Transforming growth factor beta (TGF-ß) isomers influence cell detachment of MG-63 bone cells.

Bone repair and wound healing are modulated by different stimuli. There is evidence that Transforming Growth Factor-beta (TGF-ß) super-family of cytokines have significant effects on bone structure by regulating the replication and differentiation of chondrocytes, osteoblasts and osteoclasts. There is also significant evidence that interactions with extracellular matrix molecules influence cell behaviour. In this study cell surface attachment was examined via a trypsinization assay using various TGF-ß isomers in which the time taken to trypsinize cells from the surface provided a means of assessing the strength of attachment. Three TGF-ß isomers (TGF-ß1, 2 and 3), four combined forms (TGF-ß(1+2), TGF-ß(1+3), TGF-ß(2+3) and TGF-ß(1+2+3)) along with four different controls (BSA, HCl, BSA/HCl and negative control) were investigated in this study. The results indicated that treatment with TGF-ß1, 2, 3 and HCl decreased cell attachment, however, this effect was significantly greater in the case of TGF-ß3 (p<0.001) indicating perhaps that TGF-ß3 does not act alone in cell detachment, but instead functions synergistically with signalling pathways that are dependent on the availability of hydrogen ions. Widefield Surface Plasmon Resonance (WSPR) microscope was also used to investigate cell surface interactions.

Tracking traction force changes of single cells on the liquid crystal surface.

Cell migration is a key contributor to wound repair. This study presents findings indicating that the liquid crystal based cell traction force transducer (LCTFT) system can be used in conjunction with a bespoke cell traction force mapping (CTFM) software to monitor cell/surface traction forces from quiescent state in real time. In this study, time-lapse photo microscopy allowed cell induced deformations in liquid crystal coated substrates to be monitored and analyzed. The results indicated that the system could be used to monitor the generation of cell/surface forces in an initially quiescent cell, as it migrated over the culture substrate, via multiple points of contact between the cell and the surface. Future application of this system is the real-time assaying of the pharmacological effects of cytokines on the mechanics of cell migration.

Sensitivity and specificity of the empirical lymphocyte genome sensitivity (LGS) assay: implications for improving cancer diagnostics.

Lymphocyte responses from 208 individuals: 20 with melanoma, 34 with colon cancer, and 4 with lung cancer (58), 18 with suspected melanoma, 28 with polyposis, and 10 with COPD (56), and 94 healthy volunteers were examined. The natural logarithm of the Olive tail moment (OTM) was plotted for exposure to UVA through 5 different agar depths (100 cell measurements/depth) and analyzed using a repeated measures regression model. Responses of patients with cancer plateaued after treatment with different UVA intensities, but returned toward control values for healthy volunteers. For precancerous conditions and suspected cancers, intermediate responses occurred. ROC analysis of mean log OTMs, for cancers plus precancerous/suspect conditions vs. controls, cancer vs. precancerous/suspect conditions plus controls, and cancer vs. controls, gave areas under the curve of 0.87, 0.89, and 0.93, respectively (P<0.001). Optimization allowed test sensitivity or specificity to approach 100% with acceptable complementary measures. This modified comet assay could represent a stand-alone test or an adjunct to other investigative procedures for detecting cancer.

Effects of different transforming growth factor beta (TGF-ß) isomers on wound closure of bone cell monolayers.

This study aimed at determining the role of the transforming growth factor-beta (TGF-ß) isomers and their combinations in bone cell behaviour using MG63 cells. The work examined how TGF-ß1, 2 and 3 and their solvent and carrier (HCl and BSA, respectively) effected cell morphology, cell proliferation and integrin expression. This study also aimed at examining how the TGF-ßs and their solvent and carrier influenced wound closure in an in vitro wound closure model and how TGF-ßs influence extracellular matrix (ECM) secretion and integrin expression. The wound healing response in terms of healing rate to the TGF-ßs and their solvent/carrier was investigated in 300 µm ± 10-30 µm SD wide model wounds induced in fully confluent monolayers of MG63 bone cells. The effect of different TGF-ß isomers and their combinations on proliferation rate and cell length of human bone cells were also assessed. Immunostaining was used to determine if TGF-ßs modifies integrin expression and ECM secretion by the bone cells. Imaging with WSPR allowed observation of the focal contacts without the need for immunostaining. The wound healing results indicated that TGF-ß3 has a significant effect on the wound healing process and its healing rate was found to be higher than the control (p < 0.001), TGF-ß1 (p < 0.001), TGF-ß2 (p < 0.001), BSA/HCl (p < 0.001) and HCl (p < 0.001) in ascending order. It was also found that TGF-ß1 and TGF-ß2 treatment significantly improved wound closure rate in comparison to the controls (p < 0.001). All TGF-ß combinations induced a faster healing rate than the control (p < 0.001). It was expected that the healing rate following treatment with TGF-ß combinations would be greater than those healing rates following treatments with TGF-ß isomers alone, but this was not the case. The results also suggest that cell morphological changes were observed significantly more in cells treated with TGF-ß(2 + 3) and TGF-ß(1 + 3) (p < 0.001). Any cell treated with TGF-ß1, TGF-ß(1 + 2) and TGF-ß(1 + 2 + 3) showed significantly less elongation compared to the control and other TGF-ß isomers. In terms of proliferation rate, TGF-ß3 and TGF-ß(2 + 3) increased cell numbers more than TGF-ß1, TGF-ß2 and other combinations. TGF-ß1 and its combinations did not show significant proliferation and attachment compared to the control. Immunostaining indicated that treatment with TGF-ß3 significantly enhanced the secretion of collagen type I, fibronectin and integrins α3 and ß1. The WSPR experiments also indicated that TGF-ßs influenced the distribution of focal contacts. In conclusion, combining TGF-ß3 with any other TGF-ß isomer resulted in a faster model wound closure rate (p < 0.001), while treatment with TGF-ß1 in any TGF-ß combination reduced the healing rate (p < 0.001). It can therefore be concluded that the presence of TGF-ß1 has an inhibitory effect on bone wound healing while TGF-ß3 had the opposite effect and increased the rate of wound closure in a 2 dimensional cell culture environment.