-3350-S103003Received: September 24, 2015 Accepted: December 14, 2015Open Research Online The Open University's repository of research publications and other research outputs For guidance on citations see FAQs. Version: Version of Record Link(s) to article on publisher's website: Copyright and Moral Rights for the articles on this site are retained by the individual authors and/or other copyright owners. For more information on Open Research Online's data policy on reuse of materials please consult the policies page. 16 http://scidoc.org/IJFS.php International Journal of Food Science, Nutrition and Dietetics (IJFS) *Corresponding Author: Citation: Stramek A, Wassif W, Evans K, Barkans J, MacQueen HA (2015) Predictive Value of Biomarkers for Normal and Pathological Age-ing in a Rat Model. Int J Food Sci Nutr Diet. S3:003, 16-22.
Sub-optimal diets are widely known to be associated with disease. This study investigated two questions: first, whether a Western affluent diet affects long-term health in the rat, and second, whether, and to what extent, early changes in blood biomarkers can predict specific pathologies. A rat model was used in which animals were fed from weaning either a control diet or a high fat, low protein, energy dense Western affluent diet. Rats were harvested at either 12 or 18 months, and at these time points blood samples were taken and various clinical biomarkers measured using a hospital analyser or by ELISA. Predictive biomarkers should show small changes at early time points and larger changes at later time-points, and be associated with disease(s). Tissues were examined for overt pathology, and data were mined to establish links between pathologies and specific blood markers that might be used predictively. Results showed that the Western affluent diet is associated with ill-health, with a relative risk of developing disease 4.5 times higher than for the control diet. Furthermore we conclude that triacylglycerol, HDL cholesterol, the ratio of total cholesterol to HDL cholesterol, creatinine and alanine aminotransferase were good predictors of disease as across the cohorts they showed altered levels before the clinical development of pathologies, in agreement with our hypothesis. Special Issue on "Dietary patterns and health risks" Stramek A, Wassif W, Evans K, Barkans J, MacQueen HA (2015) Predictive Value of Biomarkers for Normal and Pathological Ageing in a Rat Model. Int J Food Sci Nutr Diet. S3:003, 16-22.

Introduction

The ageing process in mammals is complex and is often accompanied by a number of chronic and acute pathologies whose incidence increases with age. Moreover life events can impact substantially on both the ageing process itself and on the incidence and impact to the individual of the medical conditions, and a number of conditions can be influenced by lifestyle factors such as diet and exercise. Sub-optimal diet has been linked to a large proportion of the global burden of disease [1]. If clinical conditions can be alleviated by non-pharmacological interventions such as a change in diet, as is the case for Type II diabetes [2][3][4], a significant cost saving could be made in sustaining an ageing population.

Over the course of evolution mechanisms have developed that allow individuals to withstand periods of nutritional deficiency, and this has been discussed elsewhere [5]. However, in developed countries in the present day the more common problem is one of nutrient over-availability, and changed patterns of food consumption have led to an obesity epidemic in the human population and an increase in the incidence of conditions such as diabetes, coronary heart disease and joint disease, all of which impact significantly on quality of life. Such diseases are no longer thought of as diseases of old age, since they are increasingly prevalent in much younger age groups. We and others have studied the effects of diet on markers of disease [6][7][8][9]. Rats are frequently used in such research because they bring all the benefits of a well-understood laboratory model, and are not fussy eaters. Rats are also subject to many deleterious effects of ageing, and long-term studies using rats reveal the gradual onset of age-related pathologies.

The experimental diet that we have used here models the kind of sub-optimal diet that is increasingly eaten in affluent countries: it is high in fat and low in protein and fibre compared to an optimal control diet. The two diets used here are equivalent in carbohydrate content, but the high fat content of the Western affluent diet makes it more energy-dense than the control diet, which is a chow made by the same manufacturer but optimised for rodent maintenance [10]. In addition to cardiovascular problems [11], high-fat diets of this kind are known to promote the development of fatty liver disease and other metabolic abnormalities [8], to affect the structure of the kidney [12], and to affect the http://scidoc.org/IJFS.php performance of the immune system [13]. All of these parameters are important for the development of chronic health conditions, and at a population level there is interest in identifying specific biomarkers that can be used to predict the onset of cardiovascular, hepatic and renal pathologies, and hence to manage the risks associated with them.

Here we describe a study undertaken to investigate two questions: first, whether a Western affluent diet affects long-term health in the rat, and second, to what extent early changes in blood biomarkers can predict specific pathologies. This information could be used to predict future illness so that dietary or other interventions may be made before the clinical onset of the disease. In order to be a reliable predictor a biomarker must be linked to a known abnormality, and must be detectable above background level at a time before the abnormality has become a clinical problem.

