Antonio Gigante

Chitosan, hyaluronan and chondroitin sulfate in tissue engineering for cartilage regeneration: A review

Injection of hyaluronan into osteoarthritic joints restores the viscoelasticity, augments the flow of joint fluid, normalizes endogenous hyaluronan synthesis, and improves joint function. Chitosan easily forms polyelectrolyte complexes with hyaluronan and chondroitin sulfate. Synergy of chitosan with hyaluronan develops enhanced performances in regenerating hyaline cartilage, typical results being structural integrity of the hyaline-like neocartilage, and reconstitution of the subchondral bone, with positive cartilage staining for collagen-II and GAG in the treated sites. Chitosan qualifies for the preparation of scaffolds intended for the regeneration of cartilage: it yields mesoporous cryogels; it provides a friendly environment for chondrocytes to propagate, produce typical ECM, and assume the convenient phenotype; it is a good carrier for growth factors; it inactivates metalloproteinases thus preventing collagen degradation; it is suitable for the induction of the chondrogenic differentiation of mesenchymal stem cells; it is a potent means for hemostasis and platelet delivery.

Chitosan stabilizes platelet growth factors and modulates stem cell differentiation toward tissue regeneration

The idea of using chitosan as a functional delivery aid to support simultaneously PRP, stem cells and growth factors (GF) is associated with the intention to use morphogenic biomaterials to modulate the natural healing sequence in bone and other tissues. For example, chitosan-chondroitin sulfate loaded with platelet lysate was included in a poly(D,L-lactate) foam that was then seeded with human adipose-derived stem cells and cultured in vitro under osteogenic stimulus: the platelet lysate provided to the bone tissue the most suitable assortment of GF which induces the osteogenic differentiation of the mesenchymal stem cells. PDGF, FGF, IGF and TGF-β were protagonists in the repair of callus fractures. The release of GF from the composites of chitosan-PRP and either nano-hydroxyapatite or tricalcium phosphate was highly beneficial for enhancing MSC proliferation and differentiation, thus qualifying chitosan as an excellent vehicle. A number of biochemical characteristics of chitosan exert synergism with stem cells in the regeneration of soft tissues.

The Effects of Patellar Taping on Patellofemoral Incongruence A Computed Tomography Study

Patellar taping for the purpose of patellar medialization is a nonoperative rehabilitation technique used in the treatment of patellofemoral pain. Despite early reports of excellent success rates, the indications for this treatment and its efficacy on patellofemoral pain are unclear. The present computed tomography study was undertaken to evaluate the effect of patellar taping on patellofemoral incongruence. Sixteen female patients (age range, 16 to 25 years) with anterior knee pain related to patellofemoral incongruence underwent computed tomography examination with their quadriceps muscles relaxed and contracted both before and after patellar taping. Patellar lateralization was measured as lateral patellar displacement, and patellar tilt was measured as lateral patellar angle. Patellar taping did not significantly affect patellofemoral lateralization or tilt. The results of this study do not support the use of this method for passive correction of patellofemoral incongruence. Although patellar taping may well be effective in controlling anterior knee pain during physical therapy, it does not do so by medializing the patella.

Membrane-seeded autologous chondrocytes: cell viability and characterization at surgery

The implantation of chondrocytes, seeded on matrices such as hyaluronic acid or collagen membranes, is a method that is being widely used for the treatment of chondral defects. The aim of the present study was to evaluate the distribution, viability and phenotype expression of the cells seeded on a collagen membrane just at the time of the implantation. Twelve patients who were suffering from articular cartilage lesions were treated by the MACI® procedure. The residual part of each membrane was tested by colorimetric assay (MTT) and histochemical and ultrastructural analyses were carried out. In all of the samples a large number of viable cells, quite homogenously distributed, was detected. The cells expressed the markers of the differentiated hyaline chondrocytes. These data reassure in that the MACI procedure provides a suitable engineered tissue for cartilage repair, in line with the clinical evidences emerging in the literature.

Arthroscopic Knee Cartilage Repair With Covered Microfracture and Bone Marrow Concentrate

In recent years several single-stage cartilage repair approaches have been devised to treat focal cartilage lesions. These usually associate microfracture (MFX) and a coverage scaffold. We describe a novel arthroscopic technique that combines MFX, autologous bone marrow concentrate (BMC), and a protective scaffold. Bone marrow aspirate from the iliac crest is centrifuged to obtain BMC. The cartilage defect is debrided, MFX holes are created, and the final defect is measured by use of a bent K-wire. The scaffold is then shaped to match the defect, immersed in BMC, introduced into the joint with a grasper, and fixed in place with a mixture of fibrin glue and BMC. This technique aims to augment the original single-stage procedure with a number of mesenchymal stem cells and growth factors contained in the BMC, to increase the defect filling and the rate of hyaline-like cartilage regeneration. The procedure combining MFX, BMC, and a protective scaffold is inexpensive and reproducible and has already shown the ability to regenerate hyaline-like cartilage. Its use as an alternative to autologous chondrocyte implantation requires further investigation.

