Context-aware deep learning enables high-efficacy localization of high concentration microbubbles for super-resolution ultrasound localization microscopy.

Ultrasound localization microscopy (ULM) enables deep tissue microvascular imaging by localizing and tracking intravenously injected microbubbles circulating in the bloodstream. However, conventional localization techniques require spatially isolated microbubbles, resulting in prolonged imaging time to obtain detailed microvascular maps. Here, we introduce LOcalization with Context Awareness (LOCA)-ULM, a deep learning-based microbubble simulation and localization pipeline designed to enhance localization performance in high microbubble concentrations. In silico, LOCA-ULM enhanced microbubble detection accuracy to 97.8% and reduced the missing rate to 23.8%, outperforming conventional and deep learning-based localization methods up to 17.4% in accuracy and 37.6% in missing rate reduction. In in vivo rat brain imaging, LOCA-ULM revealed dense cerebrovascular networks and spatially adjacent microvessels undetected by conventional ULM. We further demonstrate the superior localization performance of LOCA-ULM in functional ULM (fULM) where LOCA-ULM significantly increased the functional imaging sensitivity of fULM to hemodynamic responses invoked by whisker stimulations in the rat brain.

Experimental and theoretical model of microvascular network remodeling and blood flow redistribution following minimally invasive microvessel laser ablation.

Overly dense microvascular networks are treated by selective reduction of vascular elements. Inappropriate manipulation of microvessels could result in loss of host tissue function or a worsening of the clinical problem. Here, experimental, and computational models were developed to induce blood flow changes via selective artery and vein laser ablation and study the compensatory collateral flow redistribution and vessel diameter remodeling. The microvasculature was imaged non-invasively by bright-field and multi-photon laser microscopy, and optical coherence tomography pre-ablation and up to 30 days post-ablation. A theoretical model of network remodeling was developed to compute blood flow and intravascular pressure and identify vessels most susceptible to changes in flow direction. The skin microvascular remodeling patterns were consistent among the five specimens studied. Significant remodeling occurred at various time points, beginning as early as days 1-3 and continuing beyond day 20. The remodeling patterns included collateral development, venous and arterial reopening, and both outward and inward remodeling, with variations in the time frames for each mouse. In a representative specimen, immediately post-ablation, the average artery and vein diameters increased by 14% and 23%, respectively. At day 20 post-ablation, the maximum increases in arterial and venous diameters were 2.5× and 3.3×, respectively. By day 30, the average artery diameter remained 11% increased whereas the vein diameters returned to near pre-ablation values. Some arteries regenerated across the ablation sites via endothelial cell migration, while veins either reconnected or rerouted flow around the ablation site, likely depending on local pressure driving forces. In the intact network, the theoretical model predicts that the vessels that act as collaterals after flow disruption are those most sensitive to distant changes in pressure. The model results correlate with the post-ablation microvascular remodeling patterns.

Diabetes as a Pancreatic Microvascular Disease-A Pericytic Perspective.

Diabetes is not only an endocrine but also a vascular disease. Vascular defects are usually seen as consequence of diabetes. However, at the level of the pancreatic islet, vascular alterations have been described before symptom onset. Importantly, the cellular and molecular mechanisms underlying these early vascular defects have not been identified, neither how these could impact the function of islet endocrine cells. In this review, we will discuss the possibility that dysfunction of the mural cells of the microvasculature-known as pericytes-underlies vascular defects observed in islets in pre-symptomatic stages. Pericytes are crucial for vascular homeostasis throughout the body, but their physiological and pathophysiological functions in islets have only recently started to be explored. A previous study had already raised interest in the "microvascular" approach to this disease. With our increased understanding of the crucial role of the islet microvasculature for glucose homeostasis, here we will revisit the vascular aspects of islet function and how their deregulation could contribute to diabetes pathogenesis, focusing in particular on type 1 diabetes (T1D).

A Vascular Feature Detection and Matching Method Based on Dual-Branch Fusion and Structure Enhancement.

