Objectives Mesenchymal stem cells (MSCs) are multipotent cells obtained from adult tissues, mainly from bone marrow, but also from muscle, brain, fat, skin, gut. Dental pulps MSCs (MSCDPs) represent an easily accessible and little explored/exploited MSCs. We have shown that MSCDPs are able to both proliferate and differentiate in vitro into many histotype precursors. Several studies reported a clinical benefit by MSCs injection after myocardial infarction; however, few data are reported about the early homing of these cells in the damaged organ. We aimed to assess the early homing behavior of MSCDPs in an ex vivo infarcted heart model. Materials and methods Dental pulps were extracted from male rat’s incisor teeth and digested through a collagenase/dispase solution. Obtained suspension was enriched to get a MSCDPs culture. Cells were cultured in RPMI-1640 supplemented with 10% FCS and antibiotics. For the ex vivo studies, we used isolate rat hearts perfused according to Langendorff’s model. Ischemic injuries were performed by 30 minutes anterior descendent coronary artery ligation, followed by 4 hours of reperfusion. After 5 min of reperfusion 106 MSCDPs, previously stained with 2.5 µM carboxyfluorescein diacetate, were injected into the left ventricle apex. Stained MSCDPSs were also injected in non-ischemic hearts and used as control. To evaluate the localization and the extension of injured area, hearts were perfused with trypan blue dye. Identification of MSCDPs was performed by direct observation of thin sections of OCT-embedded hearts. Immunohistochemistry analysis was carried out in the same sections by means of specific antibodies (Connexin-43, CX43 and α-Actinin, ACTN). Results We observed significant differences between migration capabilities of MSCDPs in ischemic hearts with respect to controls: in infarcted hearts they migrated very early towards the injured areas. Moreover MSCDPs elongated side by side with cardiomyocytes, whereas in controls they remained in the injection site to form round-shaped cells clusters. In control hearts CX43 expression was confined among MSCDPs, whereas in ischemic ones it was localized also between implanted cells and myocardial tissue. Moreover, ischemic hearts showed some ACTN positive MSCDPs, otherwise negative in controls. Conclusions Data suggest that in ischemic condition MSCDPs can effectively migrate throughout the injury and integrate within the myocardium, as confirmed by the co-localization of MSCDPs and trypan blue and by the formation of GAP-junction between implanted cells and surrounding cardiomyocytes. We can infer that MSCDPs could be an interesting approach for stem cell therapy of ischemic heart.
Homing behavior of dental pulp mesenchymal stem cells in rat ischemic heart model
DI SCIPIO, FEDERICA;FOLINO, Anna;SALAMONE, PAOLINA;SPRIO, ANDREA ELIO;PENNA, Claudia;PAGLIARO, Pasquale;RASTALDO, Raffaella;BERTA, Giovanni Nicolao
2009-01-01
Abstract
Objectives Mesenchymal stem cells (MSCs) are multipotent cells obtained from adult tissues, mainly from bone marrow, but also from muscle, brain, fat, skin, gut. Dental pulps MSCs (MSCDPs) represent an easily accessible and little explored/exploited MSCs. We have shown that MSCDPs are able to both proliferate and differentiate in vitro into many histotype precursors. Several studies reported a clinical benefit by MSCs injection after myocardial infarction; however, few data are reported about the early homing of these cells in the damaged organ. We aimed to assess the early homing behavior of MSCDPs in an ex vivo infarcted heart model. Materials and methods Dental pulps were extracted from male rat’s incisor teeth and digested through a collagenase/dispase solution. Obtained suspension was enriched to get a MSCDPs culture. Cells were cultured in RPMI-1640 supplemented with 10% FCS and antibiotics. For the ex vivo studies, we used isolate rat hearts perfused according to Langendorff’s model. Ischemic injuries were performed by 30 minutes anterior descendent coronary artery ligation, followed by 4 hours of reperfusion. After 5 min of reperfusion 106 MSCDPs, previously stained with 2.5 µM carboxyfluorescein diacetate, were injected into the left ventricle apex. Stained MSCDPSs were also injected in non-ischemic hearts and used as control. To evaluate the localization and the extension of injured area, hearts were perfused with trypan blue dye. Identification of MSCDPs was performed by direct observation of thin sections of OCT-embedded hearts. Immunohistochemistry analysis was carried out in the same sections by means of specific antibodies (Connexin-43, CX43 and α-Actinin, ACTN). Results We observed significant differences between migration capabilities of MSCDPs in ischemic hearts with respect to controls: in infarcted hearts they migrated very early towards the injured areas. Moreover MSCDPs elongated side by side with cardiomyocytes, whereas in controls they remained in the injection site to form round-shaped cells clusters. In control hearts CX43 expression was confined among MSCDPs, whereas in ischemic ones it was localized also between implanted cells and myocardial tissue. Moreover, ischemic hearts showed some ACTN positive MSCDPs, otherwise negative in controls. Conclusions Data suggest that in ischemic condition MSCDPs can effectively migrate throughout the injury and integrate within the myocardium, as confirmed by the co-localization of MSCDPs and trypan blue and by the formation of GAP-junction between implanted cells and surrounding cardiomyocytes. We can infer that MSCDPs could be an interesting approach for stem cell therapy of ischemic heart.
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