Single-cell analysis has been considered as a promising way to uncover the underlying mechanisms guiding the mysteries of life activities, which con siderably complements traditio nal en semble assays and yields novel...Single-cell analysis has been considered as a promising way to uncover the underlying mechanisms guiding the mysteries of life activities, which con siderably complements traditio nal en semble assays and yields novel in sights into cell biology. The adve nt of atomic force microscopy (AFM) provides a potent tool for investigati ng the structures and properties of n ative biological samples at the micro/na no scale un der near-physiological conditions, which promotes the studies of single-cell behaviors. In the past decades, AFM has achieved great success in single-cell observation and manipulation for biomedical applications, demonstrating the excellent capabilities of AFM in addressing biological issues at the single-cell level with unprecedented spatiotemporal resolution. In this article, we review the recent advances in single-cell analysis that has been made with the utilization of AFM, and provide perspectives for future progression.展开更多
We develop a two dimensional model of a vesicle adhered on a curved substrate via long-range molecular interactions while subjected to a detachment force. The relationship between the force and displacement of the ves...We develop a two dimensional model of a vesicle adhered on a curved substrate via long-range molecular interactions while subjected to a detachment force. The relationship between the force and displacement of the vesicle is investigated as a function of the substrate shape. It is shown that both the force- displacement relationship and the maximum force at pull-off are significantly dependent on the substrate shape. The results suggest that probes with different tip shapes may be designed for cell manipulation. For example, we demonstrate that a vesicle can be pulled off a fiat surface using a probe with a curved tip.展开更多
Fluidic force microscopy(FluidFM),which combines atomic force microscopy(AFM)with microchanneled cantilevers connected to a pressure controller,Is a technique allowing the realization of force-sensitive nanopipette un...Fluidic force microscopy(FluidFM),which combines atomic force microscopy(AFM)with microchanneled cantilevers connected to a pressure controller,Is a technique allowing the realization of force-sensitive nanopipette under aqueous conditions.FluidFM has unique advantages in simultaneous three-dimensional manipulations and mechanical measurements of biological specimens at the micro-/nanoscale.Over the past decade,FluidFM has shown its potential in biophysical assays particularly in the investigations at single-cell level,offering novel possibilities for discovering the underlying mechanisms guiding life activities.Here,we review the utilization of FluidFM to address biomechanical and biophysical issues in the life sciences.Firstly,the fundamentals of FluidFM are represented.Subsequently,the applications of FluidFM for biophysics at single-cell level are surveyed from several facets,including single-cell manipulations,single-cell force spectroscopy,and single-cell electrophysiology.Finally,the challenges and perspectives for future progressions are provided.展开更多
Immunotherapy based on natural killer(NK)cells is a promising approach for treating a variety of cancers.Unlike T cells,NK cells recognize target cells via a major histocompatibility complex(MHC)-independent mechanism...Immunotherapy based on natural killer(NK)cells is a promising approach for treating a variety of cancers.Unlike T cells,NK cells recognize target cells via a major histocompatibility complex(MHC)-independent mechanism and,without being sensitized,kill the cells directly.Several strategies for obtaining large quantities of NK cells with high purity and high cytotoxicity have been developed.These strategies include the use of cytokine−antibody fusions,feeder cells or membrane particles to stimulate the proliferation of NK cells and enhance their cytotoxicity.Various materials,including peripheral blood mononuclear cells(PBMCs),umbilical cord blood(UCB),induced pluripotent stem cells(iPSCs)and NK cell lines,have been used as sources to generate NK cells for immunotherapy.Moreover,genetic modification technologies to improve the proliferation of NK cells have also been developed to enhance the functions of NK cells.Here,we summarize the recent advances in expansion strategies with or without genetic manipulation of NK cells derived from various cellular sources.We also discuss the closed,automated and GMP-controlled large-scale expansion systems used for NK cells and possible future NK cell-based immunotherapy products.展开更多
