Lijia Liang

In situ surface-enhanced Raman scattering spectroscopy exploring molecular changes of drug-treated cancer cell nucleus.

Investigating the molecular changes of cancer cell nucleus with drugs treatment is crucial for the design of new anticancer drugs, the development of novel diagnostic strategies, and the advancement of cancer therapy efficiency. In order to better understand the action effects of drugs, accurate location and in situ acquisition of the molecular information of the cell nuclei are necessary. In this work, we report a microspectroscopic technique called dark-field and fluorescence coimaging assisted surface-enhanced Raman scattering (SERS) spectroscopy, combined with nuclear targeting nanoprobes, to in situ study Soma Gastric Cancer (SGC-7901) cell nuclei treated with two model drugs, e.g., DNA binder (Hoechst33342) and anticancer drug (doxorubicin, Dox) via spectral analysis at the molecular level. Nuclear targeting nanoprobes with an assembly structure of thiol-modified polyethylene glycol polymers (PEG) and nuclear localizing signal peptides (NLS) around gold nanorods (AuNRs) were prepared to achieve the amplified SERS signals of biomolecules in the cell nuclei. With the assistance of dark field/fluorescence imaging with simultaneous location, in situ SERS spectra in one cell nucleus were measured and analyzed to disclose the effects of Hoechst33342 and Dox on main biomolecules in the cell nuclei. The experimental results show that this method possesses great potential to investigate the targets of new anticancer drugs and the real-time monitoring of the dynamic changes of cells caused by exogenous molecules.

The use of Au@SiO2 shell-isolated nanoparticle-enhanced Raman spectroscopy for human breast cancer detection

AbstractThis study uses the powerful fingerprint features of Raman spectroscopy to distinguish different types of breast tissues including normal breast tissues (NB), fibroadenoma (FD), atypical ductal hyperplasia (ADH), ductal carcinoma in situ (DCIS), and invasive ductal carcinoma (IDC). Thin frozen tissue sections of fresh breast tissues were measured by Raman spectroscopy. Due to the inherent low sensitivity of Raman spectra, Au@SiO2 shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS) technique was utilized to provide supplementary and more informative spectral features. A total of 619 Raman spectra were acquired and compared to 654 SHINERS spectra. The maximum enhancement effect of distinct and specific bands was characterized for different tissue types. When applying the new criteria, excellent separation of FD, DCIS, and IDC was obtained for all tissue types. Most importantly, we were able to distinguish ADH from DCIS. Although only a preliminary distinction was characterized between ADH and NB, the results provided a good foundation of criteria to further discriminate ADH from NB and shed more light toward a better understanding of the mechanism of ADH formation. This is the first report to detect the premalignant (ADH and DCIS) breast tissue frozen sections and also the first report exploiting SHINERS to detect and distinguish breast tissues. The results presented in this study show that SHINERS can be applied to accurately and efficiently identify breast lesions. Further, the spectra can be acquired in a minimally invasive procedure and analyzed rapidly facilitating early and accurate diagnosis in vivo/in situ. FigureHuman breast cancer detection with Au@SiO2 SHINERS

Tracing the Therapeutic Process of Targeted Aptamer/Drug Conjugate on Cancer Cells by Surface-Enhanced Raman Scattering Spectroscopy

Targeted delivery of chemotherapeutic agents to pathology areas can improve drug efficiency and reduce serious side effects on normal regions. However, their treatment mechanism on cells or cell nuclei is still mysterious due to the lack of in situ characterization methods. In this paper, the specific diagnosis and treatment processes of a targeted antitumor agent (doxorubicin, Dox) functionalized aptamer complex (TLS11a-GC-Dox) toward HepG2 cells, a human hepatocellular carcinoma cell line, were tracked in real time by the surface-enhanced Raman scattering (SERS) spectroscopic technique and dark-field imaging with the assistance of gold nanorod-based nuclear targeted probes, which possess remarkable SERS enhancement ability, specific targeting, and excellent biological compatibility. This is the first time to explore the acting mechanism of an aptamer-based targeted drug on cell nucleus based on the spectral information on components inside the cell nucleus. The results demonstrate that this aptamer/drug conjugate has targeting and sustained-release actions and its therapeutic effect is achieved by the gradual damage of relevant proteins and DNA in nuclei. Better understanding of the mechanism of aptamer-drug conjugates acting on cancer cells is conductive to increasing cancer therapy efficiency and is also helpful for the design of highly effective drug delivery methods.