How to obtain internal cavity features and perform image matching is a great challenge for laparoscopic 3D reconstruction. This paper proposes a method for detecting and associating vascular features based on dual-branch weighted fusion vascular structure enhancement. Our proposed method is divided into three stages, including analyzing various types of minimally invasive surgery (MIS) images and designing a universal preprocessing framework to make our method generalized. We propose a Gaussian weighted fusion vascular structure enhancement algorithm using the dual-branch Frangi measure and MFAT (multiscale fractional anisotropic tensor) to address the structural measurement differences and uneven responses between venous vessels and microvessels, providing effective structural information for vascular feature extraction. We extract vascular features through dual-circle detection based on branch point characteristics, and introduce NMS (non-maximum suppression) to reduce feature point redundancy. We also calculate the ZSSD (zero sum of squared differences) and perform feature matching on the neighboring blocks of feature points extracted from the front and back frames. The experimental results show that the proposed method has an average accuracy and repeatability score of 0.7149 and 0.5612 in the Vivo data set, respectively. By evaluating the quantity, repeatability, and accuracy of feature detection, our method has more advantages and robustness than the existing methods.

Towards standardising retinal OCT angiography image analysis with open-source toolbox OCTAVA.

Quantitative assessment of retinal microvasculature in optical coherence tomography angiography (OCTA) images is important for studying, diagnosing, monitoring, and guiding the treatment of ocular and systemic diseases. However, the OCTA user community lacks universal and transparent image analysis tools that can be applied to images from a range of OCTA instruments and provide reliable and consistent microvascular metrics from diverse datasets. We present a retinal extension to the OCTA Vascular Analyser (OCTAVA) that addresses the challenges of providing robust, easy-to-use, and transparent analysis of retinal OCTA images. OCTAVA is a user-friendly, open-source toolbox that can analyse retinal OCTA images from various instruments. The toolbox delivers seven microvascular metrics for the whole image or subregions and six metrics characterising the foveal avascular zone. We validate OCTAVA using images collected by four commercial OCTA instruments demonstrating robust performance across datasets from different instruments acquired at different sites from different study cohorts. We show that OCTAVA delivers values for retinal microvascular metrics comparable to the literature and reduces their variation between studies compared to their commercial equivalents. By making OCTAVA publicly available, we aim to expand standardised research and thereby improve the reproducibility of quantitative analysis of retinal microvascular imaging. Such improvements will help to better identify more reliable and sensitive biomarkers of ocular and systemic diseases.

Clinical Significance of Recurrent Disc Hemorrhage and Choroidal Microvasculature Dropout on Optical Coherence Tomography Angiography in Glaucoma.

Purpose: The purpose of this study was to investigate the clinical significance of recurrent disc hemorrhage (DH) and choroidal microvasculature dropout (MvD). Methods: A retrospective cohort study was conducted of 181 eyes with open-angle glaucoma. The clinical characteristics of patients with nonrecurrent and recurrent DH with and without MvD were investigated. Results: Fifty-eight patients (32.0%) had a single, nonrecurrent DH, and 63 (34.8%) had more than one DH. Sixty eyes (33.1%) with no history of DH were presented as a control group. MvD was more frequent in the recurrent DH group (44.4%) than in the nonrecurrent DH group (27.6%, P = 0.041). The recurrent DH with MvD group experienced more frequent central visual field (VF) progression (71.4%) than the recurrent DH without MvD group (17.1 %, P < 0.001). The recurrent DH without MvD group had a higher frequency of DH recurrence at different locations (42.9%) and more vascular symptoms (37.1%) than the recurrent DH with MvD group (14.3% and 7.1%, P = 0.013 and P = 0.005, respectively). Presence of DH, presence of MvD, vascular symptoms, and DH recurrence at different locations were the factors associated with central VF progression in multivariate analysis. Conclusions: DH occurrence and the presence of MvDs constitute critical parameters associated with central VF progression. In the presence of MvD, recurrent DH was more likely to recur at the same location as the MvD, whereas recurrent DH without MvD was related to vascular symptoms and recurred at other locations. When eyes present with recurrent DH and MvD, closer follow-up and more aggressive treatment are required to prevent the progression of central VF.

Computational Fractal-Based Analysis of Brain Tumor Microvascular Networks.

