## 3 Results and Discussion
### MicroRNA  
MicroRNAs are small nucleotide, non-coding RNAs that suppresses target mRNA translation and stability. miRNA sequencing of the sample was used to identify and quantitatively decode the entire population of microRNAs that were common between the tear and plasma samples and compare them. Initial analysis after data normalization revealed that over 400 miRNAs were identified in all the samples with 250 miRNAs commonly expressed in both the tear and plasma samples. Further analysis by filtering to a \(p\)-value cutoff of 0.01 showed 64 miRNAs to possess differential expression in the commonly expressed markers. Figure 2 shows the miRNA heatmap created by filtering the log fold change to a \(p\)-value cutoff of 0.01. Figure 2: The heatmap represents the cluster analysis of miRNA log fold change in tears and plasma (\(p<0.01\)). MicroRNAs have been demonstrated to serve as useful biomarkers for studying various diseases and conditions. Along with our results which show overlapping miRNA expression in blood plasma and tears, other studies have also shown similar overlapping effect with serum, cerebrospinal fluid, and saliva in traumatic brain injuries [18; 19]. Additionally, multiple studies have investigated the proteomic and genomic profile of tears, due to the simplicity of collection and processing, in order to identify their potential in differentiating healthy and disease states. One such study reported the miRNA expression in tears of normal and Sjogren Syndrome patients showed four upregulated and 10 down-regulated miRNA markers [20]. Exosome research has gained attention in recent years by clinicians and researchers as a source of cancer-specific biomarkers [21; 22; 23]. However, separation methods of exosomes in blood have proven inefficient as isolation of these small extracellular vesicles is severely hindered by coagulation. Efforts are underway by the International Society of Extracellular Vesicles to create a standardized protocol for isolating EVs from blood sample. Conversely, exosome isolation from tears is relatively easy compared to blood as it does not have any coagulant factors and does not require any special treatment. A recent study published from Kobe University identified miR-21 and miR-200c to be highly expressed in metastatic breast cancer patients' tear exosomes [24]. This overlap of protein and miRNA expression in plasma and tears clearly shows that tears can serve as an alternative biofluid with additional research. The ease of collection and reduced pre-processing times can be easily translated to diagnostic settings. Additionally, it opens another avenue to identify unique protein and miRNA expression that may be associated with diseased conditions that were not previously accessible in blood plasma. With the advancement of -omic technologies, we are getting closer to creating personalized, preventive medicine and having the ability to make informed decisions in treatment regimes. Research studies have been performed to identify potential biomarkers for pathological conditions beyond ocular diseases especially in Alzheimer's disease and cancer among other systemic diseases [24; 25; 26; 27; 28]. Multiple biomarkers associated with a systemic disease is becoming more common as it can achieve overall performance in sensitivity and specificity [29; 30]. The biomarker discovery information gained in similar studies will provide an optimal analytical strategy and aid future researchers to build custom microarray panels associated with a certain systemic disease. Our study serves to communicate that protein and miRNA from blood shows similar composition in tears with less processing techniques and time making it more attractive as a non-invasive source. The results reported were obtained from healthy individuals and a comparative study involving patients with a systemic disease will be conducted as part of our future study.