Comprehensive Summary Deoxyribonucleic acid(DNA)is a biomacromolecule,as well as a polymeric material,whose sequences with different manipulative structures enable them to implement a series of functions,such as reorg...Comprehensive Summary Deoxyribonucleic acid(DNA)is a biomacromolecule,as well as a polymeric material,whose sequences with different manipulative structures enable them to implement a series of functions,such as reorganization,target,and catalysis.Compared to existing traditional materials incapable of multifunctional integration,the polymeric DNA network is a form of material that can achieve functional integration while maintaining specific DNA properties.Furthermore,precise target enabled by DNA network is one of the most essential components of cellular manipulation.Hence,the DNA network is indispensable and irreplaceable to cell manipulation that it is a versatile tool for the understanding of basic laws of living life and treatments of diseases,such as cell isolation,cell delivery,and cell interference.Herein,the construction of polymeric DNA network is briefly introduced from the aspects of assembly modules,construction methods,and properties.展开更多
Metabolites can directly reflect and modulate cell responses and phenotypical changes by influencing energy balances,intercellular signals,and many other cellular functions throughout the lifespan of cells.Taking into...Metabolites can directly reflect and modulate cell responses and phenotypical changes by influencing energy balances,intercellular signals,and many other cellular functions throughout the lifespan of cells.Taking into account the heterogeneity of cells,single-cell metabolite analysis offers an insight into the functional process within one cell.Microfluidics as a powerful tool has attracted significant interest in the single-cell metabolite analysis field.The microfluidic platform is possible to observe,classify,and stimulate individual cells.It can also transport single-cell to subsequent analysis steps in a fast and controllable way to determine and analyze the composition and content of metabolites.The reviews of topics in microfluidics for single-cell metabolite analysis have been published in the past few years.However,most of them focused on metabolite analysis with mass spectrometry.Here,we covered the advances of microfluidic devices for single-cell metabolite analysis,with a focus on single-cell isolation and manipulation.What is more,we summarized the detection methods and applications of single-cell metabolites.展开更多
BACKGROUND Psoriasis is a chronic autoimmune disease that usually manifests as a red scaly epidermis,induration,and hyperproliferation of basal keratinocytes.About 2%of the world’s population suffers from psoriasis b...BACKGROUND Psoriasis is a chronic autoimmune disease that usually manifests as a red scaly epidermis,induration,and hyperproliferation of basal keratinocytes.About 2%of the world’s population suffers from psoriasis but there are no clear therapeutics yet.Recently,mesenchymal stem cells(MSCs)have been regarded as a therapeutic alternative for autoimmune diseases,as they possess immunosuppressive effects without risks.Human umbilical cord-derived MSCs effectively regulate immune cells and are characterized by low immunogenicity,which has many advantages in treating immune diseases.CASE SUMMARY The patient was a 47-year-old male,diagnosed with psoriasis in 1995.He had received various treatments for 25 years,but the psoriatic condition was not significantly improved.He was given three rounds of minimally manipulated umbilical cord-derived MSCs over 2 wk.The erythema gradually disappeared.Three months after the 1st round,all erythema completely disappeared,and the psoriasis did not recur.CONCLUSION Minimally manipulated umbilical cord-derived MSC transplantation can potentially treat patients who suffer from psoriasis.展开更多
BACKGROUND Alopecia areata(AA)is a common autoimmune disease characterized by hair loss.AA appears in extensive forms,such as progressive and diffusing hair loss(diffuse AA),a total loss of scalp hair(alopecia totalis...BACKGROUND Alopecia areata(AA)is a common autoimmune disease characterized by hair loss.AA appears in extensive forms,such as progressive and diffusing hair loss(diffuse AA),a total loss of scalp hair(alopecia totalis),and complete loss of hair over the entire body(alopecia universalis).Recently,mesenchymal stem cells(MSCs)have been identified as a therapeutic alternative for autoimmune diseases.For this reason,preclinical and case studies of AA and related diseases using MSCs have been conducted.CASE SUMMARY Case 1:A 55-year-old woman suffered from AA in two