Establishment of tooth blood supply and innervation is developmentally regulated and takes place through differential patterning processes

Teeth are richly supported by blood vessels and peripheral nerves. The aim of this study was to describe in detail the developmental time‐course and localization of blood vessels during early tooth formation and to compare that to innervation, as well as to address the putative role of vascular endothelial growth factor (VEGF), which is an essential regulator of vasculature development, in this process. The localization of blood vessels and neurites was compared using double immunofluorescence staining on sections at consecutive stages of the embryonic (E) and postnatal (PN) mandibular first molar tooth germ (E11‐PN7). Cellular mRNA expression domains of VEGF and its signaling receptor VEGFR2 were studied using sectional radioactive in situ hybridization. Expression of VEGF mRNA and the encoded protein were studied by RT‐PCR and western blot analysis, respectively, in the cap and early bell stage tooth germs, respectively. VEGFR2 was immunolocalized on tooth tissue sections. Smooth muscle cells were investigated by anti‐alpha smooth muscle actin (αSMA) antibodies. VEGF showed developmentally regulated epithelial and mesenchymal mRNA expression domains including the enamel knot signaling centers that correlated with the growth and navigation of the blood vessels expressing Vegfr2 and VEGFR2 to the dental papilla and enamel organ. Developing blood vessels were present in the jaw mesenchyme including the presumptive dental mesenchyme before the appearance of the epithelial dental placode and dental neurites. Similarly, formation of a blood vessel plexus around the bud stage tooth germ and ingrowth of vessels into dental papilla at E14 preceded ingrowth of neurites. Subsequently, pioneer blood vessels in the dental papilla started to receive smooth muscle coverage at the early embryonic bell stage. Establishment and patterning of the blood vessels and nerves during tooth formation are developmentally regulated, stepwise processes that likely involve differential patterning mechanisms. Development of tooth vascular supply is proposed to be regulated by local, tooth‐specific regulation by epithelial–mesenchymal tissue interactions and involving tooth target expressed VEGF signaling. Further investigations on tooth vascular development by local VEGF signaling, as well as how tooth innervation and development of blood vessels are integrated with advancing tooth organ formation by local signaling mechanisms, are warranted.     

High‐throughput fabrication of vascularized adipose microtissues for 3D bioprinting

For patients with soft tissue defects, repair with autologous in vitro engineered adipose tissue could be a promising alternative to current surgical therapies. A volume‐persistent engineered adipose tissue construct under in vivo conditions can only be achieved by early vascularization after transplantation. The combination of 3D bioprinting technology with self‐assembling microvascularized units as building blocks can potentially answer the need for a microvascular network. In the present study, co‐culture spheroids combining adipose‐derived stem cells (ASC) and human umbilical vein endothelial cells (HUVEC) were created with an ideal geometry for bioprinting. When applying the favourable seeding technique and condition, compact viable spheroids were obtained, demonstrating high adipogenic differentiation and capillary‐like network formation after 7 and 14 days of culture, as shown by live/dead analysis, immunohistochemistry and RT‐qPCR. Moreover, we were able to successfully 3D bioprint the encapsulated spheroids, resulting in compact viable spheroids presenting capillary‐like structures, lipid droplets and spheroid outgrowth after 14 days of culture. This is the first study that generates viable high‐throughput (pre‐)vascularized adipose microtissues as building blocks for bioprinting applications using a novel ASC/HUVEC co‐culture spheroid model, which enables both adipogenic differentiation while simultaneously supporting the formation of prevascular‐like structures within engineered tissues in vitro.     

组织工程技术在尿道重建中的应用

背景：多种疾病或创伤可导致尿道组织的缺损或狭窄,腔黏膜,或者其他自体移植物修复会带来较多并发症,治疗思路。需用其他组织来修复与重建。传统应用包皮、口而组织工程的兴起为尿道修复与重建开辟了新的目的：总结并探讨组织工程支架材料的选择、种子细胞的选择,以及目前组织工程在尿道修复中的临床应用情况。方法：由第一作者应用计算机检索PubMed数据库及万方全文数据库1994年1月至2012年5月有关尿道组织工程的文章,英文检索词为“Tissueengineering,urethra,scaffoldmaterials,stemcells,Vascularization”,中文检索词为“组织工程;尿道;支架材料;干细胞”。排除重复性研究,共保留57篇文献进行综述。选择文章内容主要为组织工程支架在尿道修复中的应用情况,同一领域文献则选择近期发表或发表在权威杂志文章。结果与结论：目前组织工程研究的主要目标仍旧是以下几点：选择最佳支架、最佳干细胞来源,促进移植物新生血管和神经长入。泌尿系统组织工程已经报道,在临床上,使用复合或者不复合种子细胞的脱细胞支架可以重建尿道。虽然在大规模应用于临床前,还需要进一步的研究和创新,有理由期待组织工程会在尿道疾病中发挥越来越大的作用。     

