A reflection-mode photoacoustic microscope using a hollow focused ultrasound transducer is developed for highresolution in vivo imaging.A confocal structure of the laser and the ultrasound is used to improve the system resolution.The axial and lateral resolutions of the system are measured to be ～ 32 μm and ～ 58 μm,respectively.Ex vivo and in vivo modes are tested to validate the imaging capability of the photoacoustic microscope.The adjacent vein and artery can be seen clearly from the reconstructed photoacoustic images.The results demonstrate that the reflectionmode photoacoustic microscope can be used for high-resolution imaging of micro-blood vessels,which would be of great benefit for monitoring the neovascularization in tumor angiogenesis.
Sirtuins comprise a family of enzymes implicated in the determination of organismal lifespan in yeast and the nematode.Human sirtuin SIRT1 has been shown to deacetylate several proteins in a NADt-dependent manner.It is reported that SIRT1 regulates physiological processes including senescence,fat metabolism,glucose homeostasis,apoptosis,and neurodegeneration.In general,SIRT1 has initially been thought to represent an exclusive nuclear protein.However,depending on the cell lines and organisms examined,a partial or temporary cytoplasmic localization was observed in murine pancreatic beta cells and neonatal rat cardiomyocytes.Since SIRT1 deacetylates both histone and nonhistone-proteins,such as a number of transcription factors,changes in subcellular localization probably play a role in the regulation of its function.In the present studies,we investigated the subcellular localization of SIRT1 in response to growth factor deprivation in African green monkey SV40-transformed kidneyfibroblast cells(COS-7).Using SIRT1-EGFPfluorescence reporter,we found that SIRT1 localized to nucleus in physiological conditions.We devised a model enabling cell senescence via growth factor deprivation and found that SIRT1 partially translocated to cytosol under the treatment,suggesting a reduced level of SIRT1 activity.We found PI3K/Akt pathway was involved in the inhibition of SIRT1's cytosolic translocation,because inhibition of these kinases significantly decreased the amount of SIRT1 maintained in nucleus.Taken together,we demonstrate that growth factor deprivation induces cytosolic translocation of SIRT1,which suggests a possible connection between cytoplasm-localized SIRT1 and the aging process and provides a new application of single moleculefluorescence imaging of the molecule events in living cells.
Microglial activation plays an important role in neurodegenerative diseases.Once activated,they have macrophage-like capabilities,which can be beneficial by phagocytosis and harmful by se-cretion of neurotoxins.However,the resident microglia always fail to trigger an effective pha-gocytic response to clear dead cells or Aβdeposits during the progression of neurodegeneration.Therefore,the regulation of microglial phagocytosis is considered a useful strategy in searchingfor neuroprotective treatments.In this study,our results showed that low-power laser iradiation(LPLI)(20 J/cm²)could enhance microglial phagocytic function in LPS-activated microglia.Wefound that LPLI-mediated microglial phagocytosis is a Rac-1-dependent actin-based process,that a constitutively activated form of Rac1(RaclQ61L)induced a higher level of actin pol-ymerization than cells transfected with wild-type Racl,whereas a dominant negative form ofRacl(RaclT17N)markedly suppressed actin polymerization.In addition,the involvement of Racl activation after LPLI treatment was also observed by using a Raichu fluorescence resonance energy transfer(FRET)-based biosensor.We also found that PI3K/Akt pathway was required inthe LPLI-induced Raci activation.Our research may provide a feasible therapeutic approach tocontrol the progression of neurodegenerative diseases.
Insulin resistance is a hallmark of the metabolic syndrome and type 2 diabetes.Dysfunction of PI-3K/Akt signaling was involved in insulin resistance.Glucose transporter 4(GLUT4)is a keyfactor for glucose uptake in muscle and adipose tissues,which is closely regulated by Pi-3K/Aktsignaling in response to insulin treatment.Low-power laser irradiation(LPLI)has been shown toregulate various physiological processes and induce the synthesis or release of multiple moleculessuch as growth factors,which(especially red and near infrared light)is mainly through theactivation of mitochondrial respiratory chain and the initiation of intracellular signaling path-ways.Nevertheless,it is unclear whether LPLI could promote glucose uptake through activationof PI-3K/Akt/GLUT4 signaling in 3T3L-1 adipocytes.In this study,we investigated how LPLIpromoted glucose uptake through activation of PI-3K/Akt/GLUT4 signaling path way.Here,we showed that GLUT4 was localized to the Golgi apparatus and translocated from cytoplasm tocytomembrane upon LPLI treatment in 3T3L-1 adipocytes,which enhanced glucose uptake.Moreover,we found that glucose uptake was mediated by the PI3-K/Akt2 signaling,but notAkt1 upon LPLI treatment with Akt isoforms gene silence and PI3-K/Akt inhibitors.Collec-tively,our results indicate that PI3-K/Akt2/GLUT4 signaling act as the key regulators forimprovement of glucose uptake under LPLI treatment in 3T3L-i adipocytes.More importantly,our findings suggest that activation of PI3-K/Akt2/GLUT4 signaling by LPLI may provideguidance in practical applications for promotion of glucose uptake in insulin-resistant adiposetissue.
We introduce a corrected sinusoidal-wave drag force method(SDFM) into optical tweezers to calibrate the trapping stiffness of the optical trap and conversion factor(CF) of photodetectors. First, the theoretical analysis and experimental result demonstrate that the correction of SDFM is necessary, especially the error of no correction is up to 11.25% for a bead of 5 μm in diameter. Second, the simulation results demonstrate that the SDFM has a better performance in the calibration of optical tweezers than the triangular-wave drag force method(TDFM) and power spectrum density method(PSDM) at the same signal-to-noise ratio or trapping stiffness. Third, in experiments, the experimental standard deviations of calibration of trapping stiffness and CF with the SDFM are about less than 50% of TDFM and PSDM especially at low laser power.Finally, the experiments of stretching DNA verify that the in situ calibration with the SDFM improves the measurement stability and accuracy.
Contrast agents are attracting a great deal of attention in photoacoustic inaging.Here weintroduce an exogenous contrast agent that provides high photoacoustic signal amplitude at thenear-infrared wavelength._Our_agents consist_of Indocyanine green(ICG)and phospholi-pid-polyethylene glycol(PL-PEG),entitled ICG-PL-PEG nanoparticles,These nanoparticleshave overcome numerous limitations of ICG,such as poor aqueous stability,concentration-dependent aggregation and lack of target specificity.ICG-PL-PEG nanoparticles are bio-compatible and relatively nontoxic.All the components of ICG-PL-PEG nanoparticles havebeen approved for human use.Upon pulsed laser irradiation,the nanoparticles are more eficient inproducing photoacoustic waves than ICG alone.The results showed that ICG-PL-PEG nano-particles act as good contrast agents for photoacoustic imaging.These unique ICG-PL-PEGnanoparticles have great potential in clinical applications.
