Nanoparticle Uptake in the Aging and Oncogenic Drosophila Midgut Measured with Surface-Enhanced Raman Spectroscopy

Abstract

Colorectal cancer remains a major global health concern. Colonoscopy, the gold-standard colorectal cancer diagnostic, relies on the visual detection of lesions and necessitates invasive biopsies for confirmation. Alternative diagnostic methods, based on nanomedicine, can facilitate early detection of malignancies. Here, we examine the uptake of surface-enhanced Raman scattering nanoparticles (SERS NPs) as a marker for intestinal tumor detection and imaging using an established Drosophila melanogaster model for gut disease. Young and old Oregon-R and w¹¹¹⁸ flies were orally administered SERS NPs and scanned without and upon gut lumen clearance to assess nanoparticle retention as a function of aging. Neither young nor old flies showed significant NP retention in their body after gut lumen clearance. Moreover, tumorigenic flies of the esg-Gal4/UAS-Ras^V12 genotype were tested for SERS NP retention 2, 4 and 6 days after Ras^V12 oncogene induction in their midgut progenitor cells. Tumorigenic flies showed a statistically significant NP retention signal at 2 days, well before midgut epithelium impairment. The signal was then visualized in scans of dissected guts revealing areas of NP uptake in the posterior midgut region of high stem cell activity.

Keywords: Drosophila melanogaster; SERS; gut disease; nanoparticle uptake; surface-enhanced Raman scattering.

Figure 1

SERS intensity at various nanoparticle relative concentrations. (a) The SERS nanoparticles demonstrate a characteristic spectrum that diminishes as they are serially diluted. The characteristic peaks appearing at 1203, 520 and 557 cm⁻¹ are highlighted. (b) A regression model was applied to the intensity of the 1203 cm⁻¹ peak, showing a logarithmic response of the nanoparticles as a function of concentration.

Figure 2

Examples of (a) OR and (b) w¹¹¹⁸ Smurf females with light diffused blue color (left) and non-Smurf (right).

Figure 3

SERS NP uptake by young and old females. (a) Raman spectra from young and old flies, w¹¹¹⁸ and OR, with and without clearance. The spectra demonstrate many Raman peaks intrinsic to the fly body. Spectra of individual flies are shown in grey and the group average in blue. The shaded bands indicate the areas of the peaks specific to the SERS NPs. (b) Regression (nn-LS) scores indicate moderate SERS signals in the flies fed with SERS NPs, with no statistically significant differences, as indicated by the p-values. Dots are individual fly measurements, the bars show the mean, and the whiskers the standard deviation. For each condition, 5 to 7 flies were scanned.

Figure 4

SERS NP uptake by mutant flies. (a) Raman spectra for esg^ts-Ras^V12 flies on day 2, 4 and 6 of induction of Ras^V12 oncogene in their midgut progenitors. Spectra of individual flies are shown in grey and the group average in blue. The shaded bands indicate the areas of the peaks specific to the SERS NPs. (b) Mean nn-LS scores and p-values as calculated for the different groups, show a small but statistically significant uptake for the 2-day condition, which decreases with disease progression. For each experimental condition, 10 flies were scanned.

Figure 5

Raman maps of excised fly midguts. (a) Photograph of excised guts for NP-fed esg^ts-Ras^V12 flies. The superimposed colormap represents the nn-LS scores obtained from the different areas. Scale bar: 10 mm. (b) Detailed view from the sample areas indicated by boxes in (a). The optical microscopy images are shown with and without the colormap superimposed, for comparison. The areas with high nn-LS scores in their posterior midgut region display a reddish metallic texture, indicative of the presence of SERS NPs, particularly evident in (bii). Scale bar: 1 mm. (c) Representative Raman spectra corresponding to the pixels from panel (b) indicated with red crosses (×). Area (bii) features an exceptionally bright signal, whereas other areas display the characteristic peaks at lower intensities. (d–f) Similar to panels (a–c) but from control flies not fed with nanoparticles. Regression signals and Raman spectra have no indication of SERS nanoparticles, as expected.