Study design and setting
This study is a prospective cohort study, which explores the characteristics and interrelationships of high-risk factors, functions, imaging examinations, genetic analysis, and biochemical indicators for CP, as well as the emotion and quality of life for caregivers. In addition, the longitudinal changes in function for CP and the emotions and quality of life for caregiver are analyzed. It was designed and will be conducted by the Department of Rehabilitation, Guangzhou Women and Children's Medical Center (GWCMC), China.
Participants and recruitment
Physicians of GWCMC make a diagnosis of CP for children according to the International diagnostic standards for CP. All departments in this hospital related to CP are encouraged to refer children for this study. In addition, we encourage hospitals in China to refer children with CP to the department of GWCMC for study. Details of this study are explained by researchers to children who met the inclusion criteria and their caregivers. After the child and their caregivers confirm their participation in the study, the signing of the informed consent form will be completed. A cohort information management system is instructed for the information recording, follow-up, and information management of CP. Figure 1 shows the details of the recruitment and follow-up of the study.
Selection criteria
Inclusion criteria: diagnosed as CP according to international diagnostic standards[2]; aged 1 to 17 years old; being able to visit the GWCMC for regular follow-ups (1-2 years old, follow-up every 2 months; 3-6 years old, follow-up every 3 months; 7-17 years old, follow-up every 6 months); parents and children (aged 8 and above) agree and sign an informed consent form.
Exclusion criteria: be diagnosed with progressive or neurodegenerative diseases; the clinical condition is unstable, making it impossible to complete clinical evaluation.
Sample size calculations
The sample size was calculated using PASS software (version 15.0), and the selected method was linear regression. The test efficacy was 0.95, the test level was 0.05. The value of β1 is 0.25, the standard deviation of the independent variable is 5.0, the standard deviation of the dependent variable is 2.0, and the calculated result is 1289. Considering a 20% dropout rate, the final sample size is 1547.
Data collection
During the first visit, the researchers will inform caregivers about the details of the present study before enrollment and signing the informed consent. Then the basis information inquiry will be completed by researchers and the baseline assessment of the participants and caregivers will be conducted. Researchers will inform the participants for the subsequent functional assessments, covering the gross and fine motor function, language and cognitive function, eating and drinking ability, social life ability, pain, sleep quality, behavior problems and quality of life of children with CP, as well as the emotion and quality of life of caregivers. Medical records of participants are accessible through the hospital information system. The results of imaging examination for each participant will be obtained from the medical imaging management system of GWCMC.
Each participant and their immediate families with blood ties will provide 4mL of peripheral venous blood. All the samples will be stored in the clinical biological resource library of GWCMC at -80 ℃. The samples will be used for the analysis of whole genome and 25 hydroxyvitamin D, transforming growth factor, creatine kinase, proinflammatory cytokines and other related blood indicators.
All the above results and information will be recorded in cohort information management system of CP. Any medical records related to the study will not be exposed without the permission of the principal investigator.
Measurements and procedures
After being diagnosed as CP, children attend the department of rehabilitation of GWCMC with their caregivers for clinical assessments, imaging examination and blood tests. Table 1 shows the overview of functional assessments for children with CP at different ages.
1.Functional assessments for children with CP
Clinical assessments involve gross and fine motor function, language and cognitive function, swallowing function, social life ability, pain, sleep quality, and quality of life. In addition to the baseline evaluation, participants and their caregivers will be regularly invited to return to the hospital for follow-ups. Follow up time is based on age groups considering the functional changes and follow-up compliance at different ages: participants aged 1-2, 3-6, and 7-17 years old receive follow-ups every 2 months, 3 months, and 6 months, respectively. All participants will be followed up until the age of 18 until the end of the study. Assessments will be conducted by well-experienced and qualified evaluators from the department of rehabilitation of GWCMC.
(1) Gross motor function
Gross motor function classification system (GMFCS) and gross motor function measurement (GMFM) will be conducted to evaluate the severity of gross motor function. The GMFCS is a five-level classification system, the severity of gross motor dysfunction in GMFCS level I to V aggravate sequentially[19]. GMFM covers five domains, including lying and rolling; sitting; crawling and kneeling; standing; and walking, running and jumping. Scores are expressed as a percentage of the maximum score [20].
