HPV Infection Significantly Accelerates Glycogen Metabolism in Cervical Cells with Large Nuclei: Raman Microscopic Study with Subcellular Resolution

Abstract

Using Raman microscopy, we investigated epithelial cervical cells collected from 96 women with squamous cell carcinoma (SCC) or belonging to groups I, IIa, IIID-1 and IIID-2 according to Munich III classification (IIID-1 and IIID-2 corresponding to Bethesda LSIL and HSIL groups, respectively). All women were tested for human papillomavirus (HPV) infection using PCR. Subcellular resolution of Raman microscopy enabled to understand phenotypic differences in a heterogeneous population of cervical cells in the following groups: I/HPV−, IIa/HPV−, IIa/HPV−, LSIL/HPV−, LSIL/HPV+, HSIL/HPV−, HSIL/HPV+ and cancer cells (SCC/HPV+). We showed for the first time that the glycogen content in the cytoplasm decreased with the nucleus size of cervical cells in all studied groups apart from the cancer group. For the subpopulation of large-nucleus cells HPV infection resulted in considerable glycogen depletion compared to HPV negative cells in IIa, LSIL (for both statistical significance, ca. 45%) and HSIL (trend, 37%) groups. We hypothesize that accelerated glycogenolysis in large-nucleus cells may be associated with the increased protein metabolism for HPV positive cells. Our work underlines unique capabilities of Raman microscopy in single cell studies and demonstrate potential of Raman-based methods in HPV diagnostics.

Dual Switch in Lipid Metabolism in Cervical Epithelial Cells during Dysplasia Development Observed Using Raman Microscopy and Molecular Methods

“…Complete statistical data on the lipid content in the cytoplasm have been included in the Supplementary Materials ( Table S4 ). Additionally, the glycogen level anti-correlates with the lipid content in the cytoplasm of cervical epithelial cells, in particular, it is the lowest in the HSIL+ and SCC+ groups ( Figure 2 B), in agreement with our previous results [ 38 ]. The negative correlation of both factors is statistically significant in the N and HSIL groups ( Figure 2 C).…”

“…When comparing the averaged signatures of cytoplasm and LDs, it is clear that bands due to lipids are present only in LDs ( Figure 1 B). Moreover, both of these classes also contain glycogen bands (481, 577, 858, 937, 1083, 1129, and 1340 cm −1 ) [ 38 ] but they are not present in all cells and their intensity significantly vary between cells. In the LDs of the studied cells, these components are intertwined and cannot be discriminated from each other with the use of the CA.…”

“…In the LDs of the studied cells, these components are intertwined and cannot be discriminated from each other with the use of the CA. As shown in our previous paper, the level of glycogen in cervical epithelial cells depends on the degree of dysplasia and HPVhr infection, in particular, is very low in large-nucleus HPVhr+ cells vs. HPVhr− cells, which may be associated with the increased protein metabolism [ 38 ]. The described Raman bands and their assignments have been included in the Supplementary Materials ( Table S3 ).…”

“…Raman spectroscopy is a label-free and non-destructive method that is used to study the chemical composition of samples, including biological ones [ 37 ]. Cervical cancer cells have been studied with the use of Raman microscopy [ 38 , 39 , 40 , 41 ] and infrared spectroscopy [ 42 ] but they did not concentrate on changes in the lipid profile of cells.…”

Vulvar Squamous Cell Carcinoma in a Patient with AIDS: A Case Study

“…In general, there is wide access to HPV immunization through vaccination [22], and the fact that currently there are no recommended screening strategies for vulvar cancer, HPV vaccination may be the only effective way for prevention of this malignancy [23]. Our patient was not aware of her HPV vaccination status which serves as a lesson emphasizing the importance of patient education about the importance and impact of HPV immunization.…”