In situSynthesis of Fluorescent Gold Nanoclusters by Nontumorigenic Microglial Cells

Abstract

To date, the directed in situ synthesis of fluorescent gold nanoclusters (AuNCs) has only been demonstrated in cancerous cells, with the theorized synthesis mechanism prohibiting AuNC formation in nontumorigenic cell lines. This limitation hinders potential biostabilized AuNC-based technology in healthy cells involving both chemical and mechanical analysis, such as the direct sensing of protein function and the elucidation of local mechanical environments. Thus, new synthesis strategies are required to expand the application space of AuNCs beyond cancer-focused cellular studies. In this contribution, we have developed the methodology and demonstrated the direct in situ synthesis of AuNCs in the nontumorigenic neuronal microglial line, C8B4. The as-synthesized AuNCs form in situ and are stabilized by cellular proteins. The clusters exhibit bright green fluorescence and demonstrate low (<10%) toxicity. Interestingly, elevated ROS levels were not required for the in situ formation of AuNCs, although intracellular reductants such as glutamate were required for the synthesis of AuNCs in C8B4 cells. To our knowledge, this is the first-ever demonstration of AuNC synthesis in nontumorigenic cells and, as such, it considerably expands the application space of biostabilized fluorescent AuNCs.

Intratumoral generation of photothermal gold nanoparticles through a vectorized biomineralization of ionic gold

“…For intracellular gold nanoparticle formation, all of the constituents (sans chloroauric acid) are innately present in cellular systems. This intracellular gold nanoparticle formation has been demonstrated previously in mammalian cells, however, only through bulk diffusive treatments, lasting more than 24 h, and typically those are in solutions which contain no cell nutrients (such as DPBS) [35][36][37][38][39][40][41][42][43][44][45][46][47] . These treatment scenarios are non-ideal for minimizing the impact on cellular health.…”

“…Further to this, if cellular machinery are used to generate gold nanoparticles, the resulting plasmon-optical properties could act as a reporter of the affecting cell phenotype. This capability to generate gold nanoparticles via cellular biomolecules has already been demonstrated previously in mammalian cells [36][37][38][39][40][41][42][43][44][45][46][47] . The precise mechanism for this cellular bioreduction of gold nanoparticles is not fully understood, however, these treatments have been known to negatively impact cell health through cellular responses via shock and stress-related proteins 42 , decrease in viability 43,44 , and even cell fixation 44 .…”

“…Ionic Au-PEG clusters, prepared with 0.24 mM Au 3+ , were capable of enabling a cellular reduction within 4 h of treatment in human breast cancer cell cultures (MCF-7) in complete cell media (containing 10% FBS, with 1x penicillin and streptomycin). Previous applications for generating gold nanoparticles intracellularly, typically apply 1 mM of ionic gold over time periods >24 h [36][37][38][39][40][41][42][43][44][45][46][47] . However, the amount of formed plasmonic gold nanoparticles were negligible or considerably reduced when the ionic gold was applied diffusively in the previous literature reports either at lower concentrations (0.1 mM 43,44 , 0.25 mM 42 ) or for shorter time periods (12 h 42 , 15 h 45 ).…”

Human Nontumorigenic Microglia Synthesize Strongly Fluorescent Au/Fe Nanoclusters, Retaining Bioavailability

“… 20 − 22 As such, any nonreduced ions are then free to diffuse out of the cell without any cellular interference, essentially being viewed as a “friend” via the presence of endogenous proteins. 4 , 19 …”

“…However, in a subsequent study, we demonstrated the formation of fluorescent AuNCs in nontumorigenic mouse microglia in a non-ROS-dependent mechanism. 4 Microglia are a major component of the central nervous system’s immune system, offering first response to exogenous drugs as well as providing immune regulations of other glial cells, namely, astrocytes and oligodendrocytes. 24 Therefore, understanding the interaction, uptake, and response of glial cells to nanoscale materials, such as the fluorescent metal nanoclusters, is critical in developing nano-biotechnologies for future brain studies.…”