Selective activation of photoactivatable fluorescent protein based on binary holography.

The ability to deliver laser doses to different target locations with high spatial and temporal resolution has been a long-sought goal in photo-stimulation and optogenetics research via, for example, photoactivatable proteins. These light-sensitive proteins undergo conformational changes upon photoactivation, serving functions such as triggering fluorescence, modulating ion channel activities, or initiating biochemical reactions within cells. Conventionally, photo-stimulation on light-sensitive proteins is performed by serially scanning a laser focus or via 2D projection, which is limited by relatively low spatiotemporal resolution. In this work, we present a programmable two-photon stimulation method based on a digital micromirror device (DMD) and binary holography to perform the activation of photoactivatable green fluorescent protein (PAGFP) in live cells. This method achieved grayscale and 3D selective PAGFP activation with subcellular resolution. In the experiments, we demonstrated the 3D activation capability and investigated the diffusion dynamics of activated PAGFP on the cell membrane. A regional difference in cell membrane diffusivity was observed, indicating the great potential of our approach in interrogating the spatiotemporal dynamics of cellular processes inside living cells.

Integrative analysis and risk model construction for super­enhancer­related immune genes in clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common type of kidney cancer associated with poor prognosis, and accounts for the majority of RCC-related deaths. The lack of comprehensive diagnostic and prognostic biomarkers has limited further understanding of the pathophysiology of ccRCC. Super-enhancers (SEs) are congregated enhancer clusters that have a key role in tumor processes such as epithelial-mesenchymal transition, metabolic reprogramming, immune escape and resistance to apoptosis. RCC may also be immunogenic and sensitive to immunotherapy. In the present study, an Arraystar human SE-long non-coding RNA (lncRNA) microarray was first employed to profile the differentially expressed SE-lncRNAs and mRNAs in 5 paired ccRCC and peritumoral tissues and to identify SE-related genes. The overlap of these genes with immune genes was then determined to identify SE-related immune genes. A model for predicting clinical prognosis and response to immunotherapy was built following the comprehensive analysis of a ccRCC gene expression dataset from The Cancer Genome Atlas (TCGA) database. The patients from TCGA were divided into high- and low-risk groups based on the median score derived from the risk model, and the Kaplan-Meier survival analysis showed that the low-risk group had a higher survival probability. In addition, according to the receiver operating characteristic curve analysis, the risