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Jeffrey Suhalim, Yilu Xie, Jeremiah Tao, Donald Brown, James Jester, Eric Potma; Characterization of lipids in human meibomian gland using coherent Raman scattering microscopy. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1486. doi: https://doi.org/.
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While the association between meibomian gland dysfunction (MGD) and dry eye disease is well known, it is unclear how meibum lipid changes during the progression of the disease. Several studies have pointed out the importance of meibum composition in MGD; however, these studies primarily focused on studying lipids expressed from the meibomian gland. Hence, no information on compositional changes pertaining to the structure of the gland is available. In this study, we demonstrate the application of coherent Raman scattering (CRS) microscopy to characterize the compositional changes within the human meibomian gland.
Eyelid tissues from blepharoplasty patients were fixed in PBS with 2% PFA. Cryostat tissue sections were placed on a microscope slide, then imaged with a multimodal CRS microscope as described in . The protein-to-lipid ratio (P/L) was obtained by tuning the laser beams to probe the vibrational signatures of protein and lipid. Specifically, the vibrations of methyl and amide-I were utilized to measure the contribution from protein, whereas the symmetric stretch and bending modes of methylene were used to probe lipid. In addition, the conjugated carbon-carbon vibration was monitored as a marker for carotenoid-like molecules. No labeling or staining was required in this study.
In general, CRS analysis of the meibomian gland starting at the acinus and moving to the central duct showed a decreasing P/L ratio. Our preliminary results also showed differences in the P/L ratio between acini of the same gland. Concomitantly, Raman vibration of conjugated carbon-carbon bond was detected, hinting the presence of carotenoid-like molecules in the gland, as previously observed by Oshima et al. for expressed meibum .
CRS imaging enables compositional analysis in context of the native structure of the gland, complementing other studies. Our findings suggest that there is a maturational change in meibum with decreasing protein content as lipid moves toward the gland orifice and that there may be varying degrees of acinar activity within the gland.  Suhalim, J.L., et al., Biophysical Journal, 2012. 102(8): p. 1988-95.  Oshima, Y., et al., Current Eye Research, 2009. 34(10): p. 824-835.
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