In this study we examined the gross morphology of target organs, and levels of biochemical markers used clinically for the diagnosis of cardiovascular, liver and kidney abnormalities. To address our first question we compared the prevalence of pathologies in a control population eating an optimal diet with that in a population eating a high fat, low protein Western affluent diet. To address the second question we examined the profile of blood biomarkers in these two groups at three time points: weaning, 12 months and 18 months of age. We sought biomarkers that showed small changes at the 12 month time point and larger changes at the 18 month time point, and that could be linked to an increased level of particular pathologies.

Materials and Methods

Animals

Animals were male Sprague-Dawley rats, bred in-house and maintained in an enriched environment on a 14h light: 10h dark cycle. Animals were kept and treated in accordance with the UK Animals (Scientific Procedures) Act, 1986, and the experiments were approved by the Open University Animal Welfare and Ethics Review Board. Rats were caged in Scantainers (Scanbur Technology, Denmark) at ambient temperature (19-23˚C) and 50 ± 10% humidity. They were given cardboard tubes and aspen wood blocks (LBS Biotechnology, UK). Rats were weaned at three weeks old directly onto the experimental or control diets, which were available ad libitum, and were maintained at two littermates per cage until the age of 12 months or 18 months, when they were used. During this period each rat was weighed weekly until 5 months of age and monthly thereafter, and examined visually for signs of ill-health. Because the rats were given wood blocks to gnaw, no dental problems were encountered. Growth curves for the animals are shown in Figure 1a. Hepatomegaly is frequently associated with high-fat diets [14], but Figure 1b shows that in this study the size of the liver remained in proportion to body mass across all the animal groups. All animals were harvested at the same time of day, 5 hours into the light phase. A total of 35 rats were used in this study. Seven weanlings were used as controls; these animals were assumed to be in optimal dietary conditions although not necessarily representative of older, weaned animals eating an optimum diet. At each time point (12 months and 18 months) 4 animals eating the control diet and 10 animals eating the experimental diet were harvested. The reason for the smaller number of control animals is that we already have substantial data on such animals [8,14] and for ethical reasons did not wish to repeat this data collection needlessly.

Diets

The control diet was RM3 and the experimental diet was Western RD (WRD), both purchased from Special Diet Services, Witham, Essex, UK. The components of both diets, taken from the manufacturer's data [10], are shown in Table 1. It can be seen that the principal differences between the diets are the content of fat (higher in WRD) and protein (lower in WRD). WRD is also lower in fibre and ash, suggesting that this diet may be less able to support gut function. The carbohydrate components of the two diets are equivalent, but the WRD is more energy-dense, as estimated from the Atwater Fuel Energy calculation [15].

Sampling

At the time of harvest, rats were weighed then deeply anaesthetised with Isofane (Novartis Animal Health UK Ltd). Approximately 4 ml of blood was obtained by cardiac puncture. Following coagulation at room temperature the clots were spun down and the supernatants were removed and stored at -80°C until analysis. Immediately following cardiac bleeding, the rats were killed by intracardiac injection of Pentoject (Animal Care Ltd, UK). Organs were then removed for visual assessment of size, shape, colour and the presence of any gross abnormalities such as ulcers or tumours. They were not routinely assessed histologically. Organs removed for examination were brain, heart, stomach, intestines, liver, pancreas, spleen and kidneys. Special Issue on "Dietary patterns and health risks"

Biochemical analyses

Serum metabolites, electrolytes, renal profile, liver profile and lipid profile were measured using a Cobas Integra 800 analyser (Roche Diagnostics, Mannheim, Germany). The analytes tested are listed in Table 2. Sera from the same animals were also used in triplicate in ELISAs to determine circulating levels of insulin. All ELISA assays were carried out using the Alpco Insulin (rat) ultrasensitive ELISA kit (Stratech Scientific Ltd, Newmarket, UK).

Statistical analyses

Data are expressed as mean ± SEM. Differences between means were assessed by ANOVAs and t-tests as appropriate, using Prism v5 (GraphPad Software, San Diego, California, USA, www.graphpad.com). Since sample sizes were small and unequal, and had unequal variances (with the variances being greater in the larger experimental diet groups) ANOVA cannot detect small but significant differences, and it is recommended to use t-tests for analysing these data [16]. Differences were considered statistically significant at the P < 0.05 level. Relative risk was calculated from the equation:

Relative risk = probability of disease in rats eating WRD/probability of disease in rats eating RM3.

