Neural Crest-Derived Chondrocytes Isolation for Tissue Engineering in Regenerative Medicine

Salamone, Monica; Rigogliuso, Salvatrice; Nicosia, Aldo; Tagliavia, Marcello; Campora, Simona; Cinà, Paolo; Bruno, Carmelo; Ghersi, Giulio

doi:10.3390/cells9040962

Cited by 9 publications

(7 citation statements)

References 32 publications

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“…In order to confirm such assumption and evaluate the differentiated state after culturing in different conditions, the expression pattern of some cartilage-specific genes, including Sox9 , Col2A1 and aggrecan ( Acan ) as well as Col1A2 as a proxy of fibrous differentiation [ 51 ] was evaluated and compared with that observed from cells grown in monolayer ( Figure 7 D) . Sox9 , Col2A1 and Acan were overexpressed (3.3-fold, 4.9-fold and 7.2-fold, respectively) in chondrocytes from MCh.…”

Section: Resultsmentioning

confidence: 99%

“…Chondrocytes from bovine nasal septa were isolated using recombinant collagenase as described elsewhere [51,58,59]. One gram of minced tissue was incubated with a mixture of 300 units of Collagenase H (Abiel, Palermo, Italy) plus Thermolysin (Promega, Milano, Italy) 250 µg in 10 mL of Dulbecco's Modified Eagle's medium (Sigma, Milan, Italy) containing 1% Pen-Strep and 1% Fungizone (Euroclone, Milan, Italy).…”

Section: Isolation Of Nasal Septal Chondrocytes From Bovine Tissuementioning

confidence: 99%

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Production of Injectable Marine Collagen-Based Hydrogel for the Maintenance of Differentiated Chondrocytes in Tissue Engineering Applications

Rigogliuso

Salamone

Barbarino

et al. 2020

IJMS

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Cartilage is an avascular tissue with limited ability of self-repair. The use of autologous chondrocyte transplants represent an effective strategy for cell regeneration; however, preserving the differentiated state, which ensures the ability to regenerate damaged cartilage, represents the main challenge during in vitro culturing. For this purpose, we produced an injectable marine collagen-based hydrogel, by mixing native collagen from the jellyfish Rhizostoma pulmo with hydroxy-phenyl-propionic acid (HPA)-functionalized marine gelatin. This biocompatible hydrogel formulation, due to the ability of enzymatically reticulate using horseradish peroxidase (HPR) and H2O2, gives the possibility of trap cells inside, in the absence of cytotoxic effects, during the cross-linking process. Moreover, it enables the modulation of the hydrogel stiffness merely varying the concentration of H2O2 without changes in the concentration of polymer precursors. The maintenance of differentiated chondrocytes in culture was then evaluated via morphological analysis of cell phenotype, GAG production and cytoskeleton organization. Additionally, gene expression profiling of differentiation/dedifferentiation markers provided evidence for the promotion of the chondrogenic gene expression program. This, combined with the biochemical properties of marine collagen, represents a promising strategy for maintaining in vitro the cellular phenotype in the aim of the use of autologous chondrocytes in regenerative medicine practices.

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Section: Resultsmentioning

confidence: 99%

Section: Isolation Of Nasal Septal Chondrocytes From Bovine Tissuementioning

confidence: 99%

Production of Injectable Marine Collagen-Based Hydrogel for the Maintenance of Differentiated Chondrocytes in Tissue Engineering Applications

Rigogliuso

Salamone

Barbarino

et al. 2020

IJMS

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“…To improve either yield or quality of extracted ad-MVFs, an isolation procedure was optimized. We herein show for the first time that highly pure recombinant collagenase H can digest native adipose tissue as obtained for viable chondrocyte isolation from cartilaginous tissues [42]. Moreover, it should be noted that the ad-MVFs isolated with this procedure showed higher median length and maintained intact microvascular morphology, including a lumen, an endothelium, and a perivascular structure (33 to 955 µm vs. 20 to 150 µm) [41].…”

Section: Discussionmentioning

confidence: 74%

3D Collagen Hydrogel Promotes In Vitro Langerhans Islets Vascularization through ad-MVFs Angiogenic Activity

et al. 2021

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Adipose derived microvascular fragments (ad-MVFs) consist of effective vascularization units able to reassemble into efficient microvascular networks. Because of their content in stem cells and related angiogenic activity, ad-MVFs represent an interesting tool for applications in regenerative medicine. Here we show that gentle dissociation of rat adipose tissue provides a mixture of ad-MVFs with a length distribution ranging from 33–955 μm that are able to maintain their original morphology. The isolated units of ad-MVFs that resulted were able to activate transcriptional switching toward angiogenesis, forming tubes, branches, and entire capillary networks when cultured in 3D collagen type-I hydrogel. The proper involvement of metalloproteases (MMP2/MMP9) and serine proteases in basal lamina and extracellular matrix ECM degradation during the angiogenesis were concurrently assessed by the evaluation of alpha-smooth muscle actin (αSMA) expression. These results suggest that collagen type-I hydrogel provides an adequate 3D environment supporting the activation of the vascularization process. As a proof of concept, we exploited 3D collagen hydrogel for the setting of ad-MVF–islet of Langerhans coculture to improve the islets vascularization. Our results suggest potential employment of the proposed in vitro system for regenerative medicine applications, such as the improving of the islet of Langerhans engraftment before transplantation.

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“…Primer sequences are listed in Table S1 , and amplicons length ranged from 100 to 180 bp. The gyrB and recA were chosen as reference genes, and a normalization factor was calculated as reported in [ 35 , 36 ]. Data analysis was carried out using the ΔΔCT method [ 37 ].…”

Section: Methodsmentioning

confidence: 99%

Targeting Virulence Genes Expression in Vibrio vulnificus by Alternative Carbon Sources

Nicosia

Salamone

Tagliavia

2022

IJMS

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Vibrio vulnificus is an opportunistic human pathogen causing self-limiting gastroenteritis, life-threatening necrotizing soft tissue infection, and fulminating septicaemia. An increasing rate of infections has been reported worldwide, characterized by sudden onset of sepsis and/or rapid progression to irreversible tissue damage or death. Timely intervention is essential to control the infection, and it is based on antibiotic therapy, which does not always result in the effective and rapid blocking of virulence. Inhibitors of essential virulence regulators have been reported in the last years, but none of them has been further developed, so far. We aimed to investigate whether exposure to some carbon compounds, mostly easily metabolizable, could result in transcriptional down-regulation of virulence genes. We screened various carbon sources already available for human use (thus potentially easy to be repurposed), finding some of them (including mannitol and glycerol) highly effective in down-regulating, in vitro and ex-vivo, the mRNA levels of several relevant -even essential- virulence factors (hlyU, lrp, rtxA, vvpE, vvhA, plpA, among others). This paves the way for further investigations aiming at their development as virulence inhibitors and to unveil mechanisms explaining such observed effects. Moreover, data suggesting the existence of additional regulatory networks of some virulence genes are reported.

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Neural Crest-Derived Chondrocytes Isolation for Tissue Engineering in Regenerative Medicine

Cited by 9 publications

References 32 publications

Production of Injectable Marine Collagen-Based Hydrogel for the Maintenance of Differentiated Chondrocytes in Tissue Engineering Applications

Production of Injectable Marine Collagen-Based Hydrogel for the Maintenance of Differentiated Chondrocytes in Tissue Engineering Applications

3D Collagen Hydrogel Promotes In Vitro Langerhans Islets Vascularization through ad-MVFs Angiogenic Activity

Targeting Virulence Genes Expression in Vibrio vulnificus by Alternative Carbon Sources

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