Isolation and culture of stem cells derived from dental pulp of permanent teeth: a successful method / Isolation and culture of stem cells derived from dental pulp of permanent teeth: a successful method

Objective: To establish cultures of cells from the pulp of permanent teeth by the explant method assessing parameters usually presented by stem cells, such as the expression of certain markers and the differentiation ability into osteogenic, adipogenic, and chondrogenic lineages. This study also aimed to assess the expression of ALDH1 (aldehyde dehydrogenase 1) enzyme activity on the isolated cells. Materials and method: The pulp tissue, obtained from wisdom teeth, was placed in a 6-well plate containing proper culture medium, and stored at 37 °C and 5% CO2 for cell proliferation and plastic adherence. Cells were tested for the expression of surface markers and for ALDH1 enzyme activity, by flow cytometry. In addition, cells were assessedfor multi-differentiation potential. Results: The isolated cells showed high expression of CD44 (98.8%), CD73 (100%), and CD90 (97.2%), and moderate expression of STRO-1 (18.4%) and ALDH1 (16.2%), by flow cytometry. Similarly, the cells showed differentiation ability into all three lineages of cells tested. Conclusion: Our results suggest that the explant method - or cell proliferation method - is suitable for the isolation and cultureof stem cells from dental pulp of permanent teeth. The isolated cells may be considered stem cells, based on the current criteria for their characterization, such as plastic adherence, expression of certain markers, and the absence of others, as well as multi-differentiation potential, which showed to be promising for the application in tissue regeneration.

Comparative analysis of the tissue inflammatory response in human cutaneous and disseminated leishmaniasis

Cutaneous leishmaniasis (CL) is the most frequent clinical form of tegumentary leishmaniasis and is characterised by a single or a few ulcerated skin lesions that may disseminate into multiple ulcers and papules, which characterise disseminated leishmaniasis (DL). In this study, cells were quantified using immunohistochemistry and haematoxylin and eosin staining (CD4+, CD68+, CD20+, plasma cells and neutrophils) and histopathology was used to determine the level of inflammation in biopsies from patients with early CL, late CL and DL (ulcers and papules). The histopathology showed differences in the epidermis between the papules and ulcers from DL. An analysis of the cells present in the tissues showed similarities between the ulcers from localised CL (LCL) and DL. The papules had fewer CD4+ T cells than the DL ulcers. Although both CD4+ cells and macrophages contribute to inflammation in early CL, macrophages are the primary cell type associated with inflammation intensity in late ulcers. The higher frequency of CD20+ cells and plasma cells in lesions demonstrates the importance of B cells in the pathogenesis of leishmaniasis. The number of neutrophils was the same in all of the analysed groups. A comparison between the ulcers from LCL and DL and the early ulcers and papules shows that few differences between these two clinical forms can be distinguished by observing only the tissue.

CD8+ T cells in situ in different clinical forms of human cutaneous leishmaniasis

Introduction Leishmania braziliensis infection induces a large spectrum of lesions that clinically manifest as nodules or papules that progress to ulcers. Although it is already known that T helper cells predominate in the lesions, cytotoxic T cells have also been reported to be present, and their role in leishmaniasis immunopathogenesis is not well known. This study investigated the amounts of CD8+ and granzyme B+ cells in different clinical forms of human cutaneous leishmaniasis (CL). Methods Forty tissue fragments from early (E-CL) and late CL (L-CL) lesions and from disseminated leishmaniasis (DL) - papules and ulcers - were characterized. The inflamed area per fragment was calculated, and the CD8 and granzyme B expression levels in the infiltrates were quantified by counting positive cells in 15 fields. The localization of CD8 and granzyme B was graded subjectively. Results Inflammation was higher in L-CL and DL ulcers. CD8 expression was increased in late ulcerated lesions compared to recent lesions. The increase in CD8+ cells also correlated with the duration of the lesion. Papules had a higher frequency of granzyme B+ cells than E-CL lesions, although the frequency was similar to those for late and DL ulcers. CD8+ cells were mostly found in the papillary dermis. Conclusions CD8+ T and granzyme B+ cells are present in the inflammatory infiltrates of CL and DL and may participate in the immunopathogenesis of Leishmania infection. .