(2) Fine motor function
Manual ability classification system (MACS) or mini-MACS, Melbourne assessment, Box and Block test and selective control scale of upper extremity will be used for the evaluation of fine motor function. Based on the performance of upper limb movement on daily activities, MACS or mini-MACS classifies the upper limb function of children as five levels. MACS and mini-MACS are shown good reliability and among children with CP aged 1-3 years and 4-18 years, respectively[21]. The Melbourne assessment is a criterion referenced tool designed to measure the quality of unilateral upper limb movement in children with a neurological impairment, covering four main elements (range of motion, accuracy, dexterity and fluency)[22]. Box and Block test mainly assesses dexterity of the upper extremity and eye-hand coordination [23]. Selective control scale of upper extremity allows evaluation of the features of impaired selective motor control at upper extremity joint levels, which has good validity and reliability in children with CP [24]. Well-experienced occupational therapists will complete all the assessments of upper limb motor function
(3) Muscle tone and contracture
The modified Ashworth scale (MAS) will be conducted for the evaluation of the muscle tone in this study. The degree of spasticity is divided into 6 levels according to the scale (0, 1, 1+, 2, 3, 4), with 0 being no increase in muscle tone and 4 being affected part(s) rigid in flexion or extension [25]. The contracture is determined based on joint deformities and range of motion by physiotherapists.
(4) Cognition and communication function
The ability of cognition will be assessed via Wechsler Intelligence Scale for Children-fourth edition (WISC-IV) or Wechsler Preschool and Primary Scale of Intelligence-fourth Edition (WPPSI-IV). WISC-IV and WPPSI-IV are the most popular intelligence tests in applied psychological practice aiming at school children and preschool children, respectively[26,27]. Communication function classification system (CFCS) allows clinicians to classify children’s communication skills into one of five levels of function[28]. Qualified speech and language therapists will conduct the assessments for participants.
(5) Eating and Drinking Ability
Eating, drinking and swallowing abilities of children with CP will be evaluated using eating and drinking ability classification system (EDACS) or mini-EDACS, which are aiming at children older than 5 years and children aged 1 or 2 years old, respectively. EDACS and mini-EDACS are classified into five distinct levels based on the key features of safety and efficiency [29,30]. The above assessments will be completed by experienced occupational therapists.
(6) Quality of life and social life ability
Quality of life and social life ability will be assessed using the Pediatric Quality of Life Inventory (PedsQL) 3.0 CP Module and social life ability scale for Chinese infant–junior school students, respectively. The quality of life assessment include sub-areas: daily activities, movement and balance, pain and hurt, fatigue, eating activities and communication[31]. Social life ability scale is comprised six domains: independent living, athletic abilities, operational abilities, communicative abilities, participation in collective activities, and self-management abilities[32]. The assessments will be reported by caregivers under the guidance of clinicians or therapists in this study.
(7) Behavior problems
The severity of behavior problems will be determine using the Conners' Parent Rating Scale. It is a scale recovering 7 factors: cognitive problems, oppositional, hyperactivity-impulsivity, anxious-shy, perfectionism, social problems, and psychosomatic[33]. In the present study, Parents will rate the scores based on daily performance of their child under the instruction of researchers.
(8) Nutritional status
Anthropometric parameters will be measured for the evaluation of nutritional status. Weight of each participant is measured using a digital weighing scale barefoot and with minimum clothing. Standing height of participants over 2 years old who are able to stand independently or under assistance is measured using a length board. For those who were unable to stand under assistance or have severe lower limb joint contracture/deformity, height is estimated according to Stevenson’s equation (height (cm)=(tibial length×3.26) + 30.8)[34]. Recumbent length of participants under 2 years old were measured using a length board. All the above anthropometric parameters for each participant are measured for three times and the average value is calculated. WHO Anthro software (version 3.2.2) for children aged 1-5 yeas or AnthroPlus software (version 1.0.4) for children over 5 years is used for the calculation of weight-for-age z-score (WAZ), height-for-age z-score (HAZ), weight-for-height z-score (WHZ) and BMI-forage z-score (BAZ). Based on the WHO growth charts and the standards of American Society for Parenteral and Enteral Nutrition, the nutritional status was classified into mild undernutrition (–2<WHZ/BAZ≤–1), moderate undernutrition (–3<WAZ/HAZ/WHZ/BAZ≤–2), severe undernutrition (WAZ/HAZ/WHZ/BAZ≤–3), overweight (< 5 years, BAZ>2; ≥5 years, BAZ>1), obesity (<5 years, BAZ>3; ≥5 years, BAZ>2) and normal[35,36].
(9 )Pain
Visual analog scale will be used for participants aged 7 and above who have a basic understanding of numbers. The scale is composed of a 100mm straight line, with one end representing "completely painless" and the other end representing "pain to the extreme". Participants will be asked to mark the corresponding position on this line to represent the intensity of the pain they suffer under the instruction of a trained nurse or physiotherapist[37]. For participants under the age of 7 or those who don’t understand the meaning of numbers, a trained nurse or physiotherapist will determine the pain intensity based on the participant’s facial expression according to the Faces Pain Rating Scale, which depicts the different facials expressions when children suffer varying degrees of pain[38].