Results and Discussion

Animal health

Rats are commonly used in dietary studies because they are not fussy eaters and are physiologically robust [17]. In this study we observed the health status of rats eating one of two diets, and at three time points. We measured the levels of standard biochemical markers used diagnostically, and related them to the pathologies that we observed. We were particularly looking at a population level for biochemical markers that showed significant changes by 18 months on a sub-optimal diet and which also showed a lesser change in the same direction at 12 months on the diet, such that the marker could be used clinically in a predictive way to allow early therapeutic interventions.

A number of gross pathologies were observed when the animals were harvested. At the start of the experiment all rats were healthy and active, with good coats and general appearance. One rat eating the control diet and one eating the WRD had to be culled before 12 months due to poor health. Two further rats eating the experimental WRD were found dead in their cages between 12 and 18 months. These four animals were omitted from the analysis. Among the animals which survived until the experi-mental end-point, 4 out of 13 rats harvested at 12 months, and 7 out of 11 rats harvested at 18 months were found to have one or more pathologies upon dissection in spite of appearing in good health judging by coat condition, food intake and activity. Overall, 12.5% of animals eating the RM3 diet (all ages) had an overt pathology, compared with 56.3% of those eating the WRD (33.3% at 12 months and 85.7% at 18 months; see Table 3). This means that over the course of the study the relative risk of developing a pathology was 4.5 times higher for rats eating the WRD than for those eating the RM3 diet. Table 3 also shows that all but one of the animals exhibiting pathologies had levels of specific biomarkers either elevated or depressed beyond one standard deviation from the population mean. The specific analyte changes were consistent with the observed pathologies; for example kidney abnormalities were accompanied by changes in creatinine and urea levels in the blood. This indicates that the analytes used do indeed reflect particular health problems. These results clearly demonstrate that rats can appear to be healthy while nevertheless harbouring a variety of pathologies, and provide further evidence that a sub-optimal Western affluent diet cannot support long-term health.

Biochemical analysis

Following biochemical analysis, the majority of analytes showed no differences between the cohorts of animals eating the two diets or at the ages examined. These analytes included C-reactive protein, a marker of immune function, so in this study we were unable to detect any changes in the immune status of the animals. Statistically significant changes were, however, observed in the following analytes: iron, ALT, AST, albumin, creatinine, cholesterol, HDL-cholesterol, TAG, cortisol, glucose and insulin. We detail these differences below.

Iron:

Compared with the level seen at weaning, iron levels decreased with age, and were significantly lower (P < 0.01) in 18 month old animals compared with weanlings ( Figure 2a). There were no significant differences between the diets. This was the only change observed in serum electrolytes, in contrast to previous work that showed a reduction in potassium level with age [8], and this may be accounted for by the different diets used in these two studies.

Cortisol: Corticosterone, not cortisol (which is measured by the clinical analyser) is the primary glucocorticoid stress hormone in the rat; nevertheless cortisol measurements can be used as a proxy measurement for stress as there is considerable overlap in the values for the two hormones [18]. There was a slight but not significant upward trend in cortisol in WRD animals compared to those eating the RM3 diet (Figure 2b). Levels of glucocorticoid Special Issue on "Dietary patterns and health risks" stress hormones have been reported to rise with age in the rat [19] and we also observed this trend, although the rise was not statistically significant. However we found that cortisol levels were high in weanlings compared with the other groups (Figure 2b). Similar findings have been reported by others, and this effect is thought to be a response to the stress of weaning [20].

Markers of liver function:

ALT rose with age (RM3 diet) but dropped in the WRD animals ( Figure 2c). This pattern was also seen to a lesser extent with AST (not shown). The significant drop in 18 month WRD animals (P < 0.001) was accompanied by a non-significant but consistent decrease at 12 months; thus ALT is a possible candidate for a disease biomarker, although it may be an imprecise marker since low ALT was associated with an atrophied brain, ulcerated kidney and stomach, and leukocytosis (Table 3). Albumin remained stable in the RM3 animals, but there was a highly significant drop (P < 0.001) in the WRD animals at 18 months which was not seen at 12 months (Figure 2d). Liver enzymes are frequently used as clinical markers of a range of conditions, and we have previously found a number of changes with age and diet in Sprague-Dawley rats [8,14]. Although in humans an enlarged liver is frequently associated with ill-health, we did not observe this phenomenon, and all the livers remained in the correct proportion to whole body mass (Figure 1b). A histological examination of some of the livers (not shown) showed glycogen and fatty deposits increasing with age, and noticeably more prominent with animals eating the high-fat diet, consistent with our previous findings and those of others [14,21]. In this study we observed that the rise in AST and ALT with age seen in control animals was substantially abolished in WRD animals, which also showed a significant drop in levels of albumin at 18 months (Figure 2c and d; P < 0.001). These results suggest a profound change in the liver metabolism of WRD animals that can be linked to general ill-health (as revealed by the pathology data) but not to any specific condition.

