5 replies to this topic

[FunkOdyssey]

Studies that have shown pro-inflammatory effects of C60 have been discounted on this forum due to the fact they were usually hydroxylated, water-soluble forms. What rationale are we using to dismiss the following study:

Distinct cytotoxic mechanisms of pristine versus hydroxylated fullerene.

AuthorsIsakovic A, et al. Show all Journal
Toxicol Sci. 2006 May;91(1):173-83. Epub 2006 Feb 13.

Affiliation
Institute of Biochemistry, School of Medicine, University of Belgrade, 11000 Belgrade, Serbia and Montenegro.

Abstract
The mechanisms underlying the cytotoxic action of pure fullerene suspension (nano-C60) and water-soluble polyhydroxylated fullerene [C60(OH)n] were investigated. Crystal violet assay for cell viability demonstrated that nano-C60 was at least three orders of magnitude more toxic than C60(OH)n to mouse L929 fibrosarcoma, rat C6 glioma, and U251 human glioma cell lines. Flow cytometry analysis of cells stained with propidium iodide (PI), PI/annexin V-fluorescein isothiocyanate, or the redox-sensitive dye dihydrorhodamine revealed that nano-C60 caused rapid (observable after few hours), reactive oxygen species (ROS)-associated necrosis characterized by cell membrane damage without DNA fragmentation. In contrast, C60(OH)n caused delayed, ROS-independent cell death with characteristics of apoptosis, including DNA fragmentation and loss of cell membrane asymmetry in the absence of increased permeability. Accordingly, the antioxidant N-acetylcysteine protected the cell lines from nano-C60 toxicity, but not C60(OH)n toxicity, while the pan-caspase inhibitor z-VAD-fmk blocked C60(OH)n-induced apoptosis, but not nano-C60-mediated necrosis. Finally, C60(OH)n antagonized, while nano-C60 synergized with, the cytotoxic action of oxidative stress-inducing agents hydrogen peroxide and peroxynitrite donor 3-morpholinosydnonimine. Therefore, unlike polyhydroxylated C60 that exerts mainly antioxidant/cytoprotective and only mild ROS-independent pro-apoptotic activity, pure crystalline C60 seems to be endowed with strong pro-oxidant capacity responsible for the rapid necrotic cell death.

PMID 16476688

[niner]

Studies that have shown pro-inflammatory effects of C60 have been discounted on this forum due to the fact they were usually hydroxylated, water-soluble forms. What rationale are we using to dismiss the following study:

I'll take a shot at it... Nano-c60 is an aggregate of pristine c60, while we're dealing with a molecular c60 species that most likely has two (or so) long chain fatty acid substituents. (Mostly oleic acid) These are pretty different animals, with different properties. The experimental setup here is cancer cells in a petri dish, while we're dealing with intact animals. The oxygen level and light level in the petri dish will be much higher than in vivo, and the concentration of the c60 products will no doubt be vastly higher than any cells in vivo will see, with the possible exception of some gut endothelium. (Or Anthony Loera...) I think that if c60-oo was toxic in vivo, Baati's rats wouldn't have lived so long, and a lot of us would not be feeling so good.

We still need a lot of careful toxicology work on c60-oo, which I presume will come in the fullness of time, but based on everything I've seen, I'm not willing to wait five or ten years for all the scientific i's to be dotted and t's to be crossed. (Just in time for c60 sales to individuals to be outlawed...)

[Adaptogen]

do you really think it will be outlawed? for what reasons?

[niner]

do you really think it will be outlawed? for what reasons?

I was only half serious. If it turned out to substantially increase human life (say maybe an extra 20 years? Or even 10), then it would break the actuarial assumptions of pensions, annuities, and Social Security. This might be considered to be too disruptive to society. Or maybe religious people would get weirded out by it. Another angle might be if large numbers of people were using it to treat diseases, which may well become a bigger market for c60 than antiaging, maybe it would be shut down by the FDA. High purity c60 is already more expensive, gram for gram, than gold. What if the supply/demand equation drove the price up by an order of magnitude or two? That would be effectively like outlawing it, for a lot of people. If I had to bet money on it, I would say that these things weren't going to happen. On the other hand, I can't rule them out completely.

[hav]

Studies that have shown pro-inflammatory effects of C60 have been discounted on this forum due to the fact they were usually hydroxylated, water-soluble forms. What rationale are we using to dismiss the following study:

Actually, the study you're thinking of, done in 2004, was discounted on other grounds. Although it involved water-soluble c60, it was the use of tetrahydrofuran to form a THF/C60 adduct that turned out to be responsible for the toxicity. The author concluded that in a followup study done 2 years later in 2006. The study you cited, done in 2005 and citing the earlier 2004 study, did the same thing, even with their so-called pristine C60 which was actually a colloidal mix of C60/C70 reacted with THF:


Distinct Cytotoxic Mechanisms of Pristine versus Hydroxylated Fullerene

C_60/70 colloid (referred to as nano-C₆₀ for reasons of convenience) was produced by evaporating tetrahydrofuran (THF) from a mixture of water and molecularly dispersed C_60/70 in THF (Sigma, St. Louis, MO), using the procedure first described by Deguchi et al. (2001) and modified by Fortner et al. (2005). The concentration of nano-C₆₀ suspension in water was adjusted by evaporation to 10 μg/ml, as determined from the absorption spectrum and using a gravimetric procedure. Polyhydroxylated fullerene, referred to as C₆₀(OH)_n, was prepared as previously described (Zhao et al., 2004) from the same C_60/70 extract of carbon soot used for the nano-C₆₀ preparation.

Howard

[niner]

Thanks Howard, good catch. Here's a recent review on the topic:

Curr Opin Biotechnol. 2011 Aug;22(4):533-7. doi: 10.1016/j.copbio.2011.05.511. Epub 2011 Jun 28.
Aqueous fullerene aggregates (nC60) generate minimal reactive oxygen species and are of low toxicity in fish: a revision of previous reports.
Henry TB, Petersen EJ, Compton RN.

School of Biomedical and Biological Sciences, University of Plymouth, Plymouth, Devon, United Kingdom. ted.henry@plymouth.ac.uk

This review aims to clarify inconsistencies in previous reports regarding the potential for aqueous aggregates of fullerenes (nC60) to generate reactive oxygen species (ROS) and cause toxicity in fish. Methods for evaluation of ROS production and toxicity of aqueous nC60 have evolved over time and limitations in initial studies have led to unintentional erroneous reports of nC60 ROS generation and toxicity. Some of these reports continue to lead to misconceptions of the environmental effects of C60. Critical review of the evidence (2007-2011) indicates that aqueous nC60 have minimal potential to produce ROS and that oxidative stress in fish is not induced by environmentally relevant exposure to nC60. Future studies should acknowledge that current evidence indicates low toxicity of nC60 and refrain from citing articles that attribute toxicity in fish to nC60 based on methods shown to be compromised by experimental artifacts. Despite low toxicity of nC60 in fish, an emerging environmental issue is that nC60 can affect environmental fate, transport, and bioavailability of co-contaminants in aquatic environments in a similar manner to that observed for other anthropogenic particulates (e.g., microplastics).

PMID: 21719272