areas of the scalp.She was given 15 rounds of minimally manipulated umbilical cord-MSCs(MM-UC-MSCs)over 6 mo.The AA gradually improved 3 mo after the first round.The patient was cured,and AA did not recur.Case 2:A 30-year-old woman,with history of local steroid hormone injections,suffered from AA in one area on the scalp.She was given two rounds of MM-UC-MSCs over 1 mo.The AA immediately improved after the first round.The patient was cured,and AA did not recur.Case 3:A 20-year-old woman,who was diagnosed with alopecia universalis at the age of 12,was given 14 rounds of MM-UC-MSCs over 12 mo.Her hair began to grow about 3 mo after the first round.The patient was cured,and alopecia universalis did not recur.CONCLUSION MM-UC-MSC transplantation potentially treats patients who suffer from AA and related diseases.展开更多
We propose a novel on-chip 3D cell rotation method based on a vibration-induced flow.When circular vibration is applied to a microchip with micropillar patterns,a highly localized whirling flow is induced around the m...We propose a novel on-chip 3D cell rotation method based on a vibration-induced flow.When circular vibration is applied to a microchip with micropillar patterns,a highly localized whirling flow is induced around the micropillars.The direction and velocity of this flow can be controlled by changing the direction and amplitude of the applied vibration.Furthermore,this flow can be induced on an open chip structure.In this study,we adopted a microchip with three micropillars arranged in a triangular configuration and an xyz piezoelectric actuator to apply the circular vibration.At the centre of the micropillars,the interference of the vibration-induced flows originating from the individual micropillars induces rotational flow.Consequently,a biological cell placed at this centre rotates under the influence of the flow.Under three-plane circular vibrations in the xy,xz or yz plane,the cell can rotate in both the focal and vertical planes of the microscope.Applying this 3D cell rotation method,we measured the rotational speeds of mouse oocytes in the focal and vertical planes as 63.7±4.0°s^(−1) and 3.5±2.1°s^(−1),respectively.Furthermore,we demonstrated the transportation and rotation of the mouse oocytes and re-positioned their nuclei into a position observable by microscope.展开更多
The development of a novel polymer-based micro robotic gripper that can be actuated in a fluidic medium is presented in this paper.Our current work is to explore new materials and designs for thermal actuators to achi...The development of a novel polymer-based micro robotic gripper that can be actuated in a fluidic medium is presented in this paper.Our current work is to explore new materials and designs for thermal actuators to achieve micromanipulation of live biological cells.We used parylene C to encapsulate a metal heater,resulting in effectively a tri-layered thermal actuator.Parylene C is a bio-compatible dielectric polymer that can serve as a barrier to various gases and chemicals. Therefore,it is suitable to serve as a thermal/electrical/chemical isolation material for protecting the metal heater from exposing to an aqueous environment.We have demonstrated parylene actuators (2mm×100μm×0.5μm)to operate in an aqueous environment using 10 to 80mW input power.The temperature of these actuators at full deflection was estimated to be～60℃,which is much lower than the typical requirement of>100℃ to actuate other conventional MEMS actuators.Danio rerio follicles in fluidic medium were captured successfully using these actuators.Moreover,these actuators were found to be responsive to moderate rise in environmental temperature,and hence,we could vary the fluidic medium temperature to actuate trimorphs on a chip without any input of electrical energy, i.e.,raising the fluidic temperature from 23℃ to 60℃ could actuate the trimorphs to grasp follicles of ～1mm size in diameter.At 60℃,the embryos inside the follicles were observed to be alive,i.e.,they were still moving in the biological fluid isolated by the follicle membrane.The smallest follicles grasped were～500μm in diameter using 800μm×130μm×0.6μm actuators.The fabrication process,modeling, and optimization of the trimorph actuators are presented.Based on the successful operation of these polymer-based actuators,we are currently developing multifinger thermal microgrippers for cellular grasping and manipulation,which can potentially be hybridly integrated with circuits for computer control.展开更多
基金National Natural Science Foundation of China (Nos. 6187325& 61503372, U1613220, and 61433017)Youth Innovation Promotion Association CAS (No. 2017243)CAS FEA International Partnership Program for Creative Research Teams.
