Effect Of 5-hydroxytryptamine On Vascular Calcification In Rats With Chronic Kidney Disease And The Underlying Mechanisms

BackgroundPathological vascular calcification is intrinsically linked with cardiovascular complications in patients suffering from chronic kidney disease(CKD).The process of vascular calcification shares similarities with bone formation.Numerous studies over the past few years have shed light on several mechanisms contributing to vascular calcification.These include factors such as the osteogenic transdifferentiation of vascular smooth muscle cells(VSMCs),apoptosis,oxidative stress,endoplasmic reticulum stress,alterations in calcium/phosphate balance,and an imbalance between pro-calcification and anti-calcification factors.Despite this understanding,we currently lack effective therapeutic strategies to curb vascular calcification.5-hydroxytryptamine(5-HT),a biogenic amine,is known to stimulate bone formation by enhancing the differentiation of osteocytes.Patients with CKD often exhibit high plasma levels of 5-HT.However,the role of 5-HT in CKD-induced vascular calcification is yet to be thoroughly understood.This study aims to delve into the impact of 5-HT on vascular calcification in CKD and unravel the underlying mechanisms.ObjectiveThis study seeks to uncover the role of 5-HT in CKD-induced vascular calcification and the underlying mechanisms utilizing in vitro,ex vivo,and in vivo models.Methods and results1.Plasma 5-HT levels were compared between two groups of CKD patients:11 without aortic calcification and 17 with aortic arch calcification.The results demonstrated significantly higher plasma 5-HT levels in the latter group,implying a potential role for 5-HT in the progression of vascular calcification in CKD patients.2.Rat VSMCs and arterial rings were exposed to varying concentrations of 5-HT(1 μM,10 μM,100 μM)in a calcifying medium.Based on alizarin red staining and formic acid quantification results,it was determined that 5-HT enhances mineral deposition in both VSMCs and aortic rings.Calcium content analysis further confirmed that 5-HT increases the calcium content of VSMCs and aortic rings.Western blot analysis provided additional evidence,showing that 5-HT elevates the protein expression of osteogenic differentiation-related molecules,Runx2 and BMP2.An in vivo experiment was also conducted using a CKD rat model.Rats were divided into sham,model,and 5-HT treatment groups,and alizarin red staining and calcium content testing demonstrated that 5-HT exacerbates aortic calcification in CKD rats.3.Treatment with 5-HT was found to augment the mRNA level of the 5-HT2B receptor(5-HT2BR),suggesting that 5-HT2BR in VSMCs is upregulated by 5-HT treatment.Subsequently,VSMCs were incubated with 5-HT in a calcifying medium using RS-127445(a selective inhibitor of 5-HT2BR)or 5-HT2BR small interfering RNA.Following this treatment,calcium content analysis and alizarin red staining revealed a decrease in mineral deposition and calcium content within VSMCs.Western blot analysis further demonstrated a reduction in protein expression levels of Runx2 and BMP2.These results imply that the pro-calcifying effect of 5-HT is mediated through 5-HT2BR.Conclusions1.5-HT facilitates osteogenic differentiation and calcification within rat VSMCs and also enhances calcification of rat arterial rings.2.5-HT accelerates the aortic calcification in CKD rats.3.5-HT promotes vascular calcification by activating 5-HT2B receptor.