Differentiation of human scalp adipose-derived mesenchymal stem cells into mature neural cells on electrospun nanofibrous scaffolds for nerve tissue engineering applications

Objective: This study aimed to isolate and culture SADS cells, investigate their neurogenic capacity and evaluate their application for nerve tissue engineering

Materials and Methods: In this experimental study, SADS cells were isolated from human adipose tissue. After 7-day treatment of SADS cells with insulin, indomethacin and isobutylmethylxanthine, neurogenic differentiation of SADS cells was investigated. During this study, Poly [?-caprolactone] [PCL] and PCL/gelatin nanofibrous scaffolds were fabricated using electrospinning and subsequently nanofibrous scaffolds were coated with platelet-rich plasma [PRP]. SADS cells were also seeded on nanofibrous scaffolds and neurogentic differentiation of these cells on nanofibers was also evaluated. Effect of PRP on proliferation and differentiation of SADS cells on scaffolds was also studied

Results: Our results showed that after 7-day treatment of SADS cells with insulin, indomethacin and isobutylmethylxanthine, SADS cells expressed markers characteristic of neural cells such as nestin and neuron specific nuclear protein [NEUN] [as early neuronal markers] as well as microtubule-associated protein 2 [MAP2] and neuronal microtubule-associated [TAU] [as mature neuronal markers] while mature astrocyte maker [GFAP] was not expressed. MTT assay and SEM results showed that incorporation of gelatin and PRP into the structure of nanofibrous scaffolds has a significant positive influence on the bioactivity of scaffolds. Our results also showed neurogentic differentiation of SADS cells on scaffolds

Conclusion: Our results demonstrated that SADS cells have potential to differentiate into early and mature progenitor neurons, in vitro. PCL/gelatin/PRP was found to be a promising substrate for proliferation of SADS cells and differentiation of these cells into neural cells which make these scaffolds a candidate for further in vivo experiments and suggest their application for nerve tissue engineering

Cytotoxicity of triple antibiotic paste and calcium hydroxide against cultured human dental pulp fibroblasts

Objective: In necrotic immature teeth, intra canal medicaments such as triple antibiotic paste [TAP] and calcium hydroxide [CH] are used for root canal disinfection and regeneration treatment. However, the effect of these medicaments on dental pulp fibroblasts has yet to be known. This study aimed to assess the cytotoxicity of CH and TAP against cultured human dental pulp fibroblasts [HDPFs] obtained from third molars

Methods: In this in vitro study, fibroblasts were obtained from the dental pulp of two third molars. Fibroblasts were exposed to 0.1, 1 and 10 mg/mL concentrations of TAP and CH. Six samples were prepared of each medicament and fibroblast viability was evaluated after 72 hours. Data were analyzed using one-way and two-way ANOVA [p<0.001]. The percentage of cell viability was calculated and the cytotoxicity of the medicament was categorized as severe [30%], moderate [30- 60%], mild [60-90%] and non-toxic [>90%]

Results: In TAP samples, only the 10 mg/mL concentration had a significant difference with the control group in terms of the percentage of cell viability and showed moderate cytotoxicity. In CH samples, the 1 and 10 mg/mL concentrations showed significant differences with the control group and were severely cytotoxic

Conclusion: Reduction in cell viability of fibroblasts by increase in concentration was significantly greater in CH compared to TAP group. Thus, in regeneration treatments, these medicaments must be used in concentrations with adequate therapeutic and insignificant adverse effects on fibroblasts

Study of human chondrocyte redifferntiation capacity in three-dimensional hydrogel culture

Articular cartilage tissue defects cannot be repaired by the proliferation of resident chondrocytes. Autologous chondrocyte transplantation [ACT] is a relatively new therapeutic approach to cover full thickness articular cartilage defects by in vitro grown chondrocytes from the joint of a patient. Therefore, we investigated the differentiation capability of human chondrocytes maintained in alginate culture. The cartilage specimens obtained from 50 patients who underwent total knee and hip operations at the teaching hospital of Isfahan University of Medical Sciences, Isfahan Iran. Isolated primary chondrocytes were first grown in monolayer cultures for 1 to 6 passages [each passage lasting about 3 days]. At each passage, monolayer cells seeded in alginate culture and investigated morphologically and immuno-cytologically for expression of cartilage-specific markers [collagen type II and cartilage-specific proteoglycans]. The chondrocytes from monolayer passages PI to P4 introduced in alginate cultures regained a chondrocyte phenotype. Cells were interconnected by typical gap junctions and after few days, they produced a cartilage-specific extracellular matrix [collagen type II and cartilage-specific proteoglycans]. In contrast, cells from monolayer passages P5 and P6 did not redifferentiate to chondrocytes in the alginate cultures. Chondrocyte culture was established for the first time in Iran. The alginate culture conditions promote the redifferentiation of dedifferentiated chondrocytes that have still a chondrogenic potential. This procedure opens up a promising approach to produce sufficient numbers of differentiated chondrocytes for ACT. Indeed, in some patients the harvested cells were used immediately and successfully for transplantation

Access to chondrocyte culture, with alginate, in Iran

In this study, chondrocyte culture was established for the first time in Iran, and calcium alginate was used for longer culture of chondrocyte in vitro. The study was programmed in order to be used for future human chondrocyte transplantation. The cartilage specimen obtained from 50 patients who underwent total knee and hip operations in Isfahan University of Medical Sciences. Cartilage specimens were used for monolayer as well as suspension culture in alginate beads. Approximately 12 +/- 1 millions cells were harvested from the 3[RD] passage. The cells were round with large euchromatic nucleus and several nucleoli and small vacuoles. The cells derived from passages 1 to 4, which were grown up then, in alginate beads, showed higher staining with alcian blue. The harvested cells in some patients were immediately and successfully used for autologus transplantation. This later work will be reported separately