Isolation of osteogenic progenitors from human amniotic fluid using a single step culture protocol

BackgroundStem cells isolated from amniotic fluid are known to be able to differentiate into different cells types, being thus considered as a potential tool for cellular therapy of different human diseases. In the present study, we report a novel single step protocol for the osteoblastic differentiation of human amniotic fluid cells.ResultsThe described protocol is able to provide osteoblastic cells producing nodules of calcium mineralization within 18 days from withdrawal of amniotic fluid samples. These cells display a complete expression of osteogenic markers (COL1, ONC, OPN, OCN, OPG, BSP, Runx2) within 30 days from withdrawal. In order to test the ability of these cells to proliferate on surfaces commonly used in oral osteointegrated implantology, we carried out cultures onto different test disks, namely smooth copper, machined titanium and Sandblasted and Acid Etching titanium (SLA titanium). Electron microscopy analysis evidenced the best cell growth on this latter surface.ConclusionThe described protocol provides an efficient and time-saving tool for the production of osteogenic cells from amniotic fluid that in the future could be used in oral osteointegrated implantology.

Genipin-Crosslinked Chitosan Gels and Scaffolds for Tissue Engineering and Regeneration of Cartilage and Bone.

The present review article intends to direct attention to the technological advances made since 2009 in the area of genipin-crosslinked chitosan (GEN-chitosan) hydrogels. After a concise introduction on the well recognized characteristics of medical grade chitosan and food grade genipin, the properties of GEN-chitosan obtained with a safe, spontaneous and irreversible chemical reaction, and the quality assessment of the gels are reviewed. The antibacterial activity of GEN-chitosan has been well assessed in the treatment of gastric infections supported by Helicobacter pylori. Therapies based on chitosan alginate crosslinked with genipin include stem cell transplantation, and development of contraction free biomaterials suitable for cartilage engineering. Collagen, gelatin and other proteins have been associated to said hydrogels in view of the regeneration of the cartilage. Viability and proliferation of fibroblasts were impressively enhanced upon addition of poly-l-lysine. The modulation of the osteocytes has been achieved in various ways by applying advanced technologies such as 3D-plotting and electrospinning of biomimetic scaffolds, with optional addition of nano hydroxyapatite to the formulations. A wealth of biotechnological advances and know-how has permitted reaching outstanding results in crucial areas such as cranio-facial surgery, orthopedics and dentistry. It is mandatory to use scaffolds fully characterized in terms of porosity, pore size, swelling, wettability, compressive strength, and degree of acetylation, if the osteogenic differentiation of human mesenchymal stem cells is sought: in fact, the novel characteristics imparted by GEN-chitosan must be simultaneously of physico-chemical and cytological nature. Owing to their high standard, the scientific publications dated 2010–2015 have met the expectations of an interdisciplinary audience.

Expression of NGF, Trka and p75 in human cartilage.

Nerve growth factor (NGF) exerts its action through two types of receptor: high-affinity tyrosine kinase A receptor (trkA) and low-affinity p75 receptor. NGF has a neurotrophic role in central and peripheral nervous system development, but there is also clear evidence of its involvement in the developing skeleton. The aim of the present immunohistochemical study was to investigate the expression and distribution of NGF, trkA, and p75 in normal cartilaginous tissues from adult subjects: articular and meniscal cartilage of the knee, cartilage from the epiglottis, and intervertebral disc tissue. Detection of NGF mRNA was also performed by in situ hybridization. Immunoreaction for NGF and the two receptors in articular chondrocytes, chondrocyte-like cells of meniscus and annulus fibrosus, and chondrocytes of the epiglottis demonstrated that they are all expressed in hyaline, fibrous and elastic cartilaginous tissues, suggesting that they could be involved in cartilage physio-pathology.

Adult mesenchymal stem cells for bone and cartilage engineering: effect of scaffold materials

Bone marrow is a useful cell source for skeletal tissue engineering approaches. In vitro differentiation of marrow mesenchymal stem cells (MSCs) to chondrocytes or osteoblasts can be induced by the addition of specific growth factors to the medium. The present study evaluated the behaviour of human MSCs cultured on various scaffolds to determine whether their differentiation can be induced by cell-matrix interactions. MSCs from bone marrow collected from the acetabulum during hip arthroplasty procedures were isolated by cell sorting, expanded and characterised by a flow cytometry system. Cells were grown on three different scaffolds (type I collagen, type I + II collagen and type I collagen + hydroxyapatite membranes) and analysed by histochemistry, immunohistochemistry and spectrophotometry (cell proliferation, alkaline phosphatase activity) at 15 and 30 days. Widely variable cell adhesion and proliferation was observed on the three scaffolds. MSCs grown on type I+II collagen differentiated to cells expressing chondrocyte markers, while those grown on type I collagen + hydroxyapatite differentiated into osteoblast-like cells. The study highlighted that human MSCs grown on different scaffold matrices may display different behaviours in terms of cell proliferation and phenotype expression without growth factor supplementation.

Subungual Exostosis of the Foot

Eleven subungual exostoses of the foot (10 on the hallux, 1 on the third toe) were studied. The initial symptom was subungual pain. When a subungual mass of fibrous tissue appeared, the nail was pushed up and in one case the mass became infected. X-rays exhibited a bone mass protruding from the terminal phalanx on the dorsomedial aspect of the toe in all cases. All patients underwent surgical excision of the lesions with partial onychectomy. Three layers were identified in five cases: a cap of fibrous tissue, a middle zone of hyaline cartilage with enchondral ossification, and a deep zone of cancellous bone. In three other cases, the histological pattern was pleomorphic and poorly characterized. The study shows that most subungual bone masses exhibited the pathological features of conventional osteochondromas. Nonetheless, a small number of lesions were pleomorphic and differed from osteochondromas, with abundant fibrous tissue merging irregularly into scattered islets of cartilage that was not organized in columns. Radical excision of the mass achieved complete relief of symptoms and recovery without recurrences in all cases.