Note: Raman microspectroscopy integrated with fluorescence and dark field imaging

A Raman detection platform integrated with both fluorescence and dark field microscopes was built for in situ Raman detection with the assistance of fluorescence and dark field imaging to locate the target micro regions. Cells and organelles can be easily found via fluorescence imaging with labeling techniques. Besides, nano-sized particles could be observed and located by dark field microscopes. Therefore, comparing with the commercial Raman spectrometers, much more researches based on Raman spectroscopy could be carried out on this integrated Raman platform, especially in the fields of analyzing biological tissues and subwavelength samples.

Exploring type II microcalcifications in benign and premalignant breast lesions by shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS)

The characteristics of type II microcalcifications in fibroadenoma (FB), atypical ductal hyperplasia (ADH), and ductal carcinoma in situ (DCIS) breast tissues has been analyzed by the fingerprint features of Raman spectroscopy. Fresh breast tissues were first handled to frozen sections and then they were measured by normal Raman spectroscopy. Due to inherently low sensitivity of Raman scattering, Au@SiO2 shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS) technique was utilized. A total number of 71 Raman spectra and 70 SHINERS spectra were obtained from the microcalcifications in benign and premalignant breast tissues. Principal component analysis (PCA) was used to distinguish the type II microcalcifications between these tissues. This is the first time to detect type II microcalcifications in premalignant (ADH and DCIS) breast tissue frozen sections, and also the first time SHINERS has been utilized for breast cancer detection. Conclusions demonstrated in this paper confirm that SHINERS has great potentials to be applied to the identification of breast lesions as an auxiliary method to mammography in the early diagnosis of breast cancer.

Tracing sialoglycans on cell membrane via surface-enhanced Raman scattering spectroscopy with a phenylboronic acid-based nanosensor in molecular recognition

Sialoglycan expression is critical for assessing various diseases progression. Especially, its abnormal levels are commonly believed to be associated with tumor and metastatic cancer types. While, complicated structures, multiple types and dynamic distributions make it challenging for in situ investigating sialoglycans at the physiological status. Herein, we developed a 4-mercaptophenylboronic acid (MPBA)-based surface-enhanced Raman scattering (SERS) nanosensor to in situ study sialoglycan levels and dynamic expression processes of different cell types based on molecular recognition between phenylboronic acid and sialoglycans at physiological condition. This nanosensor is designed by the MPBA decorated silver nanoparticle (AgNP), which is unique and multifunctional because of its three-in-one role involving the Raman signal enhancer (AgNP), the sensing reporter of MPBA and the target receptor based on the recognition of phenylboronic acid and sialoglycans. When this nanosensor binds to sialoglycans, the molecular vibrational modes of MPBA will change, which can be traced by ultrasensitive SERS technique. The superiority of this study is that we built the relation between the spectral changes of MPBA (relative intensities) in molecular recognition with the sialoglycan dynamic expression of cells. We believe that our SERS strategy could be further extended to explore crucial physiological processes and significant biological system that glycans are involved in.

Glucose-bridged silver nanoparticle assemblies for highly sensitive molecular recognition of sialic acid on cancer cells via surface-enhanced raman scattering spectroscopy

The expression levels of glycans on the surfaces of cancer and normal cells show different, however, this difference is not noticeable enough to distinguish them directly. So, herein, based on the targeted molecular recognition of the glycans on cell surfaces by 4-mercaptophenyl boronic acid (MPBA), a novel surface-enhanced Raman scattering (SERS) nanoprobe (glucose-MPBA@AgNPs) was prepared by inducing controllable assembly of MPBA decorated Ag nanoparticles (MPBA@AgNPs) in a certain level via the bridge of glucose to amplify such a limited difference in SERS measurements. On the basis of the aggregation-induced 3D SERS hot spot effect, this multi-particle nanoprobe possesses over 10 times stronger SERS enhancement ability than the individual MPBA@AgNPs. As the different sialic acid (SA) expression on the surfaces of cancer and normal cells led to the different accumulation of glucose-MPBA@AgNPs, the results we obtained (mean intensities recorded from five cells) indicate the SA amounts on two kinds of cells can provide 5-7 times signal contrast grade in SERS band intensities (P < 0.001). Compared with the monodispersed nanoprobe, our developed nanoprobe amplifies the SA expression difference on cell surfaces and supports high sensitivity for cancer cell recognition, which might be useful in providing highly effective recognition of the edges of tumor tissues in clinic field.

