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Direct observation of CD4 T cell morphologies and their cross-sectional traction force derivation on quartz nanopillar substrates using focused ion beam technique.

Kim DJ, Kim GS, Hyung JH, Lee WY, Hong CH, Lee SK - Nanoscale Res Lett (2013)

Bottom Line: Direct observations of the primary mouse CD4 T cell morphologies, e.g., cell adhesion and cell spreading by culturing CD4 T cells in a short period of incubation (e.g., 20 min) on streptavidin-functionalized quartz nanopillar arrays (QNPA) using a high-content scanning electron microscopy method were reported.Furthermore, we first demonstrated cross-sectional cell traction force distribution of surface-bound CD4 T cells on QNPA substrates by culturing the cells on top of the QNPA and further analysis in deflection of underlying QNPA via focused ion beam-assisted technique.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physics, Chung-Ang University, Seoul 156-756, Republic of Korea. sangkwonlee@cau.ac.kr.

ABSTRACT
Direct observations of the primary mouse CD4 T cell morphologies, e.g., cell adhesion and cell spreading by culturing CD4 T cells in a short period of incubation (e.g., 20 min) on streptavidin-functionalized quartz nanopillar arrays (QNPA) using a high-content scanning electron microscopy method were reported. Furthermore, we first demonstrated cross-sectional cell traction force distribution of surface-bound CD4 T cells on QNPA substrates by culturing the cells on top of the QNPA and further analysis in deflection of underlying QNPA via focused ion beam-assisted technique.

No MeSH data available.


Related in: MedlinePlus

SEM images of captured CD4 T cells on four different sizes of QNPA substrates. (a) Top and (b) tilt views. All captured cells were highlighted in blue for easy distinction.
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Figure 2: SEM images of captured CD4 T cells on four different sizes of QNPA substrates. (a) Top and (b) tilt views. All captured cells were highlighted in blue for easy distinction.

Mentions: Figure 2a,b shows SEM images (top, tilt, and enlarged views) of CD4 T cells bound on four different sizes of STR-functionalized QNPA substrates. The diameters of QNPA using four PS NPs (200, 300, 430, and 750 nm in diameter) were approximately 100, 200, 300, and 450 nm, respectively, as determined by SEM. The detailed morphologies of the captured T cells on STR-QNPA substrates were examined by quantitative SEM analysis using a cell freezing technique described previously. These results exhibit that the captured T cells were well bound on the surface with different morphologies of filopodia or lamellipodia as shown in Figure 2a,b. Interestingly, these images indicate that the morphology (e.g., width of these surface components) of the captured T cells is highly correlated with the size of QNPA in diameter from 200 to 450 nm. To ensure the evaluation of the filopodial width in the early stage of cell adhesion, we quantified at least approximately 20 cells. As a result, the widths of filopodia protruding from T cells bound on QNPA were determined to be approximately 69.00 ± 15.10, 71.60 ± 17.1, 104.40 ± 32.50, and 212.50 ± 16.00 nm corresponding to QNPA surface diameters of approximately 100, 200, 300, and 450 nm, respectively, as shown in Figures 2 and 3a. Filopodial morphologies on STR-QNPA below approximately 300 nm in diameter present a long extended shape, but it extends to be remarkably narrow as it has to be confined by adjacent STR-QNPs with 450 nm diameter. We noticed that captured CD4 T cells on the STR-QNPA surfaces exhibited striking differences in morphology on the varied diameters, even under the condition of extremely early stages of adhesion and statically stable activity of T cells (approximately 20-min incubation at 4°C). Furthermore, to assess the significance of our correlation results, p values were calculated with neighboring column data. Figure 3a exhibits that the distribution of extended filopodial width of the captured CD4 T cells were observed to increase in width by increasing the diameter of QNPA from 200 to 450 nm (**** p < 0.0001, Figure 3b,c), resulting in a good linear response between the width of T cells and diameter of QNPA (R2 = 0.994, n = 20). On the other hand, the filopodial width for 100-nm QNPA shows a similar trend in size to that of the 200-nm QNPA, exhibiting a statistically insignificant difference (* p = 0.0448, bottom part in Figure 3a,b).


