To our knowledge, this is the first major U.S. study that has examined upper limits of normal for ALT, a critical test for detection and monitoring of liver disease. The broad range across laboratories in ULN for ALT has been a vexing issue. Studies that have addressed the ULN of ALT have been concerned primarily with either the unfortunate variation across laboratories(2
) or have attempted to define and apply ULN to defined populations.(26
) Among the former, it is clear that there is more variation across laboratories than across analyzers, and that the reference populations from which ULN are defined have often been poorly characterized and may have included persons with liver disease. Variation among analyzers exists and may limit the application of an ALT ULN to the US population. However, among 11 clinical laboratories used by the Nonalcoholic Steatohepatitis (NASH) Clinical Research network (CRN), most of the variability in ALT ULN was attributed to differences in reference populations rather than to inter-analyzer variation.(3
) Among laboratories in the state of Indiana, the ALT ULN varied more than two-fold (range 31–72 IU/L); however, using standardized samples, variation among analyzers was statistically, but not clinically significant.(2
) Differences in manufacturer’s recommendations, the most common method for establishing ALT ULN, may have contributed to inter-laboratory variation. A multinational evaluation found very minor differences across three models of autoanalyzers for a reference sample with ALT activity of 39.7 U/L, which is within the range of activity that is of greatest interest for seeking inter-laboratory agreement.(34
) Although greater variation across analyzers is to be expected for enzyme activities than for chemical analytes, if should be possible to achieve greater harmonization of analytic approaches and reference ranges for ALT.
Most papers that have addressed the definition of ULN examined a low risk population only,(28
) while a few also considered the effect of hepatitis C.(26
) These studies were conducted in Europe and Asia, usually at a single center. The study cited most often and whose design was similar to the current study was performed on first-time blood donors in Milan, Italy.(26
) ALT discriminated asymptomatic donors harboring HCV from a low risk group who were not overweight or taking medication and who had normal blood glucose, triglyceride, and cholesterol concentrations. The 95th
percentile cut-offs for the low risk group (30 IU/L for men and 19 IU/L for women) were substantially lower than those for the low risk group in the current study (44 IU/L for men and 32 IU/L for women). Because of the lower cut-offs, sensitivity for detection of hepatitis C of 76% in the Italian study was higher than in the current study (64% for men, 59% for women). In contrast, a study of French blood donors with BMI <=23 kg/m2
percentiles of 42 IU/L for men and 31 IU/L for women, quite similar to the 95th
percentiles of the current study.(31
) An alternative approach was taken in a longitudinal study of a South Korean cohort of more than 90,000 men, who were followed for liver disease deaths over 8 years, a cut-off of 30 IU/L classified correctly the most men. (There were too few liver disease deaths among women to analyze.) While applicable to South Korea, such results would not necessarily apply to other countries, where different causes of liver disease predominate.
As a screening test for liver disease, a relatively low ULN for ALT may be limited practically by the large proportion of the general population that would have abnormal values, primarily due to the epidemic of overweight and obesity in the U.S. For example, applying the Italian blood donor cut-offs(26
) to the U.S. would have resulted in abnormal ALT in almost 40% of the population. Nevertheless, normal ranges for a marker of liver disease, such as ALT, must be developed from a reference population without liver disease. If the goal were to identify a high proportion of persons with HCV infection, then the ALT activity cut-off of 29 IU/L among men and 22 IU/L among women should be considered, as it classified correctly the highest proportions of persons at low liver disease risk or with hepatitis C. However, if these cut-offs were applied across the U.S., then nearly a third of the adult population would have abnormal ALT (). It remains to be established the relative utility of labeling a large minority of the population as having liver injury when the cost of evaluation would be high and its value uncertain. If direct virological testing were applied to screening for HCV, and ALT activity was not important for case identification, then the 95th
percentiles of 44 IU/L for low risk men and 32 IU/L for low risk women would be preferable. Even at these higher ALT activities, 11% of the U.S. population would be considered abnormal. We were not able to determine the causes of elevated ALT, which is the task of smaller studies that perform a thorough clinical evaluation. Nevertheless, given the phenotype of those participants with elevated ALT (), we would anticipate that the large majority would have fatty liver, primarily related to overweight and obesity.
While one can question ALT as a practical screening test for liver disease, ALT is quite helpful for the diagnosis and monitoring of liver disease. Therefore, knowledge of the distribution of ALT in persons at low risk of liver injury is important. Furthermore, it would be preferable that clinical trial requirements and drug labels that currently refer to some multiple of the ULN for exclusion or for discontinuation of therapy (such as 3 times the ULN) were instead based on a fixed ALT activity derived from populations without liver disease.(37
For both the low risk and HCV infected participants, women tended to have lower ALT than men, similar to findings in many other studies. For this reason, we presented separately results for men and women. However, the AUC was essentially the same for sexes combined as when analyzed separately. For screening, it may therefore not be absolutely necessary to use sex-specific cut-offs. Also, there was considerable racial-ethnic variation in the proportion of the total U.S. population with elevated ALT, raising the question of whether ALT ULN should also be specific to racial-ethnic groups. However, among persons at low risk for liver disease, racial-ethnic differences disappeared (), with the exception of a higher proportion of elevated ALT among Mexican-American men, suggesting that ethnic differences in the total population may be at least partially explained by racial-ethnic variation in liver disease risk factors.
A limitation of using NHANES was reliance on a single ALT measurement. Furthermore, two different autoanalyzers were used during the study period and the machine employed for the first three years generated a wider dispersion of ALT values. In addition, in clinical settings, ALT would be used in combination with other tests, rather than in isolation. Another limitation of the study was the lack of iron studies on some NHANES participants, therefore, persons with iron overload could not be excluded from the subgroup at low risk for liver disease; nor could those with rare causes of liver disease. The inclusion of a small number of participants with undetected liver diseases who had disease-related elevated ALT would have falsely lowered specificity. The choice of HCV infection as the sole reference disease could be considered a study limitation. However, HCV is an important cause of liver disease in the U.S., is transmissible and treatable, and is therefore the liver disease most important to detect. These study limitations are balanced by the benefits of a large, national, population-based sample, particularly the avoidance of ascertainment bias that can occur in clinical studies of selected patients, and the ability to generalize the results to the U.S. population. In addition, this may be the only study that has considered the effect of cut-offs on the likelihood of detection of a serious liver disease in the general population.
Ultimately, an effective laboratory test should reduce the morbidity and mortality from the diseases associated with the test. In the current study, the implications were demonstrated of the application of various cut-offs of ALT to the identification of an important liver disease, hepatitis C, and the proportion of the population that would be considered abnormal. Based on results from this national sample, a high proportion of the U.S. population would have elevated ALT at a level necessary to detect a high proportion of persons with HCV. We believe that defining the “diseased” group as persons positive for HCV or other liver disease, as done in the current study, is a useful approach. However, evaluation of ALT ULN requires further testing in multiple populations and settings.