文摘Single-cell analysis has been considered as a promising way to uncover the underlying mechanisms guiding the mysteries of life activities, which con siderably complements traditio nal en semble assays and yields novel in sights into cell biology. The adve nt of atomic force microscopy (AFM) provides a potent tool for investigati ng the structures and properties of n ative biological samples at the micro/na no scale un der near-physiological conditions, which promotes the studies of single-cell behaviors. In the past decades, AFM has achieved great success in single-cell observation and manipulation for biomedical applications, demonstrating the excellent capabilities of AFM in addressing biological issues at the single-cell level with unprecedented spatiotemporal resolution. In this article, we review the recent advances in single-cell analysis that has been made with the utilization of AFM, and provide perspectives for future progression.
基金The project supported by the National Natural Science Foundation of China (10525210 and 10121202)the 973 Program
文摘We develop a two dimensional model of a vesicle adhered on a curved substrate via long-range molecular interactions while subjected to a detachment force. The relationship between the force and displacement of the vesicle is investigated as a function of the substrate shape. It is shown that both the force- displacement relationship and the maximum force at pull-off are significantly dependent on the substrate shape. The results suggest that probes with different tip shapes may be designed for cell manipulation. For example, we demonstrate that a vesicle can be pulled off a fiat surface using a probe with a curved tip.
基金supported by the China Scholarship Council(CSC)(No.202004910157)support from the National Natural Science Foundation of China(Nos.61922081 and 61873258)+2 种基金the Key Research Program of Frontier Sciences CAS(No.ZDBS-LY-JSC043)the Youth Innovation Promotion Association CAS(No.2017243)the LiaoNing Revitalization Talents Program(No.XLYC1907072).
文摘Fluidic force microscopy(FluidFM),which combines atomic force microscopy(AFM)with microchanneled cantilevers connected to a pressure controller,Is a technique allowing the realization of force-sensitive nanopipette under aqueous conditions.FluidFM has unique advantages in simultaneous three-dimensional manipulations and mechanical measurements of biological specimens at the micro-/nanoscale.Over the past decade,FluidFM has shown its potential in biophysical assays particularly in the investigations at single-cell level,offering novel possibilities for discovering the underlying mechanisms guiding life activities.Here,we review the utilization of FluidFM to address biomechanical and biophysical issues in the life sciences.Firstly,the fundamentals of FluidFM are represented.Subsequently,the applications of FluidFM for biophysics at single-cell level are surveyed from several facets,including single-cell manipulations,single-cell force spectroscopy,and single-cell electrophysiology.Finally,the challenges and perspectives for future progressions are provided.
基金This work was supported by the Strategic Priority Research Program of the Chinese Academy of Sciences(Grant no.XDB29030202)the Ministry of Science and Technology of China(Grant no.2016YFC1303503).
文摘Immunotherapy based on natural killer(NK)cells is a promising approach for treating a variety of cancers.Unlike T cells,NK cells recognize target cells via a major histocompatibility complex(MHC)-independent mechanism and,without being sensitized,kill the cells directly.Several strategies for obtaining large quantities of NK cells with high purity and high cytotoxicity have been developed.These strategies include the use of cytokine−antibody fusions,feeder cells or membrane particles to stimulate the proliferation of NK cells and enhance their cytotoxicity.Various materials,including peripheral blood mononuclear cells(PBMCs),umbilical cord blood(UCB),induced pluripotent stem cells(iPSCs)and NK cell lines,have been used as sources to generate NK cells for immunotherapy.Moreover,genetic modification technologies to improve the proliferation of NK cells have also been developed to enhance the functions of NK cells.Here,we summarize the recent advances in expansion strategies with or without genetic manipulation of NK cells derived from various cellular sources.We also discuss the closed,automated and GMP-controlled large-scale expansion systems used for NK cells and possible future NK cell-based immunotherapy products.
