Lipid Droplet Proteins in Acne Skin: A Sound Target for the Maintenance of Low Comedogenic Sebum and Acne-Prone Skin Health

Abstract

In adipocytes and sebocytes, lipid droplet proteins control the storage of lipids in organized droplets and their release on demand. The contribution of lipid droplet proteins to the pathogenesis of acne is plausible because they control the levels of comedogenic free fatty acids. The expression of two lipid droplet proteins, CIDEA and PLIN2, was analyzed in the skin of patients with acne by immunohistochemistry and western blotting. The design of clinical protocols allowed correlating the expression of CIDEA and PLIN2 with both comedogenesis and the release of free fatty acids. Both proteins were detected by immunohistochemistry in the sebaceous glands of patients with acne, with a disturbed expression pattern of PLIN2 compared with that in the controls. Higher levels of PLIN2 and CIDEA, as detected by western blotting in the infundibulum, significantly correlated with lower ongoing comedogenesis over 48 weeks of Silybum marianum fruit extract application. Accordingly, free fatty acid release from sebum triglycerides was significantly decreased, as shown with two distinct methods. The data are consistent with the expected role of PLIN2 and CIDEA in the prevention of comedogenic free fatty acid release. Modulation of PLIN2 and CIDEA expression appears as a sound target for the maintenance of low comedogenic sebum and acne-prone skin health.

Keywords: FFA, free fatty acid; LD, lipid droplet; SMFE, Silybum marianum fruit extract; TG, triglyceride.

Figure 1

Summary diagram of data. PLIN2 and CIDEA are anchored on the phospholipid layer of lipid droplets. The lipid core contains mainly triglycerides, FFAs, S, as well as CWEs (Kory et al., 2016; Olzmann and Carvalho, 2019). During comedogenesis, the expression of PLIN2 and CIDEA is decreased, and a triglyceride lipase is activated, releasing FFAs from triglycerides. This process is partly reversed by topical application of SMFE. CWE, cholesterol and wax ester; FFA, free fatty acid; S, squalene; SMFE, Silybum marianum fruit extract.

Figure 2

Correlation between microcomedone index and lipid droplet proteins in cohort 1 (CSSB biopsies). (a) PLIN2 and CIDEA are expressed in a nonmetric multidimensional scaling of 50 CSSB biopsies from 23 patients on the basis of the value of the MCI in decreasing order; for each sample, the sum of corresponding MCI value and the expression values for CIDEA and PLIN2 gives 100%, and the bars represent the respective percentages for MCI, CIDEA, and PLIN2 (Fontao et al., 2020). (b) The means ± SD of CIDEA and PLIN2 expressions are determined for 50 CSSB biopsies from 23 patients, with MCI values above and below 15. Positive control for (c) PLIN2 and (d) CIDEA expressions were analyzed in human Sebo cultures, human hepatocyte cell line HepG2, mouse liver extract (liver), and human CSSB extracts from three samples. For each of the gels in c and d, after the transfer to nitrocellulose membranes, the membranes were spliced to incubate the respective pieces with (c) PLIN2 and GAPDH antibodies or (d) CIDEA and GAPDH antibodies; white lines indicate where the membranes were spliced. (e) This image shows CIDEA and GAPDH bands from a gel to explain how the densitometric assessment was performed for all protein bands of interest; as for the gels in c and d, the nitrocellulose membrane was spliced to incubate each part with CIDEA or GAPDH antibodies; when comparing CIDEA and GAPDH (housekeeping protein), the background of the gel—corresponding to the rectangle in the middle of the image—is subtracted, and the density of the protein bands are calculated by subtracting their mean gray values from that of the background. The ratio between the LD protein and GAPDH gives the expression value of the LD protein. This ratio is then multiplied by 100 when creating the graphs shown in Figure 1a and b for clarity. Statistical significance in b is set as ∗∗P < 0.01. CSSB, cyanoacrylate skin surface biopsy; LD, lipid droplet; MCI, microcomedone index; Sebo, sebocyte.

Figure 3

Assessment of acne in cohort 2 (skin surface lipids). (a) The lipid composition of sebum was analyzed by infrared spectroscopy to determine the FFAs-to-TGs ratio. (b) Noninflammatory acne lesions (open and closed comedones). (c) Acne lesion count as described by Lucky et al. (1996). (d) Acne assessment using the Investigator Global Assessment of Acne. (e–g) Data from FFAs and TGs for comedones collected from nose wings. FFA and TG were analyzed by gas chromatography‒mass spectrometry. Results are expressed as means ± SD; statistical significance was set at ∗P < 0.05 and ∗∗P < 0.01. FFA, free fatty acid; TG, triglyceride; wk, week.

Figure 4

Immunohistochemical labelling of PLIN2 and CIDEA in human skin of nonacne controls (cohort 3). As previously reported, both proteins were detected in the skin of humans without acne (Dahlhoff et al., 2015; Schneider et al., 2016). The distribution over the sebaceous gland as well as the cellular pattern were similar but not identical for (a) PLIN2 and (b) CIDEA. Bar = 100 μm in a or 200 μm in b. (c) Human skin biopsy (used for negative control); absence of staining using only the second antibody (172-1019, 1:500; Bio-Rad Laboratories, Hercules, CA). Bar = 500 μm. (d) Mouse heart (used for positive control of CIDEA staining [PA5-29654, 1:200; Thermo Fisher Scientific, Waltham, MA]). Bar = 50 μm. (e) Sebocyte cultures (Zen-Bio, Research Triangle Park, NC) (used for positive control of PLIN2 staining [SAB4200452, 1:200; Sigma-Aldrich, St. Louis, MO]).

Figure 5

Immunohistochemical labeling of PLIN2 in human skin of patients with acne and nonacne controls (cohort 3). (a) A general view and (b) a close view of patient 1 highest expression field. (c‒i) Close views of the highest expression field in the seven other patients with acne. (j) A general view of one control. (k‒o) Close views of the highest expression field of the controls. Bar = 500 μm in a, e, h, i, and k. Bar = 200 μm in b, d, f, and g. Bar = 250 μm in c, m, and o. Bar = 1,000 μm in l and n. Bar = 1,000 μm in j.