Hyaluronan based porous nano-particles enriched with growth factors for the treatment of ulcers: a placebo-controlled study

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Mechanical influence of tissue culture plates and extracellular matrix on mesenchymal stem cell behavior: A topical review

Tissue engineering applications need a continuous development of new biomaterials able to generate an ideal cell-extracellular matrix interaction. The stem cell fate is regulated by several factors, such as growth factors or transcription factors. The most recent literature has reported several publications able to demonstrate that environmental factors also contribute to the regulation of stem cell behavior, leading to the opinion that the environment plays the major role in the cell differentiation. The interaction between mesenchymal stem cells (MSCs) and extracellular environment has been widely described, and it has a crucial role in regulating the cell phenotype. In our laboratory (Tecnologica Research Institute, Crotone, Italy), we have recently studied how several physical factors influence the distribution and the morphology of MSCs isolated from dental pulp, and how they are able to regulate stem cell differentiation. Mechanical and geometrical factors are only a small part of the environmental factors able to influence stem cell behavior, however, this influence should be properly known: in fact, this assumption must be clearly considered during those studies involving MSCs; furthermore, these interactions should be considered as an important bias that involves an high number of studies on the MSCs, since in worldwide laboratories the scientists mostly use tissue culture plates for their experiments

In vitro and in vivo biological performance of modified gellan gum-based hydrogels for nucleus pulposus tissue engineering

Ionic- (iGG-MA) and photo-crosslinked (phGG-MA) methacrylated gellan gum hydrogels have been proposed as biomaterials for supporting nucleus pulposus (NP) regeneration and/or repair. In this study, the mechanical stability and biocompatibility of these hydrogels have been evaluated in vitro. Human intervertebral disc cells obtained from herniated patients were cultured within both hydrogels, for 1–21 days. Dynamic mechanical analysis and biological characterization (Live/ Dead assay, ATP and DNA quantification, PCR and immunocytochemistry) were performed after specific times of culturing. The in vitro study showed that both cell loading and culturing time do not affect the mechanical properties of hydrogels. In addition, the iGG-MA and phGG-MA hydrogels showed to be effective on supporting cells encapsulation and viability up to 21 days of culturing. In vivo biocompatibility screening was also performed, by subcutaneous implantation of both hydrogels in Lewis rats for the period of 10 and 18 days. Haematoxylin & eosin staining revealed that the hydrogels do not elicit necrosis, calcification or acute inflammatory reaction. The present study demonstrates that the iGG-MA and phGG-MA hydrogels support cells encapsulation and viability, and are well-tolerated, stable and non-cytotoxic in vitro and in vivo, thus possessing promising features for finding application as viable NP substitutes

Mechanical performance and biocompatibility study of methacrylated Gellan gum hydrogels with potential for nucleus pulposus regeneration

Methacrylated gellan gum hydrogels, obtained either by ionic- (iGGMA) and photo-crosslinking (phGG-MA), have been investigated as potential biomaterials for supporting nucleus pulposus (NP) regeneration and/or repair [1,2]. In previous work, some advantages were attributed to GG-MA hydrogels, such as: (i) the possibility to control endothelial cells infiltration and blood vessel ingrowth’s, (ii) tunable and improved mechanical properties, and (iii) in situ gelation, within seconds to few minutes. In this study, the mechanical and biological performance of these hydrogels was firstly evaluated in vitro. Human intervertebral disc (hIVD) cells obtained from herniated patients were cultured within both hydrogels, for 1 up to 21 days. Dynamic mechanical analysis and biological characterization (calcein-AM staining, ATP and DNA quantification and PCR) were performed after specific times of culturing. A biocompatibility study was also performed in vivo, by subcutaneous implantation of acellular iGG-MA and phGG-MA hydrogels in Lewis rats for the period of 10 and 18 days. Tissue response to the hydrogels implantation was determined by histological analysis (haematoxylin-eosin staining). The in vitro study showed that both cell loading and culturing time do not have an effect on the mechanical properties of the hydrogels. Regarding their biological performance, the iGG-MA and phGG-MA hydrogels showed to be effective on supporting hIVD cells encapsulation and viability up to 21 days of culturing. Human IVD cells were homogeneously distributed within the hydrogels and maintained its round-shape morphology during culturing time. The in vivo biocompatibility study showed that iGG-MA and phGG-MA hydrogels do not elicit any deleterious effect, as denoted by the absence of necrosis and calcification, or acute inflammatory reaction. A thin fibrous capsule was observed around the implanted hydrogels. The results presented in this study indicate that the iGG-MA and phGG-MA hydrogels are stable in vitro and in vivo, support hIVD cells encapsulation and viability, and were found to be well-tolerated and non-cytotoxic in vivo, thus being potential candidates for NP regeneration

