Riboflavin as a Source of Autofluorescence in Eisenia fetida Coelomocytes

Abstract

Immunocompetent cells of earthworms (coelomocytes) contain adherent amoebocytes and large eleocytes (chloragocytes); the latter are filled with numerous granules. We have previously shown that eleocytes of several (but not all) earthworm species exhibit strong autofluorescence detectable by fluorescent microscopy and flow cytometry. In the present article, the molecular origin of eleocytes autofluorescence was elucidated in coelomocytes expelled via dorsal pores in the integument of Eisenia fetida subjected to electric shock (1 min at 4.5 V). Spectrofluorometry (excitation and emission spectra and fluorescence lifetime), together with HPLC analysis of coelomocyte suspensions and supernatants, indicated that riboflavin but not FMN (flavin mononucleotide) or FAD (flavin-adenine dinucleotide) is the main fluorophore responsible for eleocyte fluorescence in this species. Additionally, lipofuscins are suspected to participate in this phenomenon.

Rapid diagnosis and intraoperative margin assessment of human lung cancer with fluorescence lifetime imaging microscopy

“…It seems the lifetimes of NADH and FAD both decreased in cancerous lung tissues. Since the fluorescence of protein-bound FAD is very weak [19] the lifetime of FAD in this study was mainly contributed by free FAD whose lifetime was reported in the range of 2.28–3.13 ns [20], [21]. The decreasing of FAD lifetime was caused by cellular micro-environment change in cancerous lung tissues.…”

Immune system participates in brain regeneration and restoration of reproduction in the earthworm Dendrobaena veneta

“…In an earlier publication, Moment (1974) implicated a stimulatory regeneration role for the so-called 'yellow pigment' derived from a certain coelomocyte cohort. More recently, the 'yellow pigmentation' of coelomocytes has been ascribed to stored riboflavin (vitamin B2) within the characteristic granulelike organelles ('chloragosomes') of attached chloragocytes and the chloragocyte-derived coelomocytes called 'eleocytes' that are prevalent in the coelomic fluid of several earthworm species, including D. veneta (Koziol et al, 2006;Plytycz and Morgan, 2011;Plytycz et al, 2006;Rorat et al, 2014). In the context, it is particularly noteworthy that Samuel et al (2012) reported that the injection of riboflavin promoted blastema formation during segment regeneration in the epigeic 'African nightcrawler' earthworm species Eudrilus eugeniae.…”

Fluorescence Lifetime Measurements and Biological Imaging

“…25). FAD emits at a longer wavelength than NAD(P)H, with only small overlap, allowing for selecting imaging conditions specifically for FAD without spectral contamination from NAD(P)H. In contrast to NAD(P)H, only the non-bound form of FAD is fluorescent and it has a lifetime of 2.3-2.9 ns 247,248. Protein binding leads to a significant decrease of the lifetime of FAD (<0.1 ns) 247,249.…”