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Physical stimuli have a great impact on the survival, behavior, and function of all life. Recently, a  theory has been proposed to understand cellular behavior in times of failure and disease, stating that  cellular processes and damage can be affected by mechanical forces. Scientific evidence suggests that  mechanical changes can affect many of the primary cellular mechanisms, as well as important aspects  of cell behavior such as cell adhesion, movement, and signal transmission. In this study, we examine  the history of the effects of mechanical stimulation from the beginning to the present, what is cell  biomechanics, introduce features and methods to study cell characteristics such as micropipette  suction and atomic force microscopy (AFM). In addition, to consider the effects of biomechanical  stimulation it should be point out to cells using stimuli which can be useful including inducing  differentiation into stem cells to produce bone cells that are applied by special devices that are briefly  examined for the beneficial effects. It also has negative aspects for the cell, such as the occurrence of  metastases in cancer cells. In this work, the effect of changes in the inclination, frequency and strain  on the cancer cell was also investigated. 

Keywords: Cell; Biomechanics; Cell biomechanics; Mechanical stimulation 

The effect of mechanical stimulation on  cell function dates back to the 17th  century; Galileo was the first to investigate  the effect of mechanical loads on bone  morphology [1-3]. Meyer et al. studied the  orientation of the trabeculae in the  cancellous bone. Culmann, in  collaboration with Venir, proposed the  theory of trajectory. Based on this theory,  the orientation of the trabeculae is in line with the main stresses applied to the bone [4-6]. Wolff introduced a law called the  Wolff Law, according to which the  orientation of the trabeculae would also  change if we changed the load on the bone;  However, the first laboratory studies to  investigate the effect of mechanical  stimulation on cell function date back to  1939. Several researchers used different  loads on these cells using cells obtained  from chick tibia and evaluated the results. After these experiments, a lot of research  was done in this field, which led to the  development of methods and a variety of  devices to measure the effects of  mechanical stimulation on performance [7- 11]. Recently, a perspective on  understanding cellular behavior in cases of  failure and disease has been proposed,  which states that cellular processes and  damage can be affected by mechanical  forces. Scientific evidence suggests that  mechanical changes can affect many of the  primary cellular mechanisms, as well as  important aspects of cell behavior such as  cell adhesion, movement, and signal  transmission [12-15]. Taken together, these  forces play an important role in the  proliferation of biological tissue, its  organization, and its response to stimuli.  At the cellular level, mechanical forces are  important in guiding how cells function,  because it is actively felt by living cells.  Cellular responses to mechanical stimuli  often result in adaptive changes that alter  shape and function [16-19]. It should be  noted that any change in cell behavior due  to biochemical processes that occur  between the human body and invasive  external factors or disease progression  significantly alters the mechanical  properties of the cell. Cancer may results  from the malfunction of biological cells and many patient cells proliferate uncontrollably and disrupt tissue organization. Cancer alters the  cytoskeleton and, by affecting the cell’s  mechanics, alters its ability to deform,  causing the ability of healthy cells to move  differently, causing these cells to move in  the tissue and cause metastasis. Therefore,  measuring mechanical properties can be  considered as an indicator of its biological  status and also help to diagnose the cause  of the disease and distinguish between two  diseases [12-16]. 

Mechanics is defined as the study of the  relationships between forces acting on cells  and their deformations. Functional  characteristics of different cell types and  the basic structure of living organisms as  one of the most important factors for the  very development of biology and  biomechanics. The mechanical properties  of cells include nonlinearity, anisotropy  and stiffness. It also has various functional  aspects including communication with  separate components of the cytoskeleton  and cell organs, cell response to external  stimulation and effect on the extracellular  matrix. To examine cellular characteristics,  different methods such as magnetic  tweezers, magnetic torsion cytometry,  optical stretcher, laser deformation  tracking, and cell inductors are used. One  of the best and most efficient methods is  atomic force microscopy [15-18].  The Atomic Force Microscope (AFM) examines the sample surface with a sharp  needle 2 microns long and often with a tip  diameter of less than 10 nm. The needle is  located at the free end of the cantilever,  about 100 to 450 microns long as shown in  Figure 1. The forces between the sample  surface and the needle cause the cantilever  to bend or deflect, and a detector measures  the amount of cantilever deflection while  the needle scrubs the sample surface;  Measures in systems where the sample  performs scanning motion as shown in Fig.  2.