Brain parenchyma microvasculature is set in disarray in the presence of tumors, and malignant brain tumors are among the most vascularized neoplasms in humans. As microvessels can be easily identified in histologic specimens, quantification of microvascularity can be used alone or in combination with other histological features to increase the understanding of the dynamic behavior, diagnosis, and prognosis of brain tumors. Different brain tumors, and even subtypes of the same tumor, show specific microvascular patterns, as a kind of "microvascular fingerprint," which is particular to each histotype. Reliable morphometric parameters are required for the qualitative and quantitative characterization of the neoplastic angioarchitecture, although the lack of standardization of a technique able to quantify the microvascular patterns in an objective way has limited the "morphometric approach" in neuro-oncology.In this chapter, we focus on the importance of computational-based morphometrics, for the objective description of tumoral microvascular fingerprinting. By also introducing the concept of "angio-space," which is the tumoral space occupied by the microvessels, we here present fractal analysis as the most reliable computational tool able to offer objective parameters for the description of the microvascular networks.The spectrum of different angioarchitectural configurations can be quantified by means of Euclidean and fractal-based parameters in a multiparametric analysis, aimed to offer surrogate biomarkers of cancer. Such parameters are here described from the methodological point of view (i.e., feature extraction) as well as from the clinical perspective (i.e., relation to underlying physiology), in order to offer new computational parameters to the clinicians with the final goal of improving diagnostic and prognostic power of patients affected by brain tumors.

The proteome of the blood-brain barrier in rat and mouse: highly specific identification of proteins on the luminal surface of brain microvessels by in vivo glycocapture.

BACKGROUND: The active transport of molecules into the brain from blood is regulated by receptors, transporters, and other cell surface proteins that are present on the luminal surface of endothelial cells at the blood-brain barrier (BBB). However, proteomic profiling of proteins present on the luminal endothelial cell surface of the BBB has proven challenging due to difficulty in labelling these proteins in a way that allows efficient purification of these relatively low abundance cell surface proteins. METHODS: Here we describe a novel perfusion-based labelling workflow: in vivo glycocapture. This workflow relies on the oxidation of glycans present on the luminal vessel surface via perfusion of a mild oxidizing agent, followed by subsequent isolation of glycoproteins by covalent linkage of their oxidized glycans to hydrazide beads. Mass spectrometry-based identification of the isolated proteins enables high-confidence identification of endothelial cell surface proteins in rats and mice. RESULTS: Using the developed workflow, 347 proteins were identified from the BBB in rat and 224 proteins in mouse, for a total of 395 proteins in both species combined. These proteins included many proteins with transporter activity (73 proteins), cell adhesion proteins (47 proteins), and transmembrane signal receptors (31 proteins). To identify proteins that are enriched in vessels relative to the entire brain, we established a vessel-enrichment score and showed that proteins with a high vessel-enrichment score are involved in vascular development functions, binding to integrins, and cell adhesion. Using publicly-available single-cell RNAseq data, we show that the proteins identified by in vivo glycocapture were more likely to be detected by scRNAseq in endothelial cells than in any other cell type. Furthermore, nearly 50% of the genes encoding cell-surface proteins that were detected by scRNAseq in endothelial cells were also identified by in vivo glycocapture. CONCLUSIONS: The proteins identified by in vivo glycocapture in this work represent the most complete and specific profiling of proteins on the luminal BBB surface to date. The identified proteins reflect possible targets for the development of antibodies to improve the crossing of therapeutic proteins into the brain and will contribute to our further understanding of BBB transport mechanisms.

Dual-exposure temporal laser speckle imaging for simultaneously accessing microvascular blood perfusion and angiography.

Laser speckle contrast imaging (LSCI) has gained significant attention in the biomedical field for its ability to map the spatio-temporal dynamics of blood perfusion in vivo. However, LSCI faces difficulties in accurately resolving blood perfusion in microvessels. Although the transmissive detecting geometry can improve the spatial resolution of tissue imaging, ballistic photons directly transmitting forward through tissue without scattering will cause misestimating in the flow speed by LSCI because of the lack of a quantitative theoretical model of transmissvie LSCI. Here, we develop a model of temporal LSCI which accounts for the effect of nonscattered light on estimating decorrelation time. Based on this model, we further propose a dual-exposure temporal laser speckle imaging method (dEtLSCI) to correct the overestimation of background speed when performing traditional transmissive LSCI, and reconstruct microvascular angiography using the scattered component extracted from total transmitted light. Experimental results demonstrated that our new method opens an opportunity for LSCI to simultaneously resolve the blood vessels morphology and blood flow speed at microvascular level in various contexts, ranging from the drug-induced vascular response to angiogenesis and the blood perfusion monitoring during tumor growth.

Microvascular Network Remodeling in the Ischemic Mouse Brain Defined by Light Sheet Microscopy.

