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Assessment of cell line models of primary human cells by Raman spectral phenotyping.

ABSTRACT: Researchers have previously questioned the suitability of cell lines as models for primary cells. In this study, we used Raman microspectroscopy to characterize live A549 cells from a unique molecular biochemical perspective to shed light on their suitability as a model for primary human pulmonary alveolar type II (ATII) cells. We also investigated a recently developed transduced type I (TT1) cell line as a model for alveolar type I (ATI) cells. Single-cell Raman spectra provide unique biomolecular fingerprints that can be used to characterize cellular phenotypes. A multivariate statistical analysis of Raman spectra indicated that the spectra of A549 and TT1 cells are characterized by significantly lower phospholipid content compared to ATII and ATI spectra because their cytoplasm contains fewer surfactant lamellar bodies. Furthermore, we found that A549 spectra are statistically more similar to ATI spectra than to ATII spectra. The spectral variation permitted phenotypic classification of cells based on Raman spectral signatures with >99% accuracy. These results suggest that A549 cells are not a good model for ATII cells, but TT1 cells do provide a reasonable model for ATI cells. The findings have far-reaching implications for the assessment of cell lines as suitable primary cellular models in live cultures.

SUBMITTER: Swain RJ

PROVIDER: S-EPMC2856139 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Assessment of cell line models of primary human cells by Raman spectral phenotyping.

Swain Robin J RJ Kemp Sarah J SJ Goldstraw Peter P Tetley Teresa D TD Stevens Molly M MM

Biophysical journal 20100401 8

Researchers have previously questioned the suitability of cell lines as models for primary cells. In this study, we used Raman microspectroscopy to characterize live A549 cells from a unique molecular biochemical perspective to shed light on their suitability as a model for primary human pulmonary alveolar type II (ATII) cells. We also investigated a recently developed transduced type I (TT1) cell line as a model for alveolar type I (ATI) cells. Single-cell Raman spectra provide unique biomolecu ...[more]

PMID: 20409492

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Project description:Human endogenous retroviruses (HERVs) integrated into the human genome millions of years ago, and their expression has been described both in healthy tissues and in pathology. In the present study, we study the differential expression of multiple HERV families in the presence of human immunodeficiency virus (HIV) infection in the peripheral blood mononuclear cells (PBMCs) of HIV-positive individuals, in HIV-infected cell lines, and in vitro HIV-infected macrophages/monocytes of healthy donors, utilizing publicly available data sets for secondary analysis using a pipeline appropriate for HERV transcription detection and comparison. We recognized significantly decreased expression of HERV-H in treatment-naïve HIV patients with viremia and increased expression of multiple HERV families in the PBMCs of HIV-infected individuals under combination antiretroviral therapy compared to healthy donors, in in vitro HIV-infected Stanford University pediatric T (SUP-T1) cells at 24 hours after HIV infection compared to non-infected SUP-T1 cells, and in in vitro HIV-infected MT4 T cells at 3 days after HIV infection compared to non-infected MT4 T cells. We recognized significantly increased expression of HERV-W, after HIV infection in monocyte-derived macrophage cell culture; no difference was found in HERV expression after HIV infection of monocyte cultures. We found a significantly decreased expression of HERV-K (HML-6) in donor-derived macrophage cultures at 36 hours after HIV infection, which does not persist at 6 days after infection. HIV infection modifies the expression of multiple HERV families in the PBMCs of HIV patients and in T cell-derived cultures, while the effect of HIV infection on HERV expression appears to be more restricted in cells of monocytic origin.IMPORTANCEIn this work, we demonstrated that human immunodeficiency virus (HIV) infection leads to the modification of the human endogenous retrovirus (HERV) expression. Differential expression of multiple HERVs was found in peripheral blood mononuclear cells derived from HIV-infected patients compared to healthy donors and HIV-infected T cell cultures compared to non-infected. The effect of HIV presence on HERV expression appears to be more restricted in cells of monocytic origin, as only deregulation of HERV-W and HERV-K (HML-6) was found in these cell cultures after their infection with HIV. Multiple factors contribute to this aberrant HERV expression, and its levels appear to be modified in a time-dependent manner. Further studies and the development of optimized in vitro protocols are warranted to elucidate the interactions between HIV and HERVs in detail.

Project description:IntroductionEnvironmental microparticle is becoming a global pollutant and the entire population is increasingly exposed to the microparticles from artificial materials. The accumulation of microparticles including microplastics and its subsequent effects need to be investigated timely to keep sustainable development of human society.ObjectivesThis study aimed to explore the accumulation of environmental particles in thrombus, the pathological structure in the blood circulation system.MethodsPatients receiving cardiovascular surgical operations were screened and twenty-six thrombi were collected, digested and filtered. Non-soluble microparticles were enriched on the filter membrane and then were analyzed and identified with Raman Spectrometer. The associations of particle status (presence or absence) or particle number in the thrombus and clinical indicators were examined. One strict quality control-particle detection system was designed to eliminate environmental contaminations.ResultsAmong twenty-six thrombi, sixteen contained eighty-seven identified particles ranging from 2.1 to 26.0 μm in size. The number of microparticles in each thrombus ranged from one to fifteen with the median reaching five. All the particles found in thrombi were irregularly block-shaped. Totally, twenty-one phthalocyanine particles, one Hostasol-Green particle, and one low-density polyethylene microplastic, which were from synthetic materials, were identified in thrombi. The rest microparticles included iron compounds and metallic oxides. After the adjustment for potential confounders, a significantly positive association between microparticle number and blood platelet levels was detected (P < 0.01).ConclusionThis study provides the first photograph and Raman spectrum evidence of microparticles in thrombi. A large number of non-soluble particles including synthetic material microparticles could accumulate in arteries, suggesting that the risk of microparticle exposure was under-estimated and the re-evaluation of its health effects is urgently needed. There will be a series of reports on assessing the health effects of microparticle exposure in humans in the future and this research provided clues for the subsequent research.

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Assessment of cell line models of primary human cells by Raman spectral phenotyping.

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Assessment of cell line models of primary human cells by Raman spectral phenotyping.

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