The laser beam is reflected back to the  cantilever towards a position-sensitive  optical detector (PSPD). The ratio of the distance between the cantilever and the  detector to the length of the cantilever acts  as a mechanical amplifier. As a result, the  system can measure the vertical motion of less than one angstrom of the cantilever  tip. 

3-1- Micropipette suction  

In this method, a pipette (a thin glass tube  is used to transfer small and precise  amounts of liquid) and is fixed on the  surface of a cell and enters the pipette by  creating a negative pressure and sucks part  of the cell into the pipette. In other words,  the pipette head is very small compared to  the size of the cell; by applying negative  pressure to the cell membrane, a small part  of the cell enters the pipette head. This  method is a fixed technique for measuring  the stiffness of mammalian cell  membranes, in which the scale of inverse  measurement of cell stiffness is considered  to be the height of the sucked part. The  pressure application technique can also be  used to determine the cell response time. In  this technique, the rest time is a scale for  measuring the viscoelastic properties of the  cell. 

The evaluation of biomechanics cells shows that round, spherical and more  rigid than cancer cells. They are usually  reluctant to detach from the source of  their formation and presence, and as  long as they are not affected by an invasive factor or a chain of adverse  reactions at the location of these cells,  they are likely to change spontaneously  in shape and structure as shown in Fig. 3.

3-2- Atomic Force Microscope (AFM)

AFM is a method that is widely used to  determine cell components and  characteristics. The AFM microscope used as an imaging tool and led to a new  approach in biological research, although  this microscope is different from the  electron light microscope. In this  technique, the specimens are inserted with  a needle at the end of a pedestal The  corresponding tine is followed by a laser,  probed. In addition, the measurement of  the force applied between the tip of the  probe and the specimen, i.e. the force displacement curve, is important to  determine the physical properties of the  specimen. As a result, because the needle  moves on the sample surface, the AFM force curve can be used to evaluate the  hardness and adhesion properties of the  cell membrane. It is worth mentioning that  its most important advantage is the  possibility of using it in aqueous and liquid  environments. Other prominent  applications include non-invasive imaging  and measurement of cellular mechanical  properties in tissues. Wu et al. used it to  measure heart cells. Researchers also used  the technique to identify the mechanisms  of force transmission in tendons to  investigate the mechanical behavior of  surfaces in a fiber tensile test. Stewart et al.  were able to obtain mechanical stress  strength and volume of pressurized animal  cells with the support of a cantilever at  constant altitude in AFM and a light  microscope. They also managed to provide  a way to ensure the stability of  measurement parameters such as volume  and tracking of intracellular activity and  interpretation of physical parameters [19- 28]. 

To provide the general characteristics of  the cell, the mechanical properties of the  nucleus, which is the main component of  the cell, are important. The forces are  transmitted from the surface of the cell and  change the overall characteristics of the  cell [29-33]. Different mechanisms have  been proposed to convert mechanical  stimulation into physiologically  understandable stimulation of the cell. In  general, the process of converting  mechanical excitation can be divided into  three parts. First, the stimulation is  received by the cell surface mechanical  receptors and transmitted to the cell. In the  next step, this stimulus is transmitted into  the cell and to the target receptor. Cells are  likely to use their chemical or skeletal  pathways to transmit stimuli. Finally, by  activating the target section, the desired  activity is performed. Mechanical receptors  in the space between inside and outside the  cell include integrins, stretch-sensitive ion  channels, and cell surface proteins. Stimulation is also transmitted in two  ways, one via the skeleton and the other  biochemically. It is noteworthy that the  transmission of a mechanical stimulus in  the form of a mechanical wave propagation  is much faster than the transfer of a  stimulus in the form of a chemical wave.  According to research, the transfer of  excitation through mechanical diffusion  takes about 1 to 5 milliseconds, while its  chemical transfer through penetration or  displacement takes about 5 to 10 seconds.  As a result, the only path of mechanical  excitation to the nucleus is not the  chemical path, and the transmission  through mechanical wave propagation and  its conversion to chemical excitation at the  core surface can also be mentioned.  However, with many researches that have  been done in the field of studying the paths of receiving, transmitting and converting  mechanical stimuli, the exact mechanism  of this process is still unknown [33-35]. Mechanical stimuli applied to the body can  cause changes in cell function that can be  positive (e. g, improvement in bone  mechanical properties) or negative (such as  damage to a vessel wall or ECM cancer  due to a change in the nature of the cancer) [6-12]. One of the most common methods  for mechanical stimulation of cells is  called bed tension. Also, micropapillary bed can be used to measure the forces  produced by cells. As mentioned earlier,  cancer is caused by malfunctioning  biological cells. In cancer, cells begin to  multiply uncontrollably, disrupting tissue structure. The cytoskeleton is the cell’s  internal scaffold, which consists of a  complex network of biopolymer molecules  that determine the shape and characteristics  of the cell’s mechanical deformation and,  along with proteins, play an important role  in cellular processes such as migration, cell  division, and mechanical conduction. The  ECM, is the latent protein in the  intercellular space that binds cells together  to form tissue. Cancer changes the  structure of the cytoskeleton and ECM [24- 28]. This structure of the protein-woven  deformation also affects the cancer cell’s  ability to contract and expand by affecting  the deformation mechanism. Therefore, the  rate at which cancer cells move can be  very different from that of normal cells,  causing the metastasis to occur; That cells  migrate throughout the tissue to various  organs of the human body and cause tumor  metamorphosis [36-38]. In contrast, the  presence of cytoskeletal defects is effective  in many diseases, such as cancerous  tumors, guiding the cytoskeleton to modify  its structure and mechanical and  biomechanical functions can provide a way to treat cancer. According to the latest  statistics from the World Cancer Registry,  the International Agency for Research on  Cancer predicts that in the next two  decades the number of new people with  cancer may increase by 47%. Contrary to  advances in treatment as well as the  findings of cancer studies, different types  of cancers have not yet responded to  treatment and can be treated for a variety  of reasons, such as environmental factors  or lifestyle. Biochemical processes  between the human body and invasive  external factors or disease progression  cause various changes that cause changes  in cell function, which in turn changes the  mechanical properties of the cell, so the  biological status index can be measured. It  should be considered mechanical  properties and also helped to diagnose the  cause of the disease and to differentiate  between the two diseases [36-38]. 