BACKGROUND: Until now, the analysis of microvascular networks in the reperfused ischemic brain has been limited due to tissue transparency challenges. METHODS: Using light sheet microscopy, we assessed microvascular network remodeling in the striatum from 3 hours to 56 days post-ischemia in 2 mouse models of transient middle cerebral artery occlusion lasting 20 or 40 minutes, resulting in mild ischemic brain injury or brain infarction, respectively. We also examined the effect of a clinically applicable S1P (sphingosine-1-phosphate) analog, FTY720 (fingolimod), on microvascular network remodeling. RESULTS: Over 56 days, we observed progressive microvascular degeneration in the reperfused striatum, that is, the lesion core, which was followed by robust angiogenesis after mild ischemic injury induced by 20-minute middle cerebral artery occlusion. However, more severe ischemic injury elicited by 40-minute middle cerebral artery occlusion resulted in incomplete microvascular remodeling. In both cases, microvascular networks did not return to their preischemic state but displayed a chronically altered pattern characterized by higher branching point density, shorter branches, higher unconnected branch density, and lower tortuosity, indicating enhanced network connectivity. FTY720 effectively increased microvascular length density, branching point density, and volume density in both models, indicating an angiogenic effect of this drug. CONCLUSIONS: Utilizing light sheet microscopy together with automated image analysis, we characterized microvascular remodeling in the ischemic lesion core in unprecedented detail. This technology will significantly advance our understanding of microvascular restorative processes and pave the way for novel treatment developments in the stroke field.

Vascularized organoid-on-a-chip: design, imaging, and analysis.

Vascularized organoid-on-a-chip (VOoC) models achieve substance exchange in deep layers of organoids and provide a more physiologically relevant system in vitro. Common designs for VOoC primarily involve two categories: self-assembly of endothelial cells (ECs) to form microvessels and pre-patterned vessel lumens, both of which include the hydrogel region for EC growth and allow for controlled fluid perfusion on the chip. Characterizing the vasculature of VOoC often relies on high-resolution microscopic imaging. However, the high scattering of turbid tissues can limit optical imaging depth. To overcome this limitation, tissue optical clearing (TOC) techniques have emerged, allowing for 3D visualization of VOoC in conjunction with optical imaging techniques. The acquisition of large-scale imaging data, coupled with high-resolution imaging in whole-mount preparations, necessitates the development of highly efficient analysis methods. In this review, we provide an overview of the chip designs and culturing strategies employed for VOoC, as well as the applicable optical imaging and TOC methods. Furthermore, we summarize the vascular analysis techniques employed in VOoC, including deep learning. Finally, we discuss the existing challenges in VOoC and vascular analysis methods and provide an outlook for future development.

Pattern recognition of microcirculation with super-resolution ultrasound imaging provides markers for early tumor response to anti-angiogenic therapy.

Rationale: Cancer treatment outcome is traditionally evaluated by tumor volume change in clinics, while tumor microvascular heterogeneity reflecting tumor response has not been fully explored due to technical limitations. Methods: We introduce a new paradigm in super-resolution ultrasound imaging, termed pattern recognition of microcirculation (PARM), which identifies both hemodynamic and morphological patterns of tumor microcirculation hidden in spatio-temporal space trajectories of microbubbles. Results: PARM demonstrates the ability to distinguish different local blood flow velocities separated by a distance of 24 µm. Compared with traditional vascular parameters, PARM-derived heterogeneity parameters prove to be more sensitive to microvascular changes following anti-angiogenic therapy. Particularly, PARM-identified "sentinel" microvasculature, exhibiting evident structural changes as early as 24 hours after treatment initiation, correlates significantly with subsequent tumor volume changes (|r| > 0.9, P < 0.05). This provides prognostic insight into tumor response much earlier than clinical criteria. Conclusions: The ability of PARM to noninvasively quantify tumor vascular heterogeneity at the microvascular level may shed new light on early-stage assessment of cancer therapy.

Pathogenic soluble tau peptide disrupts endothelial calcium signaling and vasodilation in the brain microvasculature.

