Microbubble-mediated Acoustic Radiation Force Enhanced Mesenchymal Stem Cell Homing To Repair Vascular Endothelium

PART I PREPARATION OF A NOVEL TARGETEDMICROBUBBLE-STEM CELL COMPLEXESObjective To prepare a novel targeted microbubble-stem cellcomplexes.Methods ICAM-1-targeted microbubbles (MBICAM-1) were preparedby biotin-avidin bridging chemistry method. Subsequently, MBICAM-1wascoated and modified by binding positively charged polylysine (PLL) totheir surface. Rabbit marrow mesenchymal stem cells (MSCs) were mixedwith PLL modified MBICAM-1, allowing for electrostatic interaction.Theattachment rate of MBICAM-1to MSCs was tested by flow cytometry.Results Surface potential of PLL modified MBICAM-1was positive.There was no significant difference in the morphous, diameter, andattachment rate to damaged human umbilical vein endothelial cells(HUVEC) between PLL modified MBICAM-1and premodified MBICAM-1(P＞0.05). MBICAM-1-MSCs complexes were prepared by successfullyattaching PLL modified MBICAM-1to MSCs. Meanwhile,the attachment rateof MSC to MBICAM-1is29.45±2.88%at the mix ratio of1:40.Conclusions MBICAM-1-MSCs complexes is prepared successfully.The attachment rate of MSC to MBICAM-1is maximal at the mix ratio of1:40(MSCs/MBICAM-1). PART Ⅱ MICROBUBBLE-MEDIATED ACOUSTICRADIATION FORCE ENHANCED MSCs HOMING TOREPAIR DAMAＧＥＤ VASCULAR ENDOTHELIUMObjective To study the efficacy of microbubble mediatied acousticradiation force enhanced MSCs on the homing to repair vascularendothelium.Methods42female New Zealand white rabbits were randomlydivided into control group (C)(n=6), the experimental group (n=36).Experimental group established iliac artery balloon injury model andrandomly divided into: surgery group (O), surgery+ultrasonic irradiationgroup (O+US), surgery+microbubbles+ultrasound irradiation group (O+MB+US), surgery+MSCs group (O+M), surgery+complexes group(O+TM), surgery+complexes+ultrasonic irradiation group (O+TM+US). After iliac artery balloon injury model were established, the maleMSCs or targeted microbubbles-stem cells complexes were transplantedinto the arterial injury immediately and incubated30min. Meanwhile,ultrasonic irradiation group used at a frequency of1MHZ and power0.66w,100%duty cycle pulse acted at the injuried artery5min. Real-Time PCRdetection of SRY mRNA expression. Another42female rabbits did thesame process.The male MSCs or targeted microbubbles-Stem cellcomplexes were transplanted into the injuried artery at the continuous7dtime, and then at7d,14d,21d,28d we executed the animals and detectedthe expression of eNOS mRNA by Real-Time PCR. At the28d, weexecuted the remaining all the animals. Sample were stained with HE toevaluate the morphological changes in the blood vessels aftertransplantation. The expression of PCNA and CD31and vWF were testedby immunohistochemistry method. Results SRY mRNA expression were detected in the group:(O+M)(O+TM) and (O+TM+US), the rest of the group was almost noexpression (P <0.05). Meanwhile,(O+TM+US) group was moreexpression than (O+M) and (O+TM)(P <0.05). After celltransplantation at7d,14d,21d,28d, compared with the control group, allother groups in eNOS gene expression were reduced. In addition to thecontrol group,(O+TM+US) at all time at the highest expression,(O+M) and (O+TM) followed,(O)(O+US) and (O+MB+US) was thelowest (P <0.05). After cell transplantation at28d, The most obvious groupof intimal hyperplasia were appeared in the group of (O)(O+US) and (O+MB+US) in the HE staining. The IA/MA and stenosis rate were thehighest in the group of (O)(O+US) and (O+MB+US), while,the groupof (O+TM+US) was the lowest,(O+M) and (O+TM) followed (P<0.05). Meanwhile, the expression of PCNA in the group of (O)、(O+US)and (O+MB+US) was the highest,(O+M) and (O+TM) followed,(O+TM+US) was the lowest (P <0.05). Expression of CD31and vWF inthe (O+TM+US) was the highest,(O+M) and (O+TM) followed, therest of the group was almost no expression (P <0.05).Conclusions Complexes combined with ultrasound irradiation groupmore effectively enhance cell home and repair endothelial than cell groupor complex group. It’s confirmed that mediated microbubble acousticradiation force can enhance homing MSCs to repair damaged vascularendothelium.