Regenerative medicine: today's discoveries informing the future of medical practice

NPJ Regenerative Medicine 2016; 1:16007

Regenerative medicine: are we there yet?

NPJ Regen Med 2017 Jan 5; 2:2

Oncological safety of stromal vascular fraction enriched fat grafting in two-stage breast reconstruction after nipple sparing mastectomy: long-term results of a prospective study

OBJECTIVE: Autologous fat transfer (AFT) is commonly used to treat implant palpability and prevent fibrosis and thinning in mastectomy skin flaps. A major limit to this procedure is volume retention over time, leading to the introduction of fat enrichment with stromal vascular fraction (SVF+AFT). Oncological concerns have been raised over the injection of an increased concentration of progenitors cells (ASCs) in the SVF. The aim of the study is to evaluate the long-term cancer recurrence risk of SVF+AFT cases compared to AFT, in patients undergoing Nipple Sparing Mastectomy (NSM). PATIENTS AND METHODS: A prospective study was designed to compare three groups of patients undergoing NSM followed by SVF+AFT, AFT or none (control group), after a two-stage breast reconstruction. Patients were strictly followed-up for at least 5-years from the second stage reconstructive procedure. Loco-regional and systemic recurrence rate were evaluated over time as the primary outcome. Logistic regression was used to investigate which factors were associated with recurrence events and independent variables of interest were: surgical technique, age above 50 years old, lympho-vascular invasion, oncological stage, adjuvant or neoadjuvant chemotherapy, adjuvant radiotherapy and adjuvant hormone therapy. RESULTS: 41 women were included in G1 (SVF+AFT), 64 in G2 (AFT), and 64 in G3 (control group). Loco-regional recurrence rate was 2.4% for G1, 4.7% for G2, and 1.6% for G3. Systemic recurrence was 7.3%, 3.1%, and 3.1%, respectively. Among the variables included, there were no significant risk factors influencing a recurrence event, either loco-regional or systemic. In particular, SVF+AFT (G1) did not increase the oncological recurrence. CONCLUSIONS: Our data suggest that both centrifuged and SVF-enhanced fat transfer have a similar safety level in comparison to patients who did not undergo fat grafting in breast reconstruction after NSM

Amine functionalization of cholecyst-derived extracellular matrix with generation 1 PAMAM dendrimer

This document is the unedited author's version of a Submitted Work that was subsequently accepted for publication in Biomacromolecules, copyright © American Chemical Society after peer review. To access the final edited and published work, see http://pubs.acs.org/doi/pdf/10.1021/bm701055k.A method to functionalize cholecyst-derived extracellular matrix (CEM) with free amine groups was established in an attempt to improve its potential for tethering of bioactive molecules. CEM was incorporated with Generation-1 polyamidoamine (G1 PAMAM) dendrimer by using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide and N-hydroxysuccinimide cross-linking system. The nature of incorporation of PAMAM dendrimer was evaluated using shrink temperature measurements, Fourier transform infrared (FTIR) assessment, ninhydrin assay, and swellability. The effects of PAMAM incorporation on mechanical and degradation properties of CEM were evaluated using a uniaxial mechanical test and collagenase degradation assay, respectively. Ninhydrin assay and FTIR assessment confirmed the presence of increasing free amine groups with increasing quantity of PAMAM in dendrimer-incorporated CEM (DENCEM) scaffolds. The amount of dendrimer used was found to be critical in controlling scaffold degradation, shrink temperature, and free amine content. Cell culture studies showed that fibroblasts seeded on DENCEM maintained their metabolic activity and ability to proliferate in vitro. In addition, fluorescence cell staining and scanning electron microscopy analysis of cell-seeded DENCEM showed preservation of normal fibroblast morphology and phenotype

The use of porcine small intestinal submucosa mesh (SURGISIS) as a pelvic sling in a man and a woman with previous pelvic surgery: two case reports

<p>Abstract</p> <p>Introduction</p> <p>Closing the pelvic peritoneum to prevent the small bowel dropping into the pelvis after surgery for locally recurrent rectal cancer is important to prevent adhesions deep in the pelvis or complications of adjuvant radiotherapy. Achieving this could be difficult because sufficient native tissue is unavailable; we report on the use of small intestine submucosa extra-cellular matrix mesh in the obliteration of the pelvic brim.</p> <p>Case presentation</p> <p>We describe two cases in which submucosa extra-cellular matrix mesh was used to obliterate the pelvic brim following resection of a recurrent rectal tumour; the first patient, a 78-year-old Caucasian man, presented with small bowel obstruction caused by adhesions to a recurrent rectal tumour. The second patient, an 84-year-old Caucasian woman, presented with vaginal discharge caused by an entero-vaginal fistula due to a recurrent rectal tumour.</p> <p>Conclusion</p> <p>We report on the use of submucosa extra-cellular matrix mesh as a pelvic sling in cases where primary closure of the pelvic peritoneum is unfeasible. Its use had no infective complications and added minimal morbidity to the postoperative period. This is an original case report that would be of interest to general and colorectal surgeons.</p