Validation of Musculoskeletal Ultrasound in the Assessment of Experimental Gout Synovitis

The objective of this study was to validate musculoskeletal ultrasound (US) in a rabbit model of acute gout. Acute gout was induced by intra-articular injection of monosodium urate (MSU) crystals in 10 rabbits; the 3 controls received vehicle. Rabbit knees were assessed by B-mode and power Doppler (PD) US 24 and 72?h after injections. After 72?h, all rabbits were euthanized. US discriminated between the MSU-injected and control groups with respect to the different inflammatory findings at both at 24 and 72?h and for MSU crystal-related findings after 24?h of injection. US synovial thickening, intra-synovial power Doppler signal and global joint distension significantly correlated with the synovial global histopathological score (r?=?0.47, p?=?0.0188), tissue vascularization measured by CD31 immunohistochemical-positive staining (r?=?0.46, p?=?0.0172) and tissue levels of interleukin-1β (r?=?0.53, p?=?0.0078), respectively. US is a valid method for assessment of synovial inflammation in experimental gouty arthritis in rabbits.     

Review of vascularised bone tissue‐engineering strategies with a focus on co‐culture systems

Poor angiogenesis within tissue‐engineered grafts has been identified as a main challenge limiting the clinical introduction of bone tissue‐engineering (BTE) approaches for the repair of large bone defects. Thick BTE grafts often exhibit poor cellular viability particularly at the core, leading to graft failure and lack of integration with host tissues. Various BTE approaches have been explored for improving vascularisation in tissue‐engineered constructs and are briefly discussed in this review. Recent investigations relating to co‐culture systems of endothelial and osteoblast‐like cells have shown evidence of BTE efficacy in increasing vascularization in thick constructs. This review provides an overview of key concepts related to bone formation and then focuses on the current state of engineered vascularized co‐culture systems using bone repair as a model. It will also address key questions regarding the generation of clinically relevant vascularized bone constructs as well as potential directions and considerations for research with the objective of pursuing engineered co‐culture systems in other disciplines of vascularized regenerative medicine. The final objective is to generate serious and functional long‐lasting vessels for sustainable angiogenesis that will enable enhanced cellular survival within thick voluminous bone grafts, thereby aiding in bone formation and remodelling in the long term. However, more evidence about the quality of blood vessels formed and its associated functional improvement in bone formation as well as a mechanistic understanding of their interactions are necessary for designing better therapeutic strategies for translation to clinical settings. Copyright © 2012 John Wiley & Sons, Ltd.     

Visualization of vascular ultrastructure during osteogenesis by tissue engineering technique

The aim of this paper was to observe and visualize the changes in osteoblasts by electron microscopy during osteogenesis using tissue engineering technique. We also studied the feasibility of improving tissue vascularization of the engineered bone by using small intestine submucosa (SIS) as the scaffold. Bone mesenchymal stem cells (BMSCs) were isolated by gradient centrifugation method. Bone mesenchymal stem cells were seeded in the SIS, and the scaffold-cell constructs were cultured in vitro for 2 weeks. Small intestine submucosa without BMSCs served as control. Both SIS scaffolds were then implanted subcutaneously in the dorsa of athymic mice. The implants were harvested after in vivo incubation for 4, 8 and 12 weeks. The changes in osteoblasts and vascularization were observed under a transmission electron microscope and a scanning electron microscope. The BMSCs grew quite well, differentiating on the surface of the SIS and secreting a great deal of extracellular matrices. The scaffold-cell constructs formed a lot of bone and blood vessels in vivo. The scaffold degraded after 12 weeks. No osteoblasts, but vascularization and fibroblasts were observed, in the control. The SIS can be used as a scaffold for constructing tissue-engineered bone as it can improve the formation of bone and vessels in vivo. Translated from Journal of Shanghai Jiaotong University (Medical Science), 2006, 26(2): 113–116 [译自: 上海交通大学学报 (医学版)]     

