Biomedicines, Vol. 11, Pages 60: Gamma-Synuclein Dysfunction Causes Autoantibody Formation in Glaucoma Patients and Dysregulation of Intraocular Pressure in Mice

Conceptualization, M.S.K., T.A.P. and R.K.O.; methodology, A.Y.R. and N.G.D.; validation, O.A.L., M.S.K. and R.K.O.; formal analysis, M.S.K.; investigation, A.Y.R., N.E.P., I.S.S. and V.O.S.; resources, M.S.K. and N.B.C.; data curation, M.S.K.; writing—original draft preparation, M.S.K. and T.A.P.; writing—review and editing, M.W.E. and N.B.C.; visualization, M.S.K.; supervision, M.S.K. and R.K.O.; funding acquisition, R.K.O. All authors have read and agreed to the published version of the manuscript.

Figure 1. Detection of autoantibodies to γ-synuclein in the serum of patients with glaucoma by immunoblotting: (a,b) correspond to different independent membranes divided into strips. Each strip of membrane was incubated with a serum sample from one patient. On the left, the molecular weight (kDa) is indicated according to the protein ladder. The numbers 14, 24, 26, 30, 38 indicate the numbers of serum samples from patients with glaucoma, in which autoantibodies to γ-synuclein (17 kDa) were detected. The arrow on the right marks the band corresponding to γ-synuclein.

Figure 1. Detection of autoantibodies to γ-synuclein in the serum of patients with glaucoma by immunoblotting: (a,b) correspond to different independent membranes divided into strips. Each strip of membrane was incubated with a serum sample from one patient. On the left, the molecular weight (kDa) is indicated according to the protein ladder. The numbers 14, 24, 26, 30, 38 indicate the numbers of serum samples from patients with glaucoma, in which autoantibodies to γ-synuclein (17 kDa) were detected. The arrow on the right marks the band corresponding to γ-synuclein.

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Figure 2. IOP change in γ-synuclein knockout (γ-KO) mice compared to wild-type (WT) mice after instillation of drugs affecting different neuromediators: (a) adrenergic agonist 1% phenylephrine, (b) adrenergic antagonist 0.5% timolol, (c) cholinergic agonist 1% pilocarpine, (d) cholinergic antagonist 0.1% atropine, (e) dopamine agonist 10% dopamine and (f) dopamine antagonist 0.25% haloperidol. IOP was measured before (0 h) and 1 and 2 h after drugs installation. Mean ± SE are presented. Statistical analysis was performed using ANOVA followed by Holm-Sidak’s multiple comparisons test between groups (* p  < 0.05, ** p  < 0.05, *** p  < 0.001). For each group, n = 5 (10 eyes).

Figure 2. IOP change in γ-synuclein knockout (γ-KO) mice compared to wild-type (WT) mice after instillation of drugs affecting different neuromediators: (a) adrenergic agonist 1% phenylephrine, (b) adrenergic antagonist 0.5% timolol, (c) cholinergic agonist 1% pilocarpine, (d) cholinergic antagonist 0.1% atropine, (e) dopamine agonist 10% dopamine and (f) dopamine antagonist 0.25% haloperidol. IOP was measured before (0 h) and 1 and 2 h after drugs installation. Mean ± SE are presented. Statistical analysis was performed using ANOVA followed by Holm-Sidak’s multiple comparisons test between groups (* p  < 0.05, ** p  < 0.05, *** p  < 0.001). For each group, n = 5 (10 eyes).

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Figure 3. Total protein concentration and α2-macroglobulin (α2-MG) activity are different in tear fluid of γ-synuclein knockout (γ-KO) mice than those of wild-type (WT) mice: (a) total protein concentration determined by the Lowry assay (mg/mL); (b) total activity (nmol/min × mL); (c) specific activity (nmol/min × mg) of α2-MG in tear fluid of γ-KO and WT mice. Means ± SE with individual data are presented and p values are given if differences between groups were statistically significant according to the Mann–Whitney test. For the WT mice group, n = 7; for the γ-KO group, n = 5.

Figure 3. Total protein concentration and α2-macroglobulin (α2-MG) activity are different in tear fluid of γ-synuclein knockout (γ-KO) mice than those of wild-type (WT) mice: (a) total protein concentration determined by the Lowry assay (mg/mL); (b) total activity (nmol/min × mL); (c) specific activity (nmol/min × mg) of α2-MG in tear fluid of γ-KO and WT mice. Means ± SE with individual data are presented and p values are given if differences between groups were statistically significant according to the Mann–Whitney test. For the WT mice group, n = 7; for the γ-KO group, n = 5.

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Table 1. Clinical data of patients and control subjects.

Table 1. Clinical data of patients and control subjects.

Patient
NumberGenderAge (Years)DiagnosisAutoantibodies to γ-SynucleinStage of the DiseaseSurgical Treatment of
Glaucoma Control group1M44No glaucoma symptoms-NANA2F91No glaucoma symptoms-NANA3M70No glaucoma symptoms-NANA4M85No glaucoma symptoms-NANA5M64No glaucoma symptoms-NANA6M64No glaucoma symptoms-NANA7F80No glaucoma symptoms-NANA8F59No glaucoma symptoms-NANA9F48No glaucoma symptoms-NANA10F67No glaucoma symptoms-NANA11F63No glaucoma symptoms-NANA12M62No glaucoma symptoms-NANA13F67No glaucoma symptoms-NANAPrimary open-angle glaucoma14M80POAG+4 OUYes15F78POAG-3 OUNo16M76POAG-5 OS; 2 ODYes17M78POAG-2OUYes18F78POAG-1 OUNo19M78POAG-4 OS; 1 ODNo20F86POAG-1 OUNo21F75POAG-3 OUYes22M69POAG-4OS; 3 ODYes23F59POAG-1 OS glaucoma suspect; 1 ODYes24M76POAG+3 OS; 1 ODYes25F69POAG-OU glaucoma suspectNo26M80POAG+2 OS; 1 ODYes27M69POAG-1 OS; 3 ODYes28F80POAG-2 OUNo29M53POAG-3 OUYes30M76POAG+2 OS; 3 ODYes31F85POAG-2 OS, OD anophthalmiaNo32M64POAG-3 OUNo33F68POAG-OU glaucoma suspectNo34F71POAG-3 OS; 1 ODNo35M79POAG-4 OS; 3 ODYes36F68POAG-1 OS; 2 ODYes37F71POAG-2 OUYes38M65POAG+1 OS; 2 ODNo

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