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Placental growth factor regulates the pentose phosphate pathway and antioxidant defense systems in human retinal endothelial cells.

ABSTRACT: The molecular mechanisms whereby placental growth factor (PlGF) mediates its effects in nonproliferative diabetic retinopathy (DR) are unknown. To better understand the role of PlGF in DR, we used tandem mass tags (TMT)-labeled quantitative proteomics to human retinal endothelial cells (HRECs), treated anti-PlGF antibody, and PBS as a control. Functional annotation and pathway enrichments were performed, which suggested that the differentially expressed proteins (DEPs) were involved in key metabolic processes, protein binding, and membrane, pentose phosphate pathway (PPP) and adherens junction. We conducted integrated gene profiles of our previously published transcriptomic data to the TMT-labeled proteomics data. The results showed the sixty genes were found to be changed at the proteome level. The functional annotation conducted for the sixty proteins suggested that 58.3% of proteins were involved in PPP, 25% of proteins were in interleukin-12 singling and 16.7% of proteins were involved in glycolysis and gluconeogenesis pathway. Mass spectrometry results were validated by transendothelial electrical resistance measurement by an electrical cell-impedance sensing (ECIS) and western blot analysis of VE-cadherin, G6PD. These findings suggest that the PPP proteins and antioxidants may act as a downstream target of PlGF and may play a decisive role in HREC biological functions in DR. SIGNIFICANCE: PlGF (Placental growth factor) is known to play a pivotal role in pathological angiogenesis and inflammation by stimulating endothelial cell migration and by recruiting pericytes and inflammatory cells such as microglia and macrophages. Despite the well-defined pathophysiological roles of PlGF, the underlying molecular and cellular mechanisms are not completely understood, especially the exact relationships between biochemical events and molecular pathways regulated by PlGF, whose inhibition exhibits a protective role in DR. This study provides new insights into protein expression patterns and enables the identification of many attractive candidates for investigation of PPP pathway role in the activation of the antioxidant defense system in DR. Our findings suggest that the PPP proteins and antioxidants (PRDX6, HMOX1, NQO1 and YES1) may act as downstream targets of PlGF and may play a decisive role in HREC biological functions in DR.

SUBMITTER: Saddala MS

PROVIDER: S-EPMC7439780 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:Metabolic dysfunction and neuroinflammation are increasingly implicated in Parkinson's disease (PD). The pentose phosphate pathway (PPP, a metabolic pathway parallel to glycolysis) converts glucose-6-phosphate into pentoses and generates ribose-5-phosphate and NADPH thereby governing anabolic biosynthesis and redox homeostasis. Brains and immune cells display high activity of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP. A postmortem study reveals dysregulation of G6PD enzyme in brains of PD patients. However, spatial and temporal changes in activity/expression of G6PD in PD remain undetermined. More importantly, it is unclear how dysfunction of G6PD and the PPP affects neuroinflammation and neurodegeneration in PD. METHODS:We examined expression/activity of G6PD and its association with microglial activation and dopaminergic neurodegeneration in multiple chronic PD models generated by an intranigral/intraperitoneal injection of LPS, daily subcutaneous injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) for 6?days, or transgenic expression of A53T ?-synuclein. Primary microglia were transfected with G6PD siRNAs and treated with lipopolysaccharide (LPS) to examine effects of G6PD knockdown on microglial activation and death of co-cultured neurons. LPS alone or with G6PD inhibitor(s) was administrated to mouse substantia nigra or midbrain neuron-glia cultures. While histological and biochemical analyses were conducted to examine microglial activation and dopaminergic neurodegeneration in vitro and in vivo, rotarod behavior test was performed to evaluate locomotor impairment in mice. RESULTS:Expression and activity of G6PD were elevated in LPS-treated midbrain neuron-glia cultures (an in vitro PD model) and the substantia nigra of four in vivo PD models. Such elevation was positively associated with microglial activation and dopaminergic neurodegeneration. Furthermore, inhibition of G6PD by 6-aminonicotinamide and dehydroepiandrosterone and knockdown of microglial G6PD attenuated LPS-elicited chronic dopaminergic neurodegeneration. Mechanistically, microglia with elevated G6PD activity/expression produced excessive NADPH and provided abundant substrate to over-activated NADPH oxidase (NOX2) leading to production of excessive reactive oxygen species (ROS). Knockdown and inhibition of G6PD ameliorated LPS-triggered production of ROS and activation of NF-?B thereby dampening microglial activation. CONCLUSIONS:Our findings indicated that G6PD-mediated PPP dysfunction and neuroinflammation exacerbated each other mediating chronic dopaminergic neurodegeneration and locomotor impairment. Insight into metabolic-inflammatory interface suggests that G6PD and NOX2 are potential therapeutic targets for PD.

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Placental growth factor regulates the pentose phosphate pathway and antioxidant defense systems in human retinal endothelial cells.

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