基金the National Natural Science Foundation of China(grant nos.22225505 and 22174097).
文摘Comprehensive Summary Deoxyribonucleic acid(DNA)is a biomacromolecule,as well as a polymeric material,whose sequences with different manipulative structures enable them to implement a series of functions,such as reorganization,target,and catalysis.Compared to existing traditional materials incapable of multifunctional integration,the polymeric DNA network is a form of material that can achieve functional integration while maintaining specific DNA properties.Furthermore,precise target enabled by DNA network is one of the most essential components of cellular manipulation.Hence,the DNA network is indispensable and irreplaceable to cell manipulation that it is a versatile tool for the understanding of basic laws of living life and treatments of diseases,such as cell isolation,cell delivery,and cell interference.Herein,the construction of polymeric DNA network is briefly introduced from the aspects of assembly modules,construction methods,and properties.
基金1226 Engineering Health Major Project(Nos.BWS17J028 and AWS16J018)。
文摘Metabolites can directly reflect and modulate cell responses and phenotypical changes by influencing energy balances,intercellular signals,and many other cellular functions throughout the lifespan of cells.Taking into account the heterogeneity of cells,single-cell metabolite analysis offers an insight into the functional process within one cell.Microfluidics as a powerful tool has attracted significant interest in the single-cell metabolite analysis field.The microfluidic platform is possible to observe,classify,and stimulate individual cells.It can also transport single-cell to subsequent analysis steps in a fast and controllable way to determine and analyze the composition and content of metabolites.The reviews of topics in microfluidics for single-cell metabolite analysis have been published in the past few years.However,most of them focused on metabolite analysis with mass spectrometry.Here,we covered the advances of microfluidic devices for single-cell metabolite analysis,with a focus on single-cell isolation and manipulation.What is more,we summarized the detection methods and applications of single-cell metabolites.
文摘BACKGROUND Psoriasis is a chronic autoimmune disease that usually manifests as a red scaly epidermis,induration,and hyperproliferation of basal keratinocytes.About 2%of the world’s population suffers from psoriasis but there are no clear therapeutics yet.Recently,mesenchymal stem cells(MSCs)have been regarded as a therapeutic alternative for autoimmune diseases,as they possess immunosuppressive effects without risks.Human umbilical cord-derived MSCs effectively regulate immune cells and are characterized by low immunogenicity,which has many advantages in treating immune diseases.CASE SUMMARY The patient was a 47-year-old male,diagnosed with psoriasis in 1995.He had received various treatments for 25 years,but the psoriatic condition was not significantly improved.He was given three rounds of minimally manipulated umbilical cord-derived MSCs over 2 wk.The erythema gradually disappeared.Three months after the 1st round,all erythema completely disappeared,and the psoriasis did not recur.CONCLUSION Minimally manipulated umbilical cord-derived MSC transplantation can potentially treat patients who suffer from psoriasis.