Adipose micro-grafts enhance tendinopathy healing in ovine model: An in vivo experimental perspective study

In Europe, approximatively 100 000 to 500 000 tendon repairs are performed every year. These procedures are associated with a considerable rate of postoperative complications (from 6% to 11%). Autologous micro-grafts (AAMG) and stromal vascular fraction (SVF) have been shown to improve tendon healing in 60% to 70% of treated rodents. The purpose of this study was to evaluate the effects of AAMG in a sheep model with tendinopathy. We used sheep models because, as a large animal, they are more comparable to humans. The hypothesis was that SVF injection would improve tendon healing compared with the control group, reducing inflammatory and matrix degrading, while increasing anti-inflammatory expression and collagen synthesis in the early stage of tendon injury. Sixteen Apennine sheep aged 2 to 5 years underwent 500 UI type I collagenase injection into both common calcaneal tendons (CCT) to induce tendinopathy. After 15 days (T0), one CCT in every ovine underwent randomly to 2.5 mL of AAMG obtained by mechanical disruption and the contralateral CCTs received no treatment. Clinical, ecographic, and sonographic evaluations were performed after 4 weeks (T1) and 8 weeks (T2). Histological, immunohistochemical, real-time polymerase chain reaction (RT-PCR), and biomechanical evaluations were performed at T2. At T2, the treated group showed a final tendon diameter (9.1 ± 1.4 mm) and a hardness expression (62%) that were similar to the original healthy tendon (8.1 ± 1.1 mm; 100%), with a significant recovery compared with the control group (9.5 ± 1.7 mm; 39%). Moreover, histological analysis of the treated group revealed an improvement in the fiber orientation score, fiber edema score, infiltrative-inflammatory process, and necrosis score (4.3 ± 3.3) compared with control group (8.8 ± 2.9). Immunohistochemically, the treated group showed high expression of collagen 1, Factor VIII and significantly low expression of collagen 3. These data were confirmed by RT-PCR analysis. The study findings suggested that AAMGs obtained through mechanical disruption present a safe, efficient, and reliable technique, enhancing tendon healing

In vitro concurrent endothelial and osteogenic commitment of adipose-derived stem cells and their genomical analyses through CGH array: novel strategies to increase the succesfull engraftement of a tissue engineered bone grafts

In the field of tissue engineering, adult stem cells are increasingly recognized as an important tool for in vitro reconstructed tissue-engineered grafts. In the world of cell therapies, mesenchymal stem cells from bone marrow or adipose tissue are undoubtedly the most promising progenitors for tissue engineering applications. In this setting, adipose-derived stem cells (ASC) are generally similar to those derived from bone marrow and are most conveniently extracted from tissue removed in elective cosmetic liposuction procedures; they also show a great potential for endothelization. The aim of the present work was to investigate how the co-commitment into a vascular and bone phenotype of ASC could be a usefull tools for improving the in vitro and in vivo reconstruction of a vascularized bone graft. Human ASC obtained from abdominoplasty procedures were loaded in a hydroxyapatite clinical-grade scaffold, co-differentiated and tested for proliferation, cell distribution, and osteogenic and vasculogenic gene expression. The chromosomal stability of the cultures was investigated using the CGH array for 3D cultures. ASC adhesion, distribution, proliferation and gene expression not only demonstrated a full osteogenic and vasculogenic commitment in vitro and in vivo, but also showed that endothelization strongly improves their osteogenic commitment. In the end, genetic analyses confirmed that no genomical alteration in long-term in vitro culture of ASC in 3D scaffolds occurs