Prevalence of Achilles and patellar tendinopathy and their association to intratendinous changes in adolescent athletes

Achilles (AT) and patellar tendons (PT) are commonly affected by tendinopathy in adult athletes but prevalence of symptoms and morphological changes in adolescents is unclear. The study aimed to determine prevalence of tendinopathy and intratendinous changes in ATs and PTs of adolescent athletes. A total of 760 adolescent athletes (13.0 ± 1.9 years; 160 ± 13 cm; 50 ± 14 kg) were examined. History, local clinical examination, and longitudinal Doppler ultrasound analysis for both ATs and PTs were performed including identification of intratendinous echoic changes and vascularization. Diagnosis of tendinopathy was complied clinically in case of positive history of tendon pain and tendon pain on palpation. Achilles tendinopathy was diagnosed in 1.8% and patellar tendinopathy in 5.8%. Vascularizations were visible in 3.0% of ATs and 11.4% of PTs, hypoechogenicities in 0.7% and 3.2% as well as hyperechogenicities in 0% and 0.3%, respectively. Vascularizations and hypoechogenicities were statistically significantly more often in males than in females (P ≤ 0.02). Subjects with patellar tendinopathy had higher prevalence of structural intratendinous changes than those without PT symptoms (P ≤ 0.001). In adolescent athletes, patellar tendinopathy is three times more frequent compared with Achilles tendinopathy. Longitudinal studies are necessary to investigate physiological or pathological origin of vascularizations and its predictive value in development of tendinopathy.     

Tumour angiogenesis and its relation to contrast enhancement on computed tomography: a review

Angiogenesis describes the formation of new blood vessels within tumours. The process is essential for tumour growth and metastasis. The development of new vessels leads to physiological changes, specifically increased perfusion, blood volume and capillary permeability, that alter contrast enhancement during computed tomography (CT). Functional CT techniques that quantify these physiological changes can provide greater insight into how angiogenesis alters contrast enhancement in routine practice and also serve as diagnostic tools in their own right. The functional information obtained can aid with tissue characterisation, such as type or grade of tumour, improve the detection of hepatic metastases, produce clearer delineation of tumours with benefits for radiotherapy planning and biopsy, and provide prognostic information. By providing a marker for tumour angiogenesis, quantitative contrast enhanced CT can improve the diagnostic assessment of patients with cancer.     

A novel image processing workflow for the in vivo quantification of skin microvasculature using dynamic optical coherence tomography

Background

Currently, imaging technologies that can accurately assess or provide surrogate markers of the human cutaneous microvessel network are limited. Dynamic optical coherence tomography (D‐OCT) allows the detection of blood flow in vivo and visualization of the skin microvasculature. However, image processing is necessary to correct images, filter artifacts, and exclude irrelevant signals. The objective of this study was to develop a novel image processing workflow to enhance the technical capabilities of D‐OCT.

Materials and methods

Single‐center, vehicle‐controlled study including healthy volunteers aged 18‐50 years. A capsaicin solution was applied topically on the subject's forearm to induce local inflammation. Measurements of capsaicin‐induced increase in dermal blood flow, within the region of interest, were performed by laser Doppler imaging (LDI) (reference method) and D‐OCT.

Results

Sixteen subjects were enrolled. A good correlation was shown between D‐OCT and LDI, using the image processing workflow. Therefore, D‐OCT offers an easy‐to‐use alternative to LDI, with good repeatability, new robust morphological features (dermal‐epidermal junction localization), and quantification of the distribution of vessel size and changes in this distribution induced by capsaicin. The visualization of the vessel network was improved through bloc filtering and artifact removal. Moreover, the assessment of vessel size distribution allows a fine analysis of the vascular patterns.

Conclusion

The newly developed image processing workflow enhances the technical capabilities of D‐OCT for the accurate detection and characterization of microcirculation in the skin. A direct clinical application of this image processing workflow is the quantification of the effect of topical treatment on skin vascularization.