Use of p53‐Silenced Endothelial Progenitor Cells to Treat Ischemia in Diabetic Peripheral Vascular Disease

Abstract

BackgroundPeripheral vascular disease is a major diabetes mellitus‐related complication. In this study, we noted that expressions of proapoptotic p53 gene and its downstream cascade gene such as p21 are upregulated in hyperglycemia. Therefore, we investigated whether p53‐ and p21‐silenced endothelial progenitor cells (EPCs) were able to survive in hyperglycemic milieu, and whether transplantation of either p53 knockout (KO) or p21KO or p53‐ and p21‐silenced EPCs could improve collateral vessel formation and blood flow in diabetic vaso‐occlusive peripheral vascular disease mouse models.Methods and ResultsWe transplanted p53 and p21KO mouse EPCs (mEPCs) into streptozotocin–induced diabetic (type 1 diabetes mellitus model) C57BL/6J and db/db (B6.BKS(D)‐Leprdb/J) (type 2 model) post–femoral artery occlusion. Similarly, Ad‐p53–silenced and Ad‐p21–silenced human EPCs (CD34+) cells were transplanted into streptozotocin‐induced diabetic NOD.CB17‐Prkdcscid/J mice. We measured blood flow at 3, 7, and 10 days and hindlimb muscles were obtained postsacrifice for mRNA estimation and CD31 staining. Enhanced blood flow was noted with delivery of p53 and p21KO mEPCs in streptozotocin‐induced diabetic C57BL/6J mice. Similar results were obtained when human Ad‐p53shEPCs(CD34+) and Ad‐p21shEPCs(CD34+) were transplanted into streptozotocin‐induced nonobese diabetic severe combined immunodeficiency mice. Gene expression analysis of p53 and p21KO EPCs transplanted hindlimb muscles showed increased expression of endothelial markers such as endothelial nitric oxide synthase, vascular endothelial growth factor A, and platelet endothelial cell adhesion molecule 1. Similarly, quantitative reverse transcriptase polymerase chain reaction of human Ad‐p53shEPCs (CD34+)– and Ad‐p21shEPCs (CD34+)–transplanted hindlimb muscles also showed increased expression of endothelial markers such as vascular endothelial growth factor A, noted primarily in the p53‐silenced EPCs group. However, such beneficial effect was not noted in the db/db type 2 diabetic mouse models.ConclusionsTransient silencing of p53 using adenoviral vector in EPCs may have a therapeutic role in diabetic peripheral vascular disease.

The synergistic effects of saxagliptin and metformin on CD34+ endothelial progenitor cells in early type 2 diabetes patients: a randomized clinical trial

“…This is concurrent with reduced inflammatory and apoptosis cascade markers such as IL6 and Caspase-3 mRNA expressions in CD34− cells indicating an overall reduction in inflammation and apoptosis pathway, which could be secondary to upregulation in antioxidants. This is particularly interesting as there is sufficient evidence from our previous work, to suggest significant patho-physiological role of ROS (anti-oxidant expression may increase in response to increased intracellular ROS presence) in not only EPCs but also in MSCs in a setting of hyperglycemia, as seen in diabetes [ 10 , 39 ]. The decrease of IGF-1 mRNA expression in CD34− cells in the saxagliptin intervention group compared to placebo may indicate reduced insulin resistance at the levels of MNCs, which also supports reduction of cellular inflammation.…”

“…Therefore, endothelial progenitor cells (EPCs, defined here as CD34+ cells), which are specialized cells responsible for endothelial repair and neo-angiogenesis, play an important role in diabetes. It has been shown that EPCs are impaired in number, function and gene expression in hyperglycemia and diabetes related complications [ 6 – 10 ]. Moreover, it has been reported that EPCs (CD34+) from diabetic patients failed to incorporate and repair damaged vessels [ 11 ].…”

Effects of endothelial progenitor cells transplantation on hyperlipidemia associated kidney damage in ApoE knockout mouse model

“…In a mouse model of hypertension-hypercholesterolemia, the administration of EPCs reduced hepatic lipid accumulation and consequently alleviated hypertension and dyslipidaemia [26]. In contrast, Silvestre et al found that EPC treatment aggravated the atherosclerosis burden when EPCs were transferred from young to old ApoE −/− mice [7]. Therefore, the results of prior atherosclerosis studies conflict in terms of both the potential benefits and adverse outcomes of EPC treatment.…”

“…Nevertheless, the potential therapeutic effects of EPCs on hyperlipidaemia-associated kidney damage have not been addressed. A previous study in an acute kidney injury model found that administration of EPCs enhanced microvascular endothelial regeneration and protected against kidney fibrosis [5]; however, Silvestre found that EPC treatment augmented the lesion burden when EPCs were transferred from young to old ApoE −/− mice [7]. These therapies are still controversial and inconclusive.…”

Linagliptin, when compared to placebo, improves CD34+ve endothelial progenitor cells in type 2 diabetes subjects with chronic kidney disease taking metformin and/or insulin: a randomized controlled trial

“…It has been shown that high glucose environment, as seen in diabetes mellitus (either type 1 or 2), leads to functional impairment of circulating EPCs. Their number goes down, along with their ability to form colonies and migration to the site of endothelial damage [6][7][8][9][10]. Other studies have shown that healthy CD34 + cells can effectively repair damaged endothelial cell lining [11].…”