The accumulation of the microtubule-associated tau protein in and around blood vessels contributes to brain microvascular dysfunction through mechanisms that are incompletely understood. Delivery of nutrients to active neurons in the brain relies on capillary calcium (Ca2+) signals to direct blood flow. The initiation and amplification of endothelial cell Ca2+ signals require an intact microtubule cytoskeleton. Since tau accumulation in endothelial cells disrupts native microtubule stability, we reasoned that tau-induced microtubule destabilization would impair endothelial Ca2+ signaling. We tested the hypothesis that tau disrupts the regulation of local cerebral blood flow by reducing endothelial cell Ca2+ signals and endothelial-dependent vasodilation. We used a pathogenic soluble tau peptide (T-peptide) model of tau aggregation and mice with genetically encoded endothelial Ca2+ sensors to measure cerebrovascular endothelial responses to tau exposure. T-peptide significantly attenuated endothelial Ca2+ activity and cortical capillary blood flow in vivo. Further, T-peptide application constricted pressurized cerebral arteries and inhibited endothelium-dependent vasodilation. This study demonstrates that pathogenic tau alters cerebrovascular function through direct attenuation of endothelial Ca2+ signaling and endothelium-dependent vasodilation.

Association between functional network connectivity, retina structure and microvasculature, and visual performance in patients after thalamic stroke: An exploratory multi-modality study.

BACKGROUND AND OBJECTIVE: Neuro-ophthalmologic symptoms and retinal changes have been increasingly observed following thalamic stroke, and there is mounting evidence indicating distinct alterations occurring in the vision-related functional network. However, the intrinsic correlations between these changes are not yet fully understood. Our objective was to explore the altered patterns of functional network connectivity and retina parameters, and their correlations with visual performance in patients with thalamic stroke. METHODS: We utilized resting-state functional MRI to obtain multi-modular functional connectivity (FC), and optical coherence tomography-angiography to measure various retina parameters, such as the retinal nerve fiber layer (RNFL), ganglion cell-inner plexiform layer (GCIPL), superficial vascular complex (SVC), and deep vascular complex. Visual acuity (VA) was used as a metric for visual performance. RESULTS: We included 46 patients with first-ever unilateral thalamic stroke (mean age 59.74 ± 10.02 years, 33 males). Significant associations were found between FC of attention-to-default mode and SVC, RNFL, and GCIPL, as well as between FC of attention-to-visual and RNFL (p < .05). Both RNFL and GCIPL exhibited significant associations with FC of visual-to-visual (p < .05). Only GCIPL showed an association with VA (p = .038). Stratified analysis based on a disease duration of 6 months revealed distinct and significant linking patterns in multi-modular FC and specific retina parameters, with varying correlations with VA in each subgroup. CONCLUSION: These findings provide valuable insight into the neural basis of the associations between brain network dysfunction and impaired visual performance in patients with thalamic stroke. Our novel findings have the potential to inform future targeted and individualized therapies. However, further comprehensive studies are necessary to validate our results.

The value of varying diffusion curvature MRI for assessing the microvascular invasion of hepatocellular carcinoma.

PURPOSE: Varying diffusion curvature (VDC) MRI is an emerging diffusion-weighted imaging (DWI) technique that can capture non-Gaussian diffusion behavior and reflect tissue heterogeneity. However, its clinical utility has hardly been evaluated. We aimed to investigate the value of the VDC technique in noninvasively assessing microvascular invasion (MVI) in hepatocellular carcinoma (HCC). METHODS: 74 patients with HCCs, including 39 MVI-positive and 35 MVI-negative HCCs were included into this prospective study. Quantitative metrics between subgroups, clinical risk factors, as well as diagnostic performance were evaluated. The power analysis was also carried out to determine the statistical power. RESULTS: MVI-positive HCCs exhibited significantly higher VDC-derived structural heterogeneity measure, D1 (0.680 ± 0.100 × 10-3 vs 0.572 ± 0.148 × 10-3 mm2/s, p = 0.001) and lower apparent diffusion coefficient (ADC) (1.350 ± 0.166 × 10-3 vs 1.471 ± 0.322 × 10-3 mm2/s, p = 0.0495) compared to MVI-negative HCCs. No statistical significance was observed for VDC-derived diffusion coefficient, D0 between the subgroups (p = 0.562). Tumor size (odds ratio (OR) = 1.242) and alpha-fetoprotein (AFP) (OR = 2.527) were identified as risk factors for MVI. A predictive nomogram was constructed based on D1, ADC, tumor size, and AFP, which exhibited the highest diagnostic accuracy (AUC = 0.817), followed by D1 (AUC = 0.753) and ADC (AUC = 0.647). The diagnostic performance of the nomogram-based model was also validated by the calibration curve and decision curve. CONCLUSION: VDC can aid in the noninvasive and preoperative diagnosis of HCC with MVI, which may result in the clinical benefit in terms of prognostic prediction and clinical decision-making.

