Hyperhomocysteinenia (HHcy) is associated with chronic kidney disease (CKD) which has increased cardiovascular disease (CVD) mortality and mobility. Elevated inflammatory monocyte (inf. MC) is a cellular hallmark of chronic inflammation which contributes to the burden CVD. We previously reported that HHcy induces inf. MC differentiation in mice and DNA hypomethylation in vascular cells. We assesses whether HHcy causes MC differentiation in CKD and the underlying mechanism.
Degree of CKD was determined by plasma creatinine from patients with peripheral vascular disease (VD). Estimated glomerular filtration rate was calculated with modification in age, race and gender. (VD, n=13; VD with CKD, n=13; Healthy donor without evidence of VD and CKD, n=13). Blood cells were assessed for phenotypic characterization by flow cytometry.
We found that plasma Hcy levels are elevated in VD and CKD. CD14++CD16+ inf. MC are increased in VD subjects and mild HHcy(>15μM). Plasma Hcy levels are positively correlated with inf. MC, CD40, TNF receptor family 5. We identify that CKD patient serum and Hcy (50μM) treatment increased CD40 in purified human blood MC by RT-PCR. Hcy metabolites, S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), are increased in CKD subjects. SAM/SAH ratio, an indicator of methylation status, is reduced and is negatively correlated with Hcy, inf. MC, and CD40+ inf. MC. Hcy induced MC-origin inflammatory cytokines IL-6 mRNA in MC isolated from healthy donor by RT-PCR, and potentiated inflammatory cytokine IL-6, TNFα, and IFN[[Unable to Display Character: ɤ]]-induced CD40 expression in cultured PBMCs. Hcy suppressed CD40 transcription and reduced DNA methyltransferase 1 protein levels in cultured human MC. We identified four DNA hypomethylation CpG dinucleotides at p65 and PU.1 transcription factor consensus sequences on CD40 promoter in white blood cells isolated from CKD patients by Bisulfite pyrosequencing. Finally, CD40L levels are positively correlated with plasma Hcy and inf. MC. CD40 ligation by CD40L treatment (0.4μg/l) in peripheral blood mononuclear cells (PBMC) induced inf. MC differentiation.
We conclude that HHcy potentiates IFN[[Unable to Display Character: ɤ]]-mediated CD40 expression via CD40 DNA hypomethylation in CKD and promotes CD40-CD40L mediated inf. MC differentiation.
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