# Age- and Sex-Adjusted Reference Intervals in Tear Cytokine Levels in Healthy Subjects
## 4 Discussion
An increase of IL-8 levels with age has recently been described in cerebrospinal fluid as well, but, in this case, the evolution is practically linear [36]. The role of IL-7 in immunosenescence, the aging of the immune system, has been known for long [41]. In this work, we have found a cyclic evolution of IL-7 tear levels with higher levels around 30 years old and increasing again after 70 years old. This result disagrees with the findings by [PERSON] et al. [12] which found an increase of IL-7 in middle age people (41-60 years old) with respect to young (18-40) and old (\(<\)60 years old). However, the observed behavior for IL-7 tear levels in our study could be consistentwith the findings of [PERSON] et al. [42], that evaluated plasma IL-7 levels in three age groups: young (aged 20-45), middle-aged (aged 58-62), and centenarians (range 98-100 years). Despite the fact that they found no significant differences between the age groups, the evolution pattern of IL-7 levels was similar to ours. There is more confusion regarding VEGF evolution. In serum, for example, both an increase [43] and a decrease [44] of VEGF levels with aging have been found. In plasma samples of patients from 43 to 80 years old with stable coronary artery disease a significant negative correlation was observed [45]; in contrast, recently, [PERSON] et al. [46] detected a positive correlation in cerebrospinal fluid from subjects aged from 40 years. Other authors such as [PERSON] et al. [47] found no association between the concentration of VEGF in synovial fluid and age. In our case, the relation between VEGF tear level and age can be described as an inverted-U shape curve, with maximum levels between 30-60 years old. Our results regarding the age-dependence in tear cytokine levels are also in agreement with the findings by [PERSON] et al. [16] for tear proteome in which the majority of the identified proteins in their study had the most notable increase/decrease in subjects aged 60 or more. Having age-dependent RI of tear cytokine levels can be very useful for the diagnosis of ocular disorders. Several very common eye problems are related to the phenomenon of inflammaging. For example, an association between age and DED has been assumed for a long time, and recently, [PERSON] et al. [48] have directly related inflammaging to a progressive impairment of ocular surface system function. An ocular disease in which age is an important risk factor is age-related macular degeneration, affecting people aged 65 years and older. Several inflammatory gene polymorphisms have been linked to this disorder indicating an important role of inflammation and immune-mediated processes, and, therefore, directly related to inflammaging term [49]. Finally, low-grade inflammation has been described as a causal factor in the pathogenesis of glaucoma [50]. Although glaucoma can affect people of all ages, is most common in adults in their 70s and 80s, so it is accepted that glaucoma risk increases with age [51]. Sex also has an influence on tear levels of some of the analyzed cytokines. Particularly, Fractalkine tear levels were higher in males compared to females and IL-7, IL-8, and IP-10 higher in females compared to males; whereas, EGF, IL-1 RA, MCP-1, and VEGF levels were not affected by sex. Some other studies have also shown sex-dependence in cytokine levels in other fluids. For instance, [PERSON] et al. (2006) [42], found a sex-dependence for IL-7 serum levels, with higher levels in centenarian females than in males, in agreement with our results in tears. Moreover, the study by [PERSON] et al. (2019) [16] revealed that protein-age correlations in tear film were, in most cases, more significant in males, although mammaglobin-A (SCGB2A2) protein had a higher correlation coefficient for females. Changes in sex hormones have been suggested as connected with these differences. In addition, very recently, stronger relationships between age and IL-6 and high-sensitivity C-reactive protein plasma levels have been found in females compared to males [52].