Menisco-fibular ligament — an overview: cadaveric dissection, clinical and magnetic resonance imaging diagnosis, arthroscopic visualisation and treatment

Background: Injury to the menisco-fibular ligament (MFiL) is not commonly recognised. The anatomy of the lateral meniscus is complex and structure-function relationships are only partly understood. The purpose of the present study was to evaluate the MFiL, an anatomic structure rarely discussed that stabilises the lateral meniscus at the level of the hiatus popliteus and may have a crucial role in pathology of lateral meniscus injury.Materials and methods: The MFiL was dissected from its attachment at the lateral meniscus to its insertion on fibular head in 12 human normal cadaver knees. The dimensions were determined and its anatomic position visualised throughout a 90° range of motion. Findings were documented on digital photographs and on video. Results were compared against the magnetic resonance imaging (MRI) appearance of the injured MFiL in 20 patients. Concomitant knee injuries in those patients were also analysed to determine the most frequent pattern of injuries.Results: The normal MFiL showed an inverted trapezoid-shape with a mean width proximally of 13 mm, mean width distally of 8.5 mm and a mean length of 18.4 mm. MRI visualisation of the ligament was possible even in regular sequences; however, additional radial plane sequences were also used. Arthroscopic visualisation and manipulation was optimal when the camera was inserted into the postero-lateral gutter with full knee extension.Conclusions: The MFiL stabilises the postero-lateral knee in concert with the menisco-femoral ligaments. Injury to the MFiL can be a cause of chronic postero-lateral pain syndrome with associated instability. Further anatomical and biomechanical studies are needed in order to fully evaluate its importance

Extracellular matrix hydrogels from decellularized tissues: structure and function

Extracellular matrix (ECM) bioscaffolds prepared from decellularized tissues have been used to facilitate constructive and functional tissue remodeling in a variety of clinical applications. The discovery that these ECM materials could be solubilized and subsequently manipulated to form hydrogels expanded their potential in vitro and in vivo utility; i.e. as culture substrates comparable to collagen or Matrigel, and as injectable materials that fill irregularly-shaped defects. The mechanisms by which ECM hydrogels direct cell behavior and influence remodeling outcomes are only partially understood, but likely include structural and biological signals retained from the native source tissue. The present review describes the utility, formation, and physical and biological characterization of ECM hydrogels. Two examples of clinical application are presented to demonstrate in vivo utility of ECM hydrogels in different organ systems. Finally, new research directions and clinical translation of ECM hydrogels are discusse

Immune enhancement by novel vaccine adjuvants in autoimmune-prone NZB/W F1 mice: relative efficacy and safety

<p>Abstract</p> <p>Background</p> <p>Vaccines have profoundly impacted global health although concerns persist about their potential role in autoimmune or other adverse reactions. To address these concerns, vaccine components like immunogens and adjuvants require critical evaluation not only in healthy subjects but also in those genetically averse to vaccine constituents. Evaluation in autoimmune-prone animal models of adjuvants is therefore important in vaccine development. The objective here was to assess the effectiveness of experimental adjuvants: two phytol-derived immunostimulants PHIS-01 (phytanol) and PHIS-03 (phytanyl mannose), and a new commercial adjuvant from porcine small intestinal submucosa (SIS-H), relative to a standard adjuvant alum. Phytol derivatives are hydrophobic, oil-in water diterpenoids, while alum is hydrophilic, and SIS is essentially a biodegradable and collagenous protein cocktail derived from extracellular matrices.</p> <p>Results</p> <p>We studied phthalate -specific and cross-reactive anti-DNA antibody responses, and parameters associated with the onset of autoimmune disorders. We determined antibody isotype and cytokine/chemokine milieu induced by the above experimental adjuvants relative to alum. Our results indicated that the phytol-derived adjuvant PHIS-01 exceeded alum in enhancing anti-phthalate antibody without much cross reactivity with ds-DNA. Relatively, SIS and PHIS-03 proved less robust, but they were also less inflammatory. Interestingly, these adjuvants facilitated isotype switching of anti-hapten, but not of anti-DNA response. The current study reaffirms our earlier reports on adjuvanticity of phytol compounds and SIS-H in non autoimmune-prone BALB/c and C57BL/6 mice. These adjuvants are as effective as alum also in autoimmune-prone NZB/WF1 mice, and they have little deleterious effects.</p> <p>Conclusion</p> <p>Although all adjuvants tested impacted cytokine/chemokine milieu in favor of Th1/Th2 balance, the phytol compounds fared better in reducing the onset of autoimmune syndromes. However, SIS is least inflammatory among the adjuvants evaluated.</p

Solution fibre spinning technique for the fabrication of tuneable decellularised matrix-laden fibres and fibrous micromembranes.