Nestin-GFP Transgene Reveals Neural Precursor Cells in Adult Skeletal Muscle

Background: Therapy for neural lesions or degenerative diseases relies mainly on finding transplantable active precursor cells. Identifying them in peripheral tissues accessible for biopsy, outside the central nervous system, would circumvent the serious immunological and ethical concerns impeding cell therapy. Methodology/Principal Findings: In this study, we isolated neural progenitor cells in cultured adult skeletal muscle from transgenic mice in which nestin regulatory elements control GFP expression. These cells also expressed the early neural marker Tuj1 and light and heavy neurofilament but not S100b, indicating that they express typical neural but not Schwann cell markers. GFP+/Tuj1+ cells were also negative for the endothelial and pericyte markers CD31 and a-smooth muscle actin, respectively. We established their a) functional response to glutamate in patch-clamp recordings; b) interstitial mesenchymal origin; c) replicative capacity; and d) the environment necessary for their survival after fluorescenceactivated cell sorting. Conclusions/Significance: We propose that the decline in nestin-GFP expression in muscle progenitor cells and its persistence in neural precursor cells in muscle cultures provide an invaluable tool for isolating a population of predifferentiated neural cells with therapeutic potential

Transcriptomics Comparison between Porcine Adipose and Bone Marrow Mesenchymal Stem Cells during In Vitro Osteogenic and Adipogenic Differentiation

Bone-marrow mesenchymal stem cells (BMSC) are considered the gold standard for use in tissue regeneration among mesenchymal stem cells (MSC). The abundance and ease of harvest make the adipose-derived stem cells (ASC) an attractive alternative to BMSC. The aim of the present study was to compare the transcriptome of ASC and BMSC, respectively isolated from subcutaneous adipose tissue and femur of 3 adult pigs, during in vitro osteogenic and adipogenic differentiation for up to four weeks. At 0, 2, 7, and 21 days of differentiation RNA was extracted for microarray analysis. A False Discovery Rate ≤0.05 for overall interactions effect and P<0.001 between comparisons were used to determine differentially expressed genes (DEG). Ingenuity Pathway Analysis and DAVID performed the functional analysis of the DEG. Functional analysis of highest expressed genes in MSC and genes more expressed in MSC vs. fully differentiated tissues indicated low immunity and high angiogenic capacity. Only 64 genes were differentially expressed between ASC and BMSC before differentiation. The functional analysis uncovered a potential larger angiogenic, osteogenic, migration, and neurogenic capacity in BMSC and myogenic capacity in ASC. Less than 200 DEG were uncovered between ASC and BMSC during differentiation. Functional analysis also revealed an overall greater lipid metabolism in ASC, while BMSC had a greater cell growth and proliferation. The time course transcriptomic comparison between differentiation types uncovered <500 DEG necessary to determine cell fate. The functional analysis indicated that osteogenesis had a larger cell proliferation and cytoskeleton organization with a crucial role of G-proteins. Adipogenesis was driven by PPAR signaling and had greater angiogenesis, lipid metabolism, migration, and tumorigenesis capacity. Overall the data indicated that the transcriptome of the two MSC is relatively similar across the conditions studied. In addition, functional analysis data might indicate differences in therapeutic application

Sexual Functioning and Opioid Maintenance Treatment in Women. Results From a Large Multicentre Study

Opioid maintenance treatment (OMT) is the most widespread therapy for both females and males opioid addicts. While many studies have evaluated the OMT impact on men’s sexuality, the data collected about the change in women’s sexual functioning is still limited despite the fact that it is now well-known that opioids - both endogenous and exogenous - affect the endocrine system and play an important role in sexual functioning. The present study aims to determine how OMT with buprenorphine (BUP) or methadone (MTD) affects sexual health in women; examining also any possible emerging correlation between sexual dysfunction (SD), type of opioid and patients’ mental health. This multi-center study case recruited 258 female volunteers attending Italian public Addiction Outpatients Centers that were stabilized with OMT for at least 3 months. SD was assessed with the Arizona Sexual Experience Scale. The twelve-item General Health Questionnaire was used to assess participants’ mental health conditions. The results show that 56.6% of women receiving OMT for at least 3 months presented SD without significant differences between MTD e BUP groups. The majority of the subjects with SD have a poorer quality of intimate relationships and worse mental health than the average. To the best of our knowledge, the present study is the largest report on the presence of SDs in women as a side effects of MTD and BUP used in OMT. Since SDs cause difficulties in intimate relationships, lower patients’ quality of life and interfere with OMT beneficial outcomes, we recommend that women undertaking an opioid therapy have routine screening for SD and we highlight the importance to better examine opioid-endocrine interactions in future studies in order to provide alternative potential treatments such as the choice of opioid, opioid dose reduction and hormone supplementation