Precision Ultrasound-guided Stem Cell Delivery for Vascular Repair in Aortic Diseases

Summary

This paper presents a protocol for the precise ultrasound-guided delivery of vascular stem cells to the aortic vessel wall. The technique effectively targets the vessel wall and promotes vascular repair in aortic aneurysms and dissections.

Abstract

Aortic aneurysms and dissections rupture are life-threatening cardiovascular emergencies. Early prevention and control of disease progression are essential for improving patient outcomes. Vascular stem cells play a crucial role in the repair of vascular injuries across various vascular diseases. However, intravascular stem cell injection therapies encounter challenges in ensuring efficient engraftment of exogenous stem cells within the vascular wall.

To explore a more effective and minimally invasive approach for delivering stem cells in aortic diseases, we developed an ultrasound-guided precision-targeted stem cell delivery technology. Specifically, the technique utilized an ultra-high-resolution small animal color Doppler ultrasound imaging system to precisely locate the vascular wall of the ascending aorta in aortic disease mouse models. Subsequently, 1 × 10⁶ mCherry-labeled vascular stem cells mixed within 25 µL of Matrigel were injected using a 29 G needle. The progression of the disease was monitored by small animal ultrasound. Finally, the aorta was harvested and analyzed using 3D imaging with a light sheet microscope to track injected vascular stem cells.

The above results demonstrate that the injected vascular stem cells were delivered accurately to the vascular wall of the ascending aorta with minimal invasiveness, thereby effectively promoting vascular repair in aortic aneurysms and dissections. This ultrasound-guided stem cell delivery technology offers a promising, minimally invasive method for stem cell transplantation, potentially controlling the progression of aortic aneurysms and dissections and other related cardiovascular diseases in clinics in the future.

Introduction

Aortic aneurysms and dissections (AAD) are common and highly lethal cardiovascular diseases, typically characterized by insidious onset and rapid progression. The incidence of AAD ranges from 1.3% to 8%, with an estimated 150,000 to 200,000 deaths globally each year^1,2. Due to the acute onset and subtle clinical symptoms, many patients are in critical condition by the time of diagnosis, missing the optimal window for treatment. Although pharmacological treatments, such as β-adrenergic blockers and angiotensin receptor blockers, can slow aneurysm expansion to some extent, these therapies primarily provide ....

Protocol

All experimental procedures are conducted in accordance with the regulations of the Animal Research Committee of Capital Medical University and are approved by the Animal Welfare and Ethics Committee of Capital Medical University.

1. Preparation of injected vascular stem cells

NOTE: To verify the postinjection state and effects of vascular stem cells, Sca1⁺ cells are isolated using immunomagnetic bead selection as previously described^17,18,19, transfected with a lentivirus carrying the mCherry r....

Results

Ultrasound-guided delivery of vascular stem cells
In this study, ultrasound-guided injection was performed using a small-animal color Doppler ultrasound imaging system (Figure 1A,B). During the procedure, mice were immobilized on a heated ultrasound platform, with adjustments made to the platform and ultrasound probe as needed (Figure 1C).

Discussion

AADs caused by genetic mutations are classified as inherited AAD, among which MFS is the most prevalent. AAD represents the most life-threatening complication of MFS²². To investigate this condition, we employed a well-established mouse model of MFS (Fbn1^C1041G/+ mice), which spontaneously develops aortic aneurysms, making it ideal for early intervention studies. This model also circumvents the additional tissue damage and experimental variability associated with other mod.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
0.1% Gelatin solution	OriCell	GLT-11301
1.5 mL microtubes	Axygen	MCT-150-C
10 μL pipette tips	Labselect	T-001-10
1000 μL pipette tips	Labselect	T-001-1000
15 mL centrifuge tube	Labselect	CT-002-15A
200 μL pipette tips	Labselect	T-001-200
5 mL micro-centrifuge tube	biosharp	BS-50-M
50 mL centrifuge tube	Labselect	CT-002-50A
Antipyrine	Macklin	A800873
Anti-Sca-1 MicroBead Kit	Miltenyi Biotec	130-123-124
Bruker Luxendo MuVi -SPIM system
CD31	R&D	AF3628	Dilution ratio 1:100
CUBIC-L			tissue clearing solution
CUBIC-RA			refractive index- matching solution
DMEM	ATCC	30-2002
donkey anti-goat secondary antibody, Alexa Fluor 488	Invitrogen	A11055	Dilution ratio 1:800
FBS	Corning	35-076-CV
HBLV-mcherry-PURO	HANBIO
Homogenizer tubes	Labselect	HGT-003-200
Image-Guided Needle Injection Apparatus	Visualsonics
Insulin Syringe	BD	328421
Isoflurane	Ameixin	LAT-YFW
Light-Sheet Microscopes	Bruker Luxendo	MuVi-SPIM
Matrigel	Corning	356234
Microscan MS400 transducer	Visualsonics
Mouse Handling and Physiological Monitoring Platform	Visualsonics
N-Butyldiethanolamine	Macklin	N802391
Nicotinamide	Macklin	N814605
PBS	biosharp	BL302A
Puralube ophthalmic ointment	EQUL	1703321138
Triton X-100	Sigma-Aldrich	T9284
Trypan Blue Stain Solution	Solarbio	C0040
Trypsin-EDTA	Gibco	25200-072
Vevo 2100 lmaging System	yiswelsonics
Vevo Integrated Rail System II	yiswelsanics