Direct observation of CD4 T cell morphologies and their cross-sectional traction force derivation on quartz nanopillar substrates using focused ion beam technique.

Kim DJ, Kim GS, Hyung JH, Lee WY, Hong CH, Lee SK - Nanoscale Res Lett (2013)

SEM images of captured CD4 T cells on four different sizes of QNPA substrates. (a) Top and (b) tilt views. All captured cells were highlighted in blue for easy distinction.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750221&req=5

Figure 2: SEM images of captured CD4 T cells on four different sizes of QNPA substrates. (a) Top and (b) tilt views. All captured cells were highlighted in blue for easy distinction.
Mentions: Figure 2a,b shows SEM images (top, tilt, and enlarged views) of CD4 T cells bound on four different sizes of STR-functionalized QNPA substrates. The diameters of QNPA using four PS NPs (200, 300, 430, and 750 nm in diameter) were approximately 100, 200, 300, and 450 nm, respectively, as determined by SEM. The detailed morphologies of the captured T cells on STR-QNPA substrates were examined by quantitative SEM analysis using a cell freezing technique described previously. These results exhibit that the captured T cells were well bound on the surface with different morphologies of filopodia or lamellipodia as shown in Figure 2a,b. Interestingly, these images indicate that the morphology (e.g., width of these surface components) of the captured T cells is highly correlated with the size of QNPA in diameter from 200 to 450 nm. To ensure the evaluation of the filopodial width in the early stage of cell adhesion, we quantified at least approximately 20 cells. As a result, the widths of filopodia protruding from T cells bound on QNPA were determined to be approximately 69.00 ± 15.10, 71.60 ± 17.1, 104.40 ± 32.50, and 212.50 ± 16.00 nm corresponding to QNPA surface diameters of approximately 100, 200, 300, and 450 nm, respectively, as shown in Figures 2 and 3a. Filopodial morphologies on STR-QNPA below approximately 300 nm in diameter present a long extended shape, but it extends to be remarkably narrow as it has to be confined by adjacent STR-QNPs with 450 nm diameter. We noticed that captured CD4 T cells on the STR-QNPA surfaces exhibited striking differences in morphology on the varied diameters, even under the condition of extremely early stages of adhesion and statically stable activity of T cells (approximately 20-min incubation at 4°C). Furthermore, to assess the significance of our correlation results, p values were calculated with neighboring column data. Figure 3a exhibits that the distribution of extended filopodial width of the captured CD4 T cells were observed to increase in width by increasing the diameter of QNPA from 200 to 450 nm (**** p < 0.0001, Figure 3b,c), resulting in a good linear response between the width of T cells and diameter of QNPA (R2 = 0.994, n = 20). On the other hand, the filopodial width for 100-nm QNPA shows a similar trend in size to that of the 200-nm QNPA, exhibiting a statistically insignificant difference (* p = 0.0448, bottom part in Figure 3a,b).

Bottom Line: Direct observations of the primary mouse CD4 T cell morphologies, e.g., cell adhesion and cell spreading by culturing CD4 T cells in a short period of incubation (e.g., 20 min) on streptavidin-functionalized quartz nanopillar arrays (QNPA) using a high-content scanning electron microscopy method were reported.Furthermore, we first demonstrated cross-sectional cell traction force distribution of surface-bound CD4 T cells on QNPA substrates by culturing the cells on top of the QNPA and further analysis in deflection of underlying QNPA via focused ion beam-assisted technique.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physics, Chung-Ang University, Seoul 156-756, Republic of Korea. sangkwonlee@cau.ac.kr.

ABSTRACT
Direct observations of the primary mouse CD4 T cell morphologies, e.g., cell adhesion and cell spreading by culturing CD4 T cells in a short period of incubation (e.g., 20 min) on streptavidin-functionalized quartz nanopillar arrays (QNPA) using a high-content scanning electron microscopy method were reported. Furthermore, we first demonstrated cross-sectional cell traction force distribution of surface-bound CD4 T cells on QNPA substrates by culturing the cells on top of the QNPA and further analysis in deflection of underlying QNPA via focused ion beam-assisted technique.

No MeSH data available.


Related in: MedlinePlus