Hemodynamic regulation allows stable growth of microvascular networks.

How do vessels find optimal radii? Capillaries are known to adapt their radii to maintain the shear stress of blood flow at the vessel wall at a set point, yet models of adaptation purely based on average shear stress have not been able to produce complex loopy networks that resemble real microvascular systems. For narrow vessels where red blood cells travel in a single file, the shear stress on vessel endothelium peaks sharply when a red blood cell passes through. We show that stable shear-stress-based adaptation is possible if vessel shear stress set points are cued to the stress peaks. Model networks that respond to peak stresses alone can quantitatively reproduce the observed zebrafish trunk microcirculation, including its adaptive trajectory when hematocrit changes or parts of the network are amputated. Our work reveals the potential for mechanotransduction alone to generate stable hydraulically tuned microvascular networks.

Using a combination of superb microvascular imaging and other auxiliary ultrasound techniques to increase the accuracy of gray-scale ultrasound for breast masses.

BACKGROUND: Breast ultrasound is highly sensitive, but its specificity is not as high for detecting malignant lesions. Auxiliary modalities like elastography, Color and Power Doppler ultrasound are used as adjuncts to yield both a high sensitivity and specificity. Superb microvascular imaging (SMI) is a newer modality with more accuracy for detecting breast lesions. In this study, our goal was to investigate the role of SMI as an adjunct to ultrasound and find a suitable combination model for the evaluation of breast masses. METHODS: In this cross-sectional study, 132 women with 172 breast masses who underwent ultrasound-guided biopsy were included.. The ultrasound features of the lesion, the strain ratio in strain elastography, the number of vessels for each lesion, their morphology and distribution in Doppler and Power Doppler ultrasound and SMI were recorded for each lesion. A vascular score and a vascular ratio were defined. RESULTS: In the histologic examination, 31 lesions (18%) were malignant and 141 lesions (82%) were benign. The vascular score was more accurate than the vascular ratio in all three modalities. The predictive ability of strain ratio was higher than Doppler and Power Doppler ultrasound and SMI. Adding SMI alone to ultrasound increased the specificity from 46.10% to 61.2% and the accuracy from 55.80% to 70.11%. In the combination of ultrasound with other modalities, the best was the combination of ultrasound, strain elastography, and SMI; which yielded a specificity and sensitivity of 100% and 74.4%, respectively. CONCLUSION: Adding SMI and STE modalities as adjuncts to ultrasound lowers the chance of missing malignant lesions and reduces unnecessary biopsies of breast lesions. A study with a larger sample size using this combination model to evaluate the accuracy with greater precision is recommended.

Engineering primitive multiscale chimeric vasculature by combining human microvessels with explanted murine vessels.

Strategies to separately manufacture arterial-scale tissue engineered vascular grafts and microvascular networks have been well-established, but efforts to bridge these two length scales to create hierarchical vasculature capable of supporting parenchymal cell functions or restoring perfusion to ischemic tissues have been limited. This work aimed to create multiscale vascular constructs by assessing the capability of macroscopic vessels isolated from mice to form functional connections to engineered capillary networks ex vivo. Vessels of venous and arterial origins from both thoracic and femoral locations were isolated from mice, and then evaluated for their abilities to sprout endothelial cells (EC) capable of inosculating with surrounding human cell-derived microvasculature within bulk fibrin hydrogels. Comparing aortae, vena cavae, and femoral vessel bundles, we identified the thoracic aorta as the rodent macrovessel that yielded the greatest degree of sprouting and interconnection to surrounding capillaries. The presence of cells undergoing vascular morphogenesis in the surrounding hydrogel attenuated EC sprouting from the macrovessel compared to sprouting into acellular hydrogels, but ultimately sprouted mouse EC interacted with human cell-derived capillary networks in the bulk, yielding chimeric vessels. We then integrated micromolded mesovessels into the constructs to engineer a primitive 3-scale vascular hierarchy comprising capillaries, mesovessels, and macrovessels. Overall, this study yielded a primitive hierarchical vasculature suitable as proof-of-concept for regenerative medicine applications and as an experimental model to better understand the spontaneous formation of host-graft vessel anastomoses.