ASSESSMENT OF MORPHOLOGIC AND MECHANICAL PROPERTIES OF T LYMPHOCYTES IN HEALTHY AND INFLAMED GINGIVAL TISSUES
Dental Research Center, Research Institute of Dental Sciences, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
BAHAREH NAZEMI SALMAN *
Department of Pediatric, School of Dentistry, Zanjan University of Medical Sciences, Zanjan, Iran.
ALIREZA TAVAKOLI NEJAD
Auckland Bioengineering Institute (ABI), Auckland, New Zealand.
Dentist, Private Practice, Qazvin University of Medical Sciences, Qazvin, Iran.
Pardis Health Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
*Author to whom correspondence should be addressed.
Objectives: The aim of this study was to investigate whether T lymphocytes of peripheral blood samples would have different mechanical characteristics from those of healthy and inflamed gingival tissue specimens. The results of this basic study can be used to differentiate the healthy and inflamed tissue via Atomic Force Microscopy (AFM). This study assessed different mechanical characteristics of T lymphocytes in healthy and inflamed gingival tissues. Review of literature pertaining to T lymphocytes till date reveals that, this is the one of the first study to assess variations in connective tissue T cells in healthy and diseased periodontal tissues.
Materials and Methods: Periodontal specimens were obtained from patients requiring crown lengthening or surgical treatment for periodontal disease. Peripheral venous blood was drawn from healthy, drug free adults/children donors and also from patients who had periodontitis and mixed immediately with heparin. Gingival specimens were collected in ROSWELL PARK MEMORIAL INSTITUTE 1640 MEDIUM 1640 plus 10% fetal bovine serum. The tissues were weighed, cut into small pieces and incubated in tissue digestion medium which consisted of ROSWELL PARK MEMORIAL INSTITUTE 1640 MEDIUM with 10% fetal bovine serum, 2.4 units/mL disphase in calcium and magnesium free balanced salt solution, 100 units/mL penicillin, 100 µg/ mL streptomycin and 2.5 µg/mL amphotericin B. Mononuclear cells were separated by Ficoll-Hypaque gradient centrifugation and then washed with phosphate buffered saline (PBS) for further processing steps. T lymphocytes were purified from mononuclear cells by using the T cell negative isolation kit from Dynal magnetic cell sorting (MACS). Cells were fixed with 0.5% glutaraldehyde for 1 minute and washed twice for 5 minutes, and then air dried for cell probing. A DME Dual ScopeTM scanning probe microscope was utilized to assess elasticity and surface topography. Cellular mechanical properties were determined by measuring force-distance curves at 5 positions on the cell surface. T-test via the Excel software version 2013 was used to analyze the results of the study with the alpha error level of 0.05.
Results: The difference between the elastic modulus of healthy donor blood T cells with the blood lymphocytes of periodontitis patients was significant (P<0.05). The specimens from infected tissues showed significant differences in the elasticity and morphological properties compared to T cells of healthy tissue. The greater the tissue infection, the more flexible the cells became. There was a significant difference between the tissue T cells of children and adults as well. Blood T cells had different morpho mechanical properties than the tissue T cell groups.
Conclusions: The results of this study showed that as greater the inflammation, greater the motility and less elastic modulus. Differences were also observed in morphology and roughness of T cells at in different groups of study and locations on each cell.
This preliminary study of mechanical properties of T cells using AFM may prove potentially useful in inflammatory disease and cancer research.
Keywords: Atomic force microscopy, t lymphocytes, periodontitis