UNLABELLED: Recreating tissue-specific microenvironments of the extracellular matrix (ECM) in vitro is of broad interest for the fields of tissue engineering and organ-on-a-chip. Here, we present biofunctional ECM protein fibres and suspended membranes, with tuneable biochemical, mechanical and topographical properties. This soft and entirely biologic membrane scaffold, formed by micro-nano-fibres using low voltage electrospinning, displays three unique characteristics for potential cell culture applications: high-content of key ECM proteins, single-layered mesh membrane, and flexibility for in situ integration into a range of device setups. Extracellular matrix (ECM) powder derived from urinary bladder, was used to fabricate the ECM-laden fibres and membranes. The highest ECM concentration in the dry protein fibre was 50 wt%, with the rest consisting of gelatin. Key ECM proteins, including collagen IV, laminin, and fibronectin, were shown to be preserved post the biofabrication process. The single fibre tensile Young's modulus can be tuned for over two orders of magnitude between ∼600 kPa and 50 MPa depending on the ECM content. Combining the fibre mesh printing with 3D printed or microfabricated structures, culture devices were constructed for endothelial layer formation, and a trans-membrane co-culture formed by glomerular cell types of podocytes and glomerular endothelial cells, demonstrating feasibility of the membrane culture. Our cell culture observation points to the importance of membrane mechanical property and re-modelling ability as a factor for soft membrane-based cell cultures. The ECM-laden fibres and membranes presented here would see potential applications in in vitro assays, and tailoring structure and biological functions of tissue engineering scaffolds. STATEMENT OF SIGNIFICANCE: Recreating tissue-specific microenvironments of the extracellular matrix (ECM) is of broad interest for the fields of tissue engineering and organ-on-a-chip. Both the biochemical and biophysical signatures of the engineered ECM interplay to affect cell response. Currently, there are limited biomaterials processing methods which allow to design ECM membrane properties flexibly and rapidly. Solvents and additives used in many existing processes also induced unwanted ECM protein degradation and toxic residues. This paper presents a solution fibre spinning technique, where careful selection of the solution combination led to well-preserved ECM proteins with tuneable composition. This technique also provides a highly versatile approach to fabricate ECM fibres and membranes, leading to designable fibre Young's modulus for over two orders of magnitude.This work is supported by the Engineering and Physical Sciences Research Council (EPSRC) UK (EP/M018989/1) and European Research Council (ERC-StG, 758865). The authors thank the studentship and funding supports from the EPSRC DTA (Z.L.), the WD Armstrong Trust (I.M.L), the Swiss National Science Foundation (P300P2_171219) and the Centre for Misfolding Disease of the University of Cambridge (F.S.R.)

Extracellular Matrix Aggregates from Differentiating Embryoid Bodies as a Scaffold to Support ESC Proliferation and Differentiation

Embryonic stem cells (ESCs) have emerged as potential cell sources for tissue engineering and regeneration owing to its virtually unlimited replicative capacity and the potential to differentiate into a variety of cell types. Current differentiation strategies primarily involve various growth factor/inducer/repressor concoctions with less emphasis on the substrate. Developing biomaterials to promote stem cell proliferation and differentiation could aid in the realization of this goal. Extracellular matrix (ECM) components are important physiological regulators, and can provide cues to direct ESC expansion and differentiation. ECM undergoes constant remodeling with surrounding cells to accommodate specific developmental event. In this study, using ESC derived aggregates called embryoid bodies (EB) as a model, we characterized the biological nature of ECM in EB after exposure to different treatments: spontaneously differentiated and retinoic acid treated (denoted as SPT and RA, respectively). Next, we extracted this treatment-specific ECM by detergent decellularization methods (Triton X-100, DOC and SDS are compared). The resulting EB ECM scaffolds were seeded with undifferentiated ESCs using a novel cell seeding strategy, and the behavior of ESCs was studied. Our results showed that the optimized protocol efficiently removes cells while retaining crucial ECM and biochemical components. Decellularized ECM from SPT EB gave rise to a more favorable microenvironment for promoting ESC attachment, proliferation, and early differentiation, compared to native EB and decellularized ECM from RA EB. These findings suggest that various treatment conditions allow the formulation of unique ESC-ECM derived scaffolds to enhance ESC bioactivities, including proliferation and differentiation for tissue regeneration applications. © 2013 Goh et al