 In this section, we discuss the results of  one of the studies performed on the ability  of cancer cells to malform under pressure  between two glass plates, or in other  words, microplates in different conditions [6-10]. In this experiment, the cell was  modeled with two separate parts (showing  the cytoplasm and showing the cell  membrane) and Maxwell’s viscoelastic  model was considered; it is possible to  consider different mechanical models for  epithelial cells, but according to research,  Maxwell viscoelastic models provide  closer proximity to laboratory results, and  also because in cell mechanics,  cytoskeletal components have the greatest  impact. They also have viscoelastic  properties. It can be said that the most  suitable material model for the cell is the  viscoelastic model. It is worth mentioning that in the study, the inner part or  cytoplasm of Maxwell viscoelastic model  was homogeneous, isotropic and  incompressible and its membrane was  considered homogeneous and isotropic. 

5-1-Membrane thickness changes 

According to research in a process of  cancer by Sphingosylphorylcholine (SPC)  protein, changes in membrane thickness  due to the presence of actin in it and the  effect of SPC on actin organization are  well known. Increasing the thickness of the  membrane leads to a decrease in the  reaction force of the cell. In other words,  there is an inverse relationship between  force and thickness. 

5-2-Effect of frequency thickness changes

Frequency is another factor that can affect  the mechanical properties of the cell,  which increases with increasing frequency,  force and reaction of the cell. 

5-3- Effect of strain changes on malignant 

Because only cancer cells can metastasize,  it can only be evaluated in this group of  cells. In this case, the strain force  increases, the reaction force may increase. 

5-4- Effect of elastic changes  

The effects of elastic changes are also directly related to the reaction force, and  with increasing membrane elasticity, the  reaction force increases [8-12]. 