文摘BACKGROUND Alopecia areata(AA)is a common autoimmune disease characterized by hair loss.AA appears in extensive forms,such as progressive and diffusing hair loss(diffuse AA),a total loss of scalp hair(alopecia totalis),and complete loss of hair over the entire body(alopecia universalis).Recently,mesenchymal stem cells(MSCs)have been identified as a therapeutic alternative for autoimmune diseases.For this reason,preclinical and case studies of AA and related diseases using MSCs have been conducted.CASE SUMMARY Case 1:A 55-year-old woman suffered from AA in two areas of the scalp.She was given 15 rounds of minimally manipulated umbilical cord-MSCs(MM-UC-MSCs)over 6 mo.The AA gradually improved 3 mo after the first round.The patient was cured,and AA did not recur.Case 2:A 30-year-old woman,with history of local steroid hormone injections,suffered from AA in one area on the scalp.She was given two rounds of MM-UC-MSCs over 1 mo.The AA immediately improved after the first round.The patient was cured,and AA did not recur.Case 3:A 20-year-old woman,who was diagnosed with alopecia universalis at the age of 12,was given 14 rounds of MM-UC-MSCs over 12 mo.Her hair began to grow about 3 mo after the first round.The patient was cured,and alopecia universalis did not recur.CONCLUSION MM-UC-MSC transplantation potentially treats patients who suffer from AA and related diseases.
基金This study was financially supported by Grant-in-Aid for JSPS Fellows Number 13J03580Grant-in-Aid for Scientific Research on Innovative Areas(No.23106002)(No.26630094).
文摘We propose a novel on-chip 3D cell rotation method based on a vibration-induced flow.When circular vibration is applied to a microchip with micropillar patterns,a highly localized whirling flow is induced around the micropillars.The direction and velocity of this flow can be controlled by changing the direction and amplitude of the applied vibration.Furthermore,this flow can be induced on an open chip structure.In this study,we adopted a microchip with three micropillars arranged in a triangular configuration and an xyz piezoelectric actuator to apply the circular vibration.At the centre of the micropillars,the interference of the vibration-induced flows originating from the individual micropillars induces rotational flow.Consequently,a biological cell placed at this centre rotates under the influence of the flow.Under three-plane circular vibrations in the xy,xz or yz plane,the cell can rotate in both the focal and vertical planes of the microscope.Applying this 3D cell rotation method,we measured the rotational speeds of mouse oocytes in the focal and vertical planes as 63.7±4.0°s^(−1) and 3.5±2.1°s^(−1),respectively.Furthermore,we demonstrated the transportation and rotation of the mouse oocytes and re-positioned their nuclei into a position observable by microscope.
基金The project supported by the Hong Kong Research Grants Council (CUHK4215/01)
文摘The development of a novel polymer-based micro robotic gripper that can be actuated in a fluidic medium is presented in this paper.Our current work is to explore new materials and designs for thermal actuators to achieve micromanipulation of live biological cells.We used parylene C to encapsulate a metal heater,resulting in effectively a tri-layered thermal actuator.Parylene C is a bio-compatible dielectric polymer that can serve as a barrier to various gases and chemicals. Therefore,it is suitable to serve as a thermal/electrical/chemical isolation material for protecting the metal heater from exposing to an aqueous environment.We have demonstrated parylene actuators (2mm×100μm×0.5μm)to operate in an aqueous environment using 10 to 80mW input power.The temperature of these actuators at full deflection was estimated to be～60℃,which is much lower than the typical requirement of>100℃ to actuate other conventional MEMS actuators.Danio rerio follicles in fluidic medium were captured successfully using these actuators.Moreover,these actuators were found to be responsive to moderate rise in environmental temperature,and hence,we could vary the fluidic medium temperature to actuate trimorphs on a chip without any input of electrical energy, i.e.,raising the fluidic temperature from 23℃ to 60℃ could actuate the trimorphs to grasp follicles of ～1mm size in diameter.At 60℃,the embryos inside the follicles were observed to be alive,i.e.,they were still moving in the biological fluid isolated by the follicle membrane.The smallest follicles grasped were～500μm in diameter using 800μm×130μm×0.6μm actuators.The fabrication process,modeling, and optimization of the trimorph actuators are presented.Based on the successful operation of these polymer-based actuators,we are currently developing multifinger thermal microgrippers for cellular grasping and manipulation,which can potentially be hybridly integrated with circuits for computer control.
