Metabolomic profiling of reactive persulfides and polysulfides in the aqueous and vitreous humors

Abstract

We investigate the metabolomic profile of reactive persulfides and polysulfides in the aqueous and vitreous humors. Eighteen eyes of 18 consecutive patients with diabetes mellitus (DM) and diabetic retinopathy underwent microincision vitrectomy combined with cataract surgery. Samples of the aqueous and vitreous humors were collected and underwent mass spectrometry-based metabolomic profiling of reactive persulfides and polysulfides (polysulfidomics). The effect of reactive polysulfide species on the viability of immortalized retinal cells (the RGC-5 cell line) under oxidative stress (induced with H2O2) was also evaluated with an Alamar Blue assay. The experiments showed that cysteine persulfides (CysSSH), oxidized glutathione trisulfide (GSSSG) and cystine were elevated in the aqueous humor, and CysSSH, Cys, and cystine were elevated in the vitreous. Furthermore, GSSSG, cystine, and CysSSH levels were correlated in the aqueous and vitreous humors. A comparison, in DM and control subjects, of plasma levels of reactive persulfides and polysulfides showed that they did not differ. In vitro findings revealed that reactive polysulfide species increased cell viability under oxidative stress. Thus, various reactive persulfides and polysulfides appear to be present in the eye, and some reactive sulfide species, which have a protective effect against oxidative stress, are upregulated in the aqueous and vitreous humors of DM eyes.

Detection of small molecule concentration gradients in ocular tissues and humours

“…However, the majority of m/z signals remain unidentified, particularly in the black bream potentially due to the lack of ocular humour analysis of teleost eyes, and their identification would allow additional interpretation of their possible biological significance to be made. In the rabbit eyeball analysis, the trends of our provisionally identified ocular humour metabolite distributions were consistent with previous studies, where cystine (the more stable, oxidised form of cysteine), uric acid, and citric acid have been detected in AH at higher abundance than VH, and taurine, as an important molecule that supports retinal photoreceptor cell function, has been detected at higher levels in the VH than AH . Complementary orthogonal LC‐MS metabolomics studies would aid greatly in identifying endogenous metabolites but are beyond the scope of this method.…”

Enzymatic Regulation and Biological Functions of Reactive Cysteine Persulfides and Polysulfides

“…Nakazawa and colleagues reported the endogenous occurrence of persulfide species in the aqueous and vitreous humour [ 7 ]. They measured persulfides and polysulfides by means of LC-MS/MS with HPE-IAM, and found that patients with diabetes mellitus (DM) had elevated levels of CysSSH, cystine, and GSSSG in the aqueous humour compared with healthy subjects.…”

“…Cysteine persulfide (CysSSH) and cysteine polysulfides (CysSS n H, n > 1) are cysteine derivatives that have sulfane sulfur atoms that are bound to cysteine thiol [ 1 , 2 , 3 , 4 ]. Various forms of persulfides/polysulfides exist in both prokaryotes and eukaryotes as low-molecular-weight compounds, such as CysSSH, homocysteine persulfide, glutathione persulfide (GSSH), glutathione trisulfide (GSSSH), oxidized glutathione trisulfide (GSSSG), bacillithiol persulfide, and coenzyme A persulfide, and as CysSSHs in proteins ( Figure 1 ) [ 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 ]. Persulfide species have been suggested to be involved in various biological processes.…”

“…H 2 S thus formed is likely followed by oxidation coupled with membrane potential generation [ 6 ]. CysSSH and related molecules can also act as strong nucleophiles and antioxidants and may have an important role in regulating oxidative stress and redox signaling in cells [ 1 , 5 , 6 , 7 , 19 , 20 ]. For instance, GSSH directly decomposed hydrogen peroxide (H 2 O 2 ) in test tubes where parental GSH did not [ 5 ].…”

“…For instance, GSSH directly decomposed hydrogen peroxide (H 2 O 2 ) in test tubes where parental GSH did not [ 5 ]. In addition, cell treatment with the GSSH donor, GSSSG, protected the cells from H 2 O 2 -induced cell death [ 7 ]. We recently demonstrated that CysSSH and related molecules have potent anti-inflammatory properties [ 13 ].…”