5-5- Effect of biomechanical stimuli on bone  cells  

As mentioned, mechanical stimulation can  also have positive effects, for example,  their effect on stem cells and their transformation into bone cells, which may explain in more detail below Stem cells are  cells that are not yet specialized and can  differentiate into other cell lines. These cells are divided into different categories,  including the ability to differentiate based  on the degree of differentiation, which is  one of the most important of these  categories. This category can also be  divided into the following categories: all  power, high power and multi power [7-12]. Bioreactor refers to any system or artificial  device built or engineered that is used in  the subject as a differentiator. There are 5  types of commercial bioreactors for  different tissues, including: Flask Spinner  SFs, Perfusion bioreactors, Wall-mounted  bioreactors (RWV: Vessel Wall Rotating),  Z-reactorP, and reactors. Biot rotating  biaxial (BXR) [1]. In addition to fluid flow  to create shear stresses, these bioreactors  improve the growth and proliferation of  cells within the scaffold by using  appropriate mass transfer. Many  researchers have studied the efficiency and  effectiveness of bioreactors to induce  differentiation, However Sim et al. worked on the effect of compressive stress on the  fate of human pneumatic stem cells. They  extracted human mesenchymal stem cells (MSCs) from bone marrow cells (BMC) and cultured them on a bed that had the  flexibility and for 5 weeks at a pressure of  5 KPa and a frequency of 1 Hz, twice daily  and each loads last for 10 minutes. In order  to be able to compare the results, they  placed the control cells in the previous  chip, except that this group of cells was not mechanically stimulated. In these  experiments, in order to be able to  investigate only the effect of mechanical  stimulation on the cell, bone growth  medium without growth factors was used [42-50]. In addition to their initial goal  mentioned earlier, Sim et al. Also  investigated the effect of different loading  amplitudes on cell fate, and the results of  their experiments showed that the rate of cell proliferation in a group was 1.5 times  higher than that of control cells. However,  the rate of reproduction decreases  compared to when bioreactors are used. One of the reasons that can be mentioned  is the reduction of food and gas transfer.  Experiments also show that this  mechanism and the use of microchips  induce the early stages of bone tissue  production [8]. The cells were subjected to  mechanical stimulation for 15 minutes, 60  minutes and 2 hours. Each was repeated 3  times and the loading results were  examined each time. The general  conclusion that can be drawn from this  study is that by increasing the time of each  loading period, it can cause damage to the cells but increase the expression of  markers. In addition, the results showed  that repetition of processes reduces cell  damage, which is due to the adaptation of  cells to mechanical stimulation, but  reduces the rate of expression of markers [9]. Using stem cells is not without  problems It is also associated with  problems, including the fact that in the  laboratory, mesenchymal stem cells lose  their differentiation and proliferation,  which must be prevented by various  chemical and physical methods [2-10]. In  general, and with respect to studies and  advances in stem cell differentiation, It can  be said that chemical stimuli are not just  the crucial stimuli of the cell and physical  stimuli such as electrical, mechanical  stimuli, even surface geometry affect the  fate of the cell. It is also noteworthy that  the application of a mechanical parameter  alone does not determine the behavior of  cells; In addition, biaxial traction, uniaxial  traction and compressive stress can help to  differentiate stem cells towards bone cells  [10-18]. The cells of the body, including  MSCs are exposed to various mechanical  forces. The type and magnitude of these  forces vary in different physiological and  pathological conditions and produce a wide  variety of responses in cells that have the  ability to alter cell function. The study of  the response of stem cells to mechanical  forces is of great importance in  understanding the function of cells and  tissues in healthy and diseased conditions.  The ability to differentiate MSCs from  other cells has made them a very important  cellular resource in tissue engineering. In  this study, using AFM we consider the  effect of mechanical loads on the stem  cells which includes examining the  behavior of the cell, affected by the  behavior of its internal components,  through stress distribution and large  deformations. Recently, models of hyper  elastic materials for the brain have been  used to express the nonlinear mechanical  behavior of brain tissue under great  deformation. Rashid et al. [39] consider the  effect of strain rate on mechanical behavior  of brain tissue, high strain rate tensile tests  on brain tissue samples. They presented a  one-term model for the hyper elastic  nonlinear behavior of brain tissue and  presented the parameters of a 6-behavior  model by inverse finite element method,  which is obtained by matching finite  element results with laboratory results.  Feng et al. [40] performed a dynamic shear  test on the white part of the brain to  determine the behavioral model of brain  tissue under various model determine the  parameters required in the hyper elastic  structural model that reflects the non isotropic mechanical behavior of the white  matter. Laksari et al. [41] performed stress limiting stress tests. 

Biochemical processes between the human  body and invasive external factors or  disease progression cause various changes  that cause changes in cell function, which  in turn causes changes in the mechanical  properties of the cell, so the biological  status index can be measured. It considered  mechanical properties and also helped to  diagnose the cause of the disease and to  differentiate between the two diseases. According to the study of mechanical  stimuli in the body can cause changes in  cell and tissue function, which can be  negative or positive. The occurrence of a  positive change depends on the optimal  choice of parameters such as amplitude,  frequency waveform and loading time.  However, if any of these parameters are  not within their proper range, they may not be able to induce the desired effect on the  cell and can even cause cell damage or  death. Therefore, optimization of  mechanical parameters is very important.  However, in addition to applying  parameters to cells for a specific purpose,  cells can be exposed to other  biomechanical stimuli and alter cell  behavior.

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