Markers of kidney function:

In view of the number of kidney abnormalities that we observed post-mortem we anticipated that there would be changes in the levels of the kidney markers that we used, particularly since levels of these markers are known to vary with the protein content of the diet [22]. Across the experimental groups there were no changes in levels of urea ( Figure  2e). Mean creatinine levels did not change with age in the RM3 animals, but increased with age in the WRD animals (Figure 2f). Individual rats with kidney abnormalities had elevated creatinine levels compared with normal members of their cohort (Table 3), accounting for the significant rise in the mean value (and SEM) seen in the 18 month WRD group. This suggests that, in addition to its current diagnostic use [3], creatinine could be used in a predictive way to allow early therapeutic intervention for kidney abnormalities. Based on these data, further study of creatinine expression during disease development is warranted.

Lipid profiles: Blood lipids are well known as markers of illhealth, and are particularly linked to cardiovascular disease. In these experiments total cholesterol increased in the control animals between weaning and 12 months of age (P < 0.05), but remained stable thereafter. The WRD 12 month group had a similar cholesterol level to the RM3 group, but at 18 months the mean had increased to a highly significant level (P < 0.001; Figure  2g). In these studies LDL cholesterol remained fairly stable (not shown) but the HDL cholesterol showed an interesting pattern.

In the control RM3 rats HDL cholesterol increased significantly by 12 months, but this increase was completely abolished in the WRD animals at both 12 and 18 months (Figure 2h). This was not unexpected, given the fat content of the WRD ( Table  1). The consequence of this change was that the total cholesterol: HDL-cholesterol ratio showed significant changes with diet, with the 12 month value being a good predictor of the 18 month value (Figure 2i). Likewise, whereas the control animals showed a non-significant increase in TAG level at 12 months compared to weanlings that then remained stable, the WRD animals showed a significant rise (P < 0.01) at 18 months that was predicted by the 12 month pattern (Figure 2j). A separately conducted multivariate analysis of volatile organic compounds in the faeces of the same animals [23] also suggested that TAG, total cholesterol, and the total cholesterol : HDL-cholesterol ratio could be reliably used as predictive markers.

Glucose and insulin: As noted above, the animals in this study were fed ad libitum, and all were harvested at the same time in the dark-light cycle. The carbohydrate content of the two diets was similar (Table 1), so in spite of the raised body mass and increased blood lipid markers exhibited by the WRD animals (Figures 1a  and 2g-j) there was no increase in circulating glucose levels. In fact by 18 months in both control and experimental animals blood glucose levels were significantly lower (P < 0.001) than at weaning and 12 months (Figure 3a).

An unexpected result was found when we examined the levels of circulating insulin. It has been shown in several species including the rat that circulating insulin levels rise with increasing glucose intake [24] and insulin levels are linked to diabetes. We observed no rise in insulin levels with either age or diet, and given  *** the carbohydrate content of the two diets noted above this was not surprising. However, insulin values fell over a wide range in all the cohorts, and the spread of values seen was similar regardless of age or diet (Figure 3b). Careful examination of the data revealed that no values fell between 0.7 and 0.9 ng/ml, suggesting that there were two insulin-secreting phenotypes: one giving values in the range 0.2 to 0.7 ng/ml (mean 0.35 ng/ml), and the other giving values in the range 0.9 to 2.3 ng/ml (mean 1.4 ng/ ml). These populations were significantly different by t-test (P < 0.001; Figure 3c). Differences in insulin levels were not associated with changes in glucose levels (Figure 3d), nor was there any correlation between insulin level and the level of any lipid markers (Figure 3e), terminal body mass (Figure 3f), or any specific gross pathology (Table 3). We consider it unlikely that the high values represent a post-prandial increase since rats are trickle feeders and do not exhibit the pattern of insulin secretion found in humans. The mean value exhibited by the higher population (1.4 ng/ml) is similar though slightly higher than that reported for baseline circulating insulin by others: 1 ng/ ml [24] or 1.09 ng/ml [25]. The mean lower value (0.35 ng/ml) is slightly below the range of 0.4-0.8 ng/ml reported for rats starved for 24 hours [25], but it should be noted that the rats in our study had not been starved and may just naturally exhibit a lower baseline circulating level. As far as we are aware this is the first report of different insulin secreting phenotypes in unstressed Sprague-Dawley rats.

