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Comparative localization of cystathionine beta synthases and cystathionine gamma lyase in canine, non-human primate and human retina.

ABSTRACT: Chronic exposure of the retina to light and high concentrations of polyunsaturated fatty acid in photoreceptor cells make this tissue susceptible to oxidative damage. As retinal degenerative diseases are associated with photoreceptor degeneration, the antioxidant activity of both hydrogen sulfide (H2S) and glutathione (GSH) may play an important role in ameliorating disease progression. H2S production is driven by cystathionine-?-lyase (CSE) and cystathionine-?-synthase (CBS), the key enzymes that also drive transsulfuration pathway (TSP) necessary for GSH production. As it is currently unclear whether localized production of either H2S or GSH contributes to retinal homeostasis, we undertook a comparative analysis of CBS and CSE expression in canine, non-human primate (NHP) and human retinas to determine if these antioxidants could play a regulatory role in age-related or disease-associated retinal degeneration. Retinas from normal dogs, NHPs and humans were used for the study. Laser capture microdissection (LCM) was performed to isolate individual layers of the canine retina and analyze CBS and CSE gene expression by qRT-PCR. Immunohistochemistry and western blotting were performed for CBS and CSE labeling and protein expression in dog, NHP, and human retina, respectively. Using qRT-PCR, western blot, and immunohistochemistry (IHC), we showed that CBS and CSE are expressed in the canine, NHP, and human retina. IHC results from canine retina demonstrated increased expression levels of CBS but not CSE with post-developmental aging. IHC results also showed non-overlapping localization of both proteins with CBS presenting in rods, amacrine, horizontal, and nerve fiber cell layers while CSE was expressed by RPE, cones and M?ller cells. Finally, we demonstrated that these enzymes localized to all three layers of canine, NHP and human retina: photoreceptors, outer plexiform layer (OPL) and notably in the ganglion cells layer/nerve fiber layer (GCL/NFL). QRT-PCR performed using RNA extracted from tissues isolated from these cell layers using laser capture microdissection (LCM) confirmed that each of CBS and CSE are expressed equally in these three layers. Together, these findings reveal that CSE and CBS are expressed in the retina, thereby supporting further studies to determine the role of H2S and these proteins in oxidative stress and apoptosis in retinal degenerative diseases.

SUBMITTER: Badiei A

PROVIDER: S-EPMC6443508 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Comparative localization of cystathionine beta synthases and cystathionine gamma lyase in canine, non-human primate and human retina.

Badiei Alireza A Sudharsan Raghavi R Santana Evelyn E Dunaief Joshua L JL Aguirre Gustavo D GD

Experimental eye research 20190114

Chronic exposure of the retina to light and high concentrations of polyunsaturated fatty acid in photoreceptor cells make this tissue susceptible to oxidative damage. As retinal degenerative diseases are associated with photoreceptor degeneration, the antioxidant activity of both hydrogen sulfide (H<sub>2</sub>S) and glutathione (GSH) may play an important role in ameliorating disease progression. H<sub>2</sub>S production is driven by cystathionine-γ-lyase (CSE) and cystathionine-β-synthase (CB ...[more]

PMID: 30653965

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Project description:Traditionally, hydrogen sulfide (H2S) was simply considered as a toxic and foul smelling gas, but recently H2S been brought into the spot light of cardiovascular research and development. Since the 1990s, H2S has been mounting evidence of physiological properties such as immune modification, vascular relaxation, attenuation of oxidative stress, inflammatory mitigation, and angiogenesis. H2S has since been recognized as the third physiological gaseous signaling molecule, along with CO and NO [65,66]. H2S is produced endogenously through several key enzymes, including cystathionine β-lyase (CBE), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (MST)/cysteine aminotransferase (CAT). These specific enzymes are expressed accordingly in various organ systems and CSE is the predominant H2S-producing enzyme in the cardiovascular system. The cystathionine γ-lyase (CSE)/H2S pathway has demonstrated various cardioprotective effects, including anti-atherosclerosis, anti-hypertension, pro-angiogenesis, and attenuation of myocardial ischemia-reperfusion injury. CSE exhibits its anti-atherosclerotic effect through 3 mechanisms, namely reduction of chemotactic factor inter cellular adhesion molecule-1 (ICAM-1) and CX3CR1, inhibition of macrophage lipid uptake, and induction of smooth muscle cell apoptosis via MAPK pathway. The CSE/H2S pathway's anti-hypertensive properties are demonstrated via aortic vasodilation through several mechanisms, including the direct stimulation of KATP channels of vascular smooth muscle cells (VSMCs), induction of MAPK pathway, and reduction of homocysteine buildup. Also, CSE/H2S pathway plays an important role in angiogenesis, particularly in increased endothelial cell growth and migration, and in increased vascular network length. In myocardial ischemia-reperfusion injuries, CSE/H2S pathway has shown a clear cardioprotective effect by preserving mitochondria function, increasing antioxidant production, and decreasing infarction injury size. However, CSE/H2S pathway's role in inflammation mitigation is still clouded, due to both pro and anti-inflammatory results presented in the literature, depending on the concentration and form of H2S used in specific experiment models.

Project description:BackgroundHuman uterus undergoes distinct molecular and functional changes during pregnancy and parturition. Hydrogen sulfide (H(2)S) has recently been shown to play a key role in the control of smooth muscle tension. The role of endogenous H(2)S produced locally in the control of uterine contractility during labour is unknown.Methodology/principal findingsHuman myometrium biopsies were obtained from pregnant women undergoing cesarean section at term. Immunohistochemistry analysis showed that cystathionine-γ-lyase (CSE) and cystathionine-β-synthetase (CBS), the principle enzymes responsible for H(2)S generation, were mainly localized to smooth muscle cells of human pregnant myometrium. The mRNA and protein expression of CBS as well as H(2)S production rate were down-regulated in labouring tissues compared to nonlabouring tissues. Cumulative administration of L-cysteine (10(-7)-10(-2) mol/L), a precursor of H(2)S, caused a dose-dependent decrease in the amplitude of spontaneous contractions in nonlabouring and labouring myometrium strips. L-cysteine at high concentration (10(-3) mol/L) increased the frequency of spontaneous contractions and induced tonic contraction. These effects of L-cysteine were blocked by the inhibitors of CBS and CSE. Pre-treatment of myometrium strips with glibenclamide, an inhibitor of ATP-sensitive potassium (K(ATP)) channels, abolished the inhibitory effect of L-cysteine on spontaneous contraction amplitude. The effects of L-cysteine on the amplitude of spontaneous contractions and baseline muscle tone were less potent in labouring tissues than that in nonlabouring strips.Conclusion/significanceH(2)S generated by CSE and CBS locally exerts dual effects on the contractility of pregnant myometrium. Expression of H(2)S synthetic enzymes is down-regulated during labour, suggesting that H(2)S is one of the factors involved in the transition of pregnant uterus from quiescence to contractile state after onset of parturition.

Dataset Information

Comparative localization of cystathionine beta synthases and cystathionine gamma lyase in canine, non-human primate and human retina.

Publications

Comparative localization of cystathionine beta synthases and cystathionine gamma lyase in canine, non-human primate and human retina.

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