## 6 Critical Appraisal and Considerations for Future Trials  
The literature summarised in this review represents the highest level of evidence of the efficacy of vitamin D supplementation for preventing or managing GDM. Despite a number of trials examining this topic over the last decade, results remain largely divergent and, in general, there is no strong evidence to support the use of vitamin D supplementation for GDM prevention or management. However, like observational studies that are limited by confounding and the inability to determine causation, RCTs are also subject to a number of limitations that likely contribute to these inconsistent results and threaten their external (e.g., low response rates) and internal (e.g., poor adherence, withdrawal, unbinding) validity. These limitations are summarised below and should be considered in the interpretation of results and in the design and execution of future trials. First, RCTs often have low response rates and small sample sizes leading to poor external validity. This is demonstrated in the trials identified in this review, whereby half had less than 100 participants and most had less than 200 participants. The most recent trial by [PERSON] et al. [44] indicated that 220 participants would be required to achieve statistical power to detect differences in insulin sensitivity and fasting glucose, with even more required to detect differences in a binary outcome such as GDM. Hence, most existing trials were underpowered which may explain some of the null results or inconsistencies identified in this review. Moreover, self-selection bias is problematic in RCT study designs since participants who volunteer to take part are more likely to be healthy or health conscious and less likely to be vitamin D-deficient [86]. While this may be less problematic in trials of pregnant women (since many women are typically more health conscious during pregnancy), there will always be some uncertainty as to whether results can be extrapolated to the general population. Together, these factors can reduce external validity and introduce bias, and are important to consider when interpreting the overall evidence. A second key issue in vitamin D RCTs pertains to 25(OH)D levels and the common use of doses which are insufficient to adequately increase circulating 25(OH)D to target ranges. Indeed, a non-linear association between vitamin D status and disease risk has been demonstrated for all-cause mortality and cardiovascular disease, and may also exist for other conditions such as GDM [86]. This strongly suggests that vitamin D trials should include only participants with 25(OH)D concentrations below the inflection point where disease risk starts to rise, since supplementation would not be expected to reduce disease risk in people with 25(OH)D levels above this inflection point. This inflection point varies depending on the disease in question, with a previous review of the literature [87] concluding that the thresholds for beneficial vitamin D status could range from a 25(OH)D of 25 nmol/L for bone diseases to up to 100 nmol/L for cancer. Importantly, inflection points or thresholds for observing benefits in the context of pregnancy or GDM have not been established. Nevertheless, the presence of vitamin D-sufficient individuals in a trial is sure to weaken its statistical power because their risk of disease remains unchanged irrespective of whether they receive vitamin D or placebo [86]. This, in turn, can mask the potential benefits of vitamin D, leading to erroneous results and conclusions. The trials reviewed herein demonstrate this concept to some extent, since the two trials [43; 44] which found no effect on most or any of the measured glycaemic outcomes (including GDM risk) both reported a non-deficient mean baseline 25(OH)D level (69 nmol/L).