(10) Sleep quality
In order to evaluate the sleep problems, sleep disturbance scale for children will be used for participants aged 6-18 years old [39]. Caregivers assesses the the occurrence of sleep disturbances over the past 6 months under the instruction of researchers.
2.Clinical assessments for caregivers
(1) Emotional problems
Self-rating anxiety scale and self-rating depression scale will be used to discriminate anxiety or depression from mood disorders, respectively. The items of the two scales cover a variety of anxiety or depression symptoms, with good psychometric credentials. Caregivers are given on a 4-point scale which range from 1 (none, or a little of the time) to 4 (most, or all of the time). Participants are instructed to base their answers on their experiences over the last week. Items include both negative and positive experiences, with the latter being reverse scored [40].
(2) Health-related quality of life
Caregivers complete the 36-Item Short Form Health Survey questionnaire (SF-36), for evaluating health-related quality of life under the instruction of researchers. The SF-36 is a self-reported scale, which measures eight scales: physical functioning, role physical, bodily pain, general health, vitality, social functioning, role emotional and mental health [41].
3.Imaging examination
During the study, each participant will be examined with brain MRI, diffusion tensor imaging (DTI), and magnetic resonance spectroscopy (MRS) once or twice to observe the brain damage. For participants at risk of epilepsy, EEG screening is required. Those with abnormal results should undergo follow-up within 3 months, and the subsequent follow-up time should be determined based on the EEG results. The frequency of hip joint X-ray examination is based on the clinical signs and GMFCS level of participants[42]. For children with signs of scoliosis, spinal X-ray examination is required, and regular follow-ups should be conducted based on the X-ray results and the age of the child. All the above imaging examinations and analysis are conducted by radiologists and technicians from GWCMC.
4. Analysis of whole-genome and biochemical indicators
Peripheral venous blood of participants and their lineal relatives (blood related parents and siblings) is collected once or twice by nurses for whole-genome analysis and detection of 25 hydroxyvitamin D, transforming growth factor, creatine kinase, proinflammatory cytokines and other related blood indicators. The blood collection operation is conducted by professional nurses, and the collection time is from 8:00 am to 12:00 am, with each person providing 4mL of blood each time. The blood is divided into 2 sterile collection tubes of 2mL and then each tube of blood sample is centrifuged into serum and blood cells. Pre processed samples are stored in the clinical biological resource library of GWCMC at -80 ℃ and waiting for analysis. Analysis of whole-genome and biochemical indicators is conducted by molecular biology and laboratory experts.
Data management and monitoring
1.Data management
The study intends to establish a cohort information management system for children with CP. The system is collected to the hospital information system and record, retrieve, and analyze relevant information, covering the modules of follow-up plan management, automatic reminders, and follow-up record management. A dedicated research coordinator is responsible for the management and data inspection for this system. To ensure data reliability, evaluators are not involved in data management. At the same time, the system is equipped with logic check function and information security protection measures, which can detect and correct abnormal data, and ensure the security of the data.
2.Reliability of the evaluation results
The evaluators involved in this study are all well-experienced with corresponding evaluation qualifications. In addition, only if the consistency of the same evaluation project is over 95% (more than 50 samples), they must undergo a test (over 95%) of the results before they participate in this study.
3.Blood sample quality control
All blood samples in this study are pre processed and stored in the biological sample library of GWCMC. Professionals are responsible for timely docking, transportation, and sample storage. The usage of samples needs to go through a strict approval process within the hospital and an informed consent is needed. Furthermore, molecular biology experts are responsible for the quality control of the analysis of whole-genome and biochemical indicators.
4.Reliability of the statistical analysis methods
We will invite 2 statistical experts to inspect and confirm the statistical methods used in this study, and conduct reliability tests on the statistical analysis results.
Statistical considerations
For continuous variables, we will use means and SDs to present normally distributed data; for other data, we will use medians and interquartile ranges (IQRs). Numbers and percentages will be used for categorical variables. Continuous variables will be compared using the t-test or Mann-Whitney test, categorical variables using the χ2 test or Fisher’s exact test. Spearman's rank correlation coefficient or Pearson Correlation Coefficient will be used to analyse the correlation among high-risk factors, functions, imaging examinations, genetic analysis, and biochemical indicators for CP, as well as the emotion and quality of life for caregivers. Logistic regression binary or multivariate logistic regression, linear/non-linear mixed-effects regression models and structural-equation models will be used to explore causal inference among the above indicators. Additionally, we will perform longitudinal structural-equation modelling or growth curve analysis to evaluate the temporal effects of function for CP and the emotions and quality of life for caregiver. All of the analyses performed in this study will be considered statistically significant at a level of 5%, and 95% CIs will be calculated for all of the data.