Organelle-Targeting Gold Nanorods for Macromolecular Profiling of Subcellular Organelles and Enhanced Cancer Cell Killing

Subcellular organelles, for example, nucleus, mitochondria, and lysosome, are the vital organelles with responsibilities that maintain cell operation and metabolism. Owing to their roles in energy production and programmed cell death, these organelles have become prime therapeutic targets in different diseases and states. In this study, biocompatible, organelle-targeting nanoprobes were developed by modifying gold nanorods (AuNRs) with specific targeting peptides. These nanoprobes were employed to directly profile subcellular biomolecules and vital organelles by surface-enhanced Raman scattering (SERS) spectroscopy. Macromolecular spectral profiles of subcellular organelles were achieved and compared. Further, these organelle-targeting AuNRs were used for the photothermal treatment of cancer cells (HepG2, HeLa, and MCF-7 cell lines). The cell viability assays show that the nucleus- and mitochondria-targeting AuNRs provide higher photothermal efficiencies under an 808 nm laser relative to the lysosome-targeting ones. This study makes critical insights into the spectral profiles of subcellular organelles and also inspires people in the development of high-efficacy cancer therapeutic strategies by subcellular organelle-targeting drugs.

Abstract P2-05-15: Exploring type II microcalcifications in benign, premalignant and malignant breast lesions by shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS)

Background: Type II microcalcifications, most often seen in proliferative lesions, including both benign and malignant pathologies. But whether the emergence of type II microcalcifications associated with cell canceration is still not clear now. Raman spectroscopy is a powerful, non-invasive analytical tool which can provide detailed and meaningful information about biochemical composition of tissues at molecular level. Our aim was to find the differences and relationships of type II microcalcifications between fibroadenoma, ADH tissues, and DCIS, IDC in breast based on their various biochemical characteristics by Raman spectroscopy. Methods: The frozen sections were collected from 15 patients (all female; ages 25-57) who underwent surgical resection or mammotome biopsy at the Department of Breast Surgery, the First Hospital of Jilin University. After operation the samples were immediately frozen at -20∼-25° and two contiguous sections (6 µm thickness) were cut from a sample by freezing microtome. One was stained with haematoxylin and eosin for routine histopathological analysis and found the microcalcification locations by three expert breast pathologists; the same position of the other section was detected by Microscopic confocal Raman spectrometer (HORIBA JY Lab800, 633nm) with its mirror image (the HE San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P2-05-15.

In situ, accurate, surface-enhanced Raman scattering detection of cancer cell nucleus with synchronous location by an alkyne-labeled biomolecular probe

AbstractA surface-enhanced Raman scattering (SERS) method for in situ detection and analysis of the intranuclear biomolecular information of a cell has been developed based on a small, biocompatible, nuclear-targeting alkyne-tagged deoxyribonucleic acid (DNA) probe (5-ethynyl-2′-deoxyuridine, EDU) that can specially accumulate in the cell nucleus during DNA replications to precisely locate the nuclear region without disturbance in cell biological activities and functions. Since the specific alkyne group shows a Raman peak in the Raman-silent region of cells, it is an interior label to visualize the nuclear location synchronously in real time when measuring the SERS spectra of a cell. Because no fluorescent-labeled dyes were used for locating cell nuclei, this method is simple, nondestructive, non- photobleaching, and valuable for the in situ exploration of vital physiological processes with DNA participation in cell organelles. Graphical abstractA universal strategy was developed to accurately locate the nuclear region and obtain precise molecular information of cell nuclei by SERS