Living cells sense and induce mechanical forces within their extracellular environment. Recent studies suggest that these mechanical cues activate signaling pathways and reform the structure of the cells. However, understanding of how these mechanical forces are transmitted to the cells and how the intracellular biochemical reactions change the mechanical behavior of the cells remains unclear. In addition, creating a microenvironment where the cell mechanics can be controlled is a challenge. In this dissertation, a set of experimental and computational approaches for investigation of cell biomechanics in a variety of physiological conditions were developed. In addition, novel methods for changing the microenvironment of the cells and controlling the mechanical behavior of the cells were presented. Firstly, the nanoscale poroelasticity of human mammary basal/claudin low carcinoma cell (MDA-MB-231) was investigated using indentation-based atomic force microscopy. The cell poroelastic behavior (i.e., the diffusion coefficient) was quantified at different indenting velocities (0.2, 2, 10, 20, 100, 200 μm/s) and indentation depths (635, 965, and 1313 nm) by fitting the force-relaxation curves using a poroelastic model. Cell treated with cytoskeleton inhibitors are measured to investigate the effect of the cytoskeletal components on the cell poroelasticity. Our results demonstrated that MDA-MB-231 cells behaved less poroelastic (i.e., with lower diffusion coefficient) at higher indenting velocities due to the local stiffening up and dramatic pore size reduction caused by faster force load and was more pronounced when the local cytoplasm porous structure was stretched by higher indentation. Furthermore, inhibition of cytoskeletal components resulted in pronounced poroelastic relaxation when compared with the control, and affected the nonlinearity of cell poroelasticity at different depth range inside of the cell. Then, the effect of substrate mechanics with different stiffness on the nonlinear biomechanical behavior of living cells was investigated using AFM. the actin filament (F-actin) cytoskeleton of the cells was fluorescently stained to investigate the adaptation of F-actin cytoskeleton structure to the substrate mechanics. It was found that living cells sense and adapt to substrate mechanics: the cellular Young's modulus, shear modulus, apparent viscosity, and their nonlinearities (mechanical property vs. measurement depth relation) were adapted to the substrates' nonlinear mechanics. Moreover, the positive correlation between the cellular poroelasticity and the indentation remained the same despite the substrate stiffness nonlinearity, but was indeed more pronounced for the cells seeded on the softer substrates. Comparison of the F-actin cytoskeleton morphology confirmed that the substrate affects the cell mechanics by regulating the intracellular structure. Next, the effect of the substrate morphology on the biomechanical behavior of living cells was thoroughly investigated using indentation-based atomic force microscopy. The results showed that the cellular biomechanical behavior was affected by the substrate morphology significantly. The elasticity and viscosity of the cells on the patterned substrates were much lower compared to those of the ones cultured on flat. The poroelastic diffusion coefficient of the cells was higher on the patterned substrates. In addition, fluorescence images confirmed that cell mechanical behavior and morphology can be controlled using substrates with properly designed topography. Finally, to investigate the cell uptake of NPs, a dynamic cell culture substrate was designed in two steps: 1. the polyaniline polymer (PANI) was deposited on the cell culture petri dish, and 2. the PANI substrate was coated by PDMS with base to curing agent ratio of 10:3 (PDMS/PANI). The substrates were characterized using Fourier Transform Infrared (FTIR) and Atomic Force Microscopy (AFM). It was found that the PDMS/PANI substrate expansion was positively correlated with the applied voltage to the PANI. In addition, the PDMS/PANI substrate was implemented for NPs delivery. Our results showed that the uptake of NPs by the cells cultured on PDMS/PANI substrate increases by expansion of the substrates. Moreover, our results suggest that the PDMS/PANI substrate is a promising device that can be used for controlling intra- and extracellular behavior of the cells.