Conclusion

This study focused on two questions: first, whether a Western affluent diet affects long-term health in the rat, and second, whether early changes in blood biomarkers can predict specific patholo-gies. The first question has an unequivocal answer, in that the health of animals eating this diet was notably worse than that of animals eating a control, optimised diet, as measured by gross pathologies observed post-mortem. The relative risk was 4.5 over the course of the study.

Regarding the second question, we have found limited feasibility for using known biomarkers as predictors of future disease, at least in the Sprague-Dawley rat. Five markers: HDL cholesterol, total cholesterol: HDL cholesterol (driven by the change in HDL cholesterol), triacylglycerols, ALT and creatinine clearly show a suitable pattern of change. The changes for these markers are consistent and statistically significant, although it is unclear at present whether they correlate with specific pathologies, and more work is required to confirm this. Our data bring together studies of individual animals in which biomarker analysis can be correlated with specific health status, and cohort studies in which trends can be more easily identified. Future work in this area should use longitudinal studies with repeated sampling from the same animals.

We also report a previously unrecognised phenotypic diversity in insulin production by Sprague-Dawley rats.

Figure 1 .
1(a) Growth of rats eating the control (closed symbols) or experimental (open symbols) diet over the period from weaning to 18 months of age; (b) relationship between the liver mass and whole body mass in animals at the time of harvest.

Figure 3 .Figure 2 .
32Circulating insulin and glucose in all groups of rats. (a) circulating glucose levels in all groups of rats; (b) insulin levels in control (closed symbols) or experimental (open symbols) groups; (c) two populations of insulin secretors; (d) relationship between circulating glucose and insulin levels; (e) relationship between levels of insulin and levels of lipid mark-ers; (f) relationship between insulin levels and body mass at the time of harvest. *** P < 0.001. Changes in biomarkers of health in blood samples from all groups of rats.

Table 1 .
1Components of the diets used in this study (from manufacturer's data).RM3 WRD
Fat (% w/w)
4.2
21.4
Protein (% w/w)
22.45
17.5
Fibre (% w/w)
4.42
3.5
Ash (% w/w)
8.05
4.1
Carbohydrate (% w/w) 50.4
50.0
Energy (kcal/g)
3.29
4.63

Table 2 :
2Analytes investigated in this study and their clinical significance. AP, alkaline phosphatase; AST, aspartate ami-
notransferase; ALT, alanine aminotransferase; GGT, γ -glutamyl transferase; HDL, high density lipoprotein; LDL, low
density lipoprotein; TAG, triacylglycerol; CRP, C-reactive protein.

Serum electrolytes Kidney function Liver function
Lipid profile
Other analytes
Sodium
Urea
Albumin
Cholesterol
Glucose
Potassium
Creatinine
Bilirubin
HDL-cholesterol
Insulin
Calcium
AP
LDL-cholesterol
CRP
Magnesium
AST
TAG
Oestradiol
Phosphate
ALT
Testosterone
Iron
GGT
Cortisol
Transferrin
Ferritin

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AcknowledgementsThe authors declare no conflict of interest. We would like to thank Dr Jane Loughlin for helpful comments on the manuscript. This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors. We acknowledge The Open University Biomedical Research Network for pumppriming funding.
High creatinine, urea, phosphate, magnesium, cortisol; Low ALT, AST, testosterone. Ast High, High AST, magnesium 2. High creatinine, urea, phosphate, magnesium, cortisol; Low ALT, AST, testosterone

High creatinine, urea, calcium, magnesium, cholesterol, TAG, cortisol. Low albumin, iron 5. High cholesterol: HDL, TAG, cortisolHigh creatinine, urea, calcium, magnesium, cholesterol, TAG, corti- sol; Low albumin, iron 5. High cholesterol: HDL, TAG, cor- tisol;

. Alt Low, Low ALT, albumin, AST

High calcium, cholesterol, TAG Special Issue on "Dietary patterns and health risks. High calcium, cholesterol, TAG Special Issue on "Dietary patterns and health risks"

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Dietary patterns and health risks. Hong KongThe Hong Kong Polytechnic UniversityTheme Edited by: Shun-Wan ChanSpecial Issue on "Dietary patterns and health risks" Theme Edited by: Shun-Wan Chan, The Hong Kong Polytechnic University, Hong Kong.