Hypoxia-Inducible Factor Drives Vascularization of Modularly Assembled Engineered Tissue.

IMPACT STATEMENT: Using two inhibitory methods, we demonstrated that hypoxia-inducible factor (HIF) plays an important role in vascularizing and oxygenating modularly-assembled engineered tissues. Each inhibitory technique elucidated a different mechanism by which this occurred. Whereas systemic inhibition negatively impacted early recruitment of host-derived cells, genetic inhibition in grafted endothelial cells was detrimental to their survival. Taken together, our study suggests that methods of HIF-mediated mechanisms could be harnessed to tune the extent and rate of vascularization in engineered tissue constructs.

Angiogenic patterning by STEEL, an endothelial-enriched long noncoding RNA.

Endothelial cell (EC)-enriched protein coding genes, such as endothelial nitric oxide synthase (eNOS), define quintessential EC-specific physiologic functions. It is not clear whether long noncoding RNAs (lncRNAs) also define cardiovascular cell type-specific phenotypes, especially in the vascular endothelium. Here, we report the existence of a set of EC-enriched lncRNAs and define a role for spliced-transcript endothelial-enriched lncRNA (STEEL) in angiogenic potential, macrovascular/microvascular identity, and shear stress responsiveness. STEEL is expressed from the terminus of the HOXD locus and is transcribed antisense to HOXD transcription factors. STEEL RNA increases the number and integrity of de novo perfused microvessels in an in vivo model and augments angiogenesis in vitro. The STEEL RNA is polyadenylated, nuclear enriched, and has microvascular predominance. Functionally, STEEL regulates a number of genes in diverse ECs. Of interest, STEEL up-regulates both eNOS and the transcription factor Kruppel-like factor 2 (KLF2), and is subject to feedback inhibition by both eNOS and shear-augmented KLF2. Mechanistically, STEEL up-regulation of eNOS and KLF2 is transcriptionally mediated, in part, via interaction of chromatin-associated STEEL with the poly-ADP ribosylase, PARP1. For instance, STEEL recruits PARP1 to the KLF2 promoter. This work identifies a role for EC-enriched lncRNAs in the phenotypic adaptation of ECs to both body position and hemodynamic forces and establishes a newer role for lncRNAs in the transcriptional regulation of EC identity.

Harnessing gene and drug delivery for vascularizing engineered tissue platforms.

Enhancement of tissue vascularization is a therapeutic target for many ischemic conditions, and is crucial for successful engraftment of therapeutic cells for tissue regeneration. The authors present opportunities for using these platforms for dissecting the role of angiogenic mechanisms and highlight recent gene and drug delivery strategies for enhancing vascularization of engineered tissues. Modular tissue engineering is featured as an example.

In vivo remodelling of vascularizing engineered tissues.

A critical aspect of creating vascularized tissues is the remodelling that occurs in vivo, driven in large part by the host response to the tissue construct. Rather than a simple inflammatory response, a beneficial tissue remodelling response results in the formation of vascularised tissue. The characteristics and dynamics of this response are slowly being elucidated, especially as they are modulated by the complex interaction between the biomaterial and cellular components of the tissue constructs and the host. This process has elements that are similar to both wound healing and tumour development, and its features are illustrated by reference to the bottom-up generation of a tissue using modular constructs. These modular constructs consist of mesenchymal stromal cells (MSC) embedded in endothelial cell (EC)-covered collagen gel rods that are a few hundred microns in size. Particular attention is paid to the role of hypoxia and macrophage recruitment, as well as the paracrine effects of the MSC and EC in this host response.

Tuning graft- and host-derived vascularization in modular tissue constructs: a potential role of HIF1 activation.

A better understanding of the factors governing the vascularization of engineered tissues is crucial for their advancement as therapeutic platforms. Here, we studied the effect of implant volume and cell densities on the in vivo vascularization of modular engineered tissue constructs. Sub-millimeter collagen modules containing adipose-derived mesenchymal stromal cells (adMSC) and enveloped by human umbilical vein endothelial cells (HUVEC) were subcutaneously implanted in severe-combined immunodeficient mice with a beige-mutation (SCID-bg) mice. Implant volume and cell density was varied relative to a base case, defined as a 0.01 mL implant containing 1.5×10(7) adMSC/mL and 3.9×10(6) HUVEC/mL. At 7 and 14 days post-transplantation, the constructs were harvested for immunohistochemical analysis of total (CD31(+)) and graft-derived (UEA1(+)) vessel formation, hypoxia-inducible factor 1-alpha (HIF1α) expression, infiltration of host-derived leukocytes (CD45), and macrophages (F4/80). Implant volume and cell density affected the relative contributions of host- versus graft-derived vascularization, highlighting that different mechanisms underlie the two processes. Graft-derived vessel formation was most rapid and robust in implants with high HIF1α expression, namely large volume implants and implants with high adMSC and HUVEC density (p<0.01 compared to base case at day 7). Many HIF1α(+) cells were vessel-lining HUVEC, suggesting that HIF1 activation may be key to vessel assembly in the graft. Host vessel ingrowth, however, dominated the vascularization of small volume implants (of high and low adMSC density alike), which showed low HIF1α expression at day 7. Host vessels were sustained to day 14 when adMSC density alone was increased, presumably due to increased paracrine secretions. This study points to a potential role of HIF1 activation in the vascularization of tissue constructs, which may be harnessed to engineer robust vessels for therapeutic applications.

Development of a smart garment to reduce kyphosis during daily living.

Many of the aches and pains of adults are the result of the long-term effects of bad posture or body misalignment. Postural kyphosis in adolescence, which is an excessive rounding of the upper spine, may be one of the effects of poor standing and sitting habits. A smart garment, consisting of a harness and two data-sensor loggers, was developed to monitor and provide vibration feedback to wearers to improve their posture during daily activities. Laboratory tests verified that the garment could provide an accuracy of 2 ± 2° during static measurement and 3 ± 2° during stable or slowly changing posture activities and 4 ± 4° during rapidly changing posture activities. Four volunteers wore the system for 3 h per day and for 4 consecutive days. The feedback was provided on the last 2 days and the kyphotic angle reduced by 8 ± 1° and 8 ± 2° on the last 2 days, respectively. Although the long-term effects of reminding the subjects' posture is still not clear, a short-term result shows promise that the smart garment may be able to improve the kyphosis.

Vertebral rotation measurement: a summary and comparison of common radiographic and CT methods.

Current research has provided a more comprehensive understanding of Adolescent Idiopathic Scoliosis (AIS) as a three-dimensional spinal deformity, encompassing both lateral and rotational components. Apart from quantifying curve severity using the Cobb angle, vertebral rotation has become increasingly prominent in the study of scoliosis. It demonstrates significance in both preoperative and postoperative assessment, providing better appreciation of the impact of bracing or surgical interventions. In the past, the need for computer resources, digitizers and custom software limited studies of rotation to research performed after a patient left the scoliosis clinic. With advanced technology, however, rotation measurements are now more feasible. While numerous vertebral rotation measurement methods have been developed and tested, thorough comparisons of these are still relatively unexplored. This review discusses the advantages and disadvantages of six common measurement techniques based on technology most pertinent in clinical settings: radiography (Cobb, Nash-Moe, Perdriolle and Stokes' method) and computer tomography (CT) imaging (Aaro-Dahlborn and Ho's method). Better insight into the clinical suitability of rotation measurement methods currently available is presented, along with a discussion of critical concerns that should be addressed in future studies and development of new methods.