Identification of Biomarker Hyaluronan on Colon Cancer Extracellular Vesicles Using Correlative AFM and Spectroscopy

Abstract

Extracellular vesicles (EVs), naturally occurring nanosized vesicles secreted from cells, are essential for intercellular communication. They carry unique biomolecules on the surface or interior that are of great interest as biomarkers for various pathological conditions such as cancer. In this work, we use highresolution atomic force microscopy (AFM) and spectroscopy (AFS) techniques to demonstrate differences between EVs derived from colon cancer cells and colon epithelial cells at the singlevesicle level. We observe that EV populations are significantly increased in the cancer cell media compared to the normal cell EVs. We show that both EVs display an EV marker, CD9, while EVs derived from the cancer cells are slightly higher in density. Hyaluronan (HA) is a nonsulfated glycosaminoglycan linked to malignant tumor growth according to recent reports. Interestingly, at the single-vesicle level, colon cancer EVs exhibit significantly increased HA surface densities compared to the normal EVs. Spectroscopic measurements such as Fourier transform infrared (FT-IR), circular dichroism (CD), and Raman spectroscopy unequivocally support the AFM and AFS measurements. To our knowledge, it represents the first report of detecting HA-coated EVs as a potential colon cancer biomarker. Taken together, this sensitive approach will be useful in identifying biomarkers in the early stages of detection and evaluation of cancer.

Cancer-associated fibroblasts derived extracellular vesicles promote angiogenesis of colorectal adenocarcinoma cells through miR-135b-5p/FOXO1 axis

“… 23 Consistently, EVs derived from cancer cells are essential for inducing malignant behaviors in COAD cells. 24 Briefly, our findings elicited that CAFs-EVs induced COAD cell-mediated HUVECs malignant biological behaviors and angiogenesis.…”

The biology, function, and applications of exosomes in cancer

“…Paul et al. 172 used high-resolution atomic force microscopy (AFM) and spectroscopy (AFS) techniques to demonstrate differences between EVs derived from colon cancer cells and colon epithelial cells at the single-vesicle level. Compared with normal cells, the EVs of colon cancer cells have a significantly increased surface density of hyaluronic acid (HA).…”

FT-IR Spectral Signature of Sensitive and Multidrug-Resistant Osteosarcoma Cell-Derived Extracellular Nanovesicles

“…In particular, all EVs curves showed the amide I absorption band located at ≈1649 cm −1 , associated with the C=O stretching mode of the peptide bond, and the amide II absorption located at ≈1455 cm −1 , which was primarily ascribable to N-H vibrations of peptide groups. A band is also assessed at 1407 cm −1 , which can be attributed to symmetric and asymmetric vibration of COO− [ 23 ].…”

“…In fact, this technique is able to detect subtle changes in the molecular structure of nucleic acids, lipids, proteins, and carbohydrates in biological samples, thus defining a specific biomolecule fingerprinting. FT-IR spectroscopy has been utilized to identify and distinguish subpopulations of EVs derived from melanoma cells with different malignant grades [ 28 ], and it has been fruitfully applied to reveal differences at the single vesicle level between EVs derived from colon normal epithelial cells and colon cancer cells [ 23 ]. Moreover, FT-IR spectra were used to characterize EVs isolated from biological fluids and consistently different spectral signatures were identified for salivary cancer and healthy individual derived-EVs [ 29 ], for blood from prostate cancer and healthy patient derived-EVs [ 30 ], and for blood derived-EVs isolated from Alzheimer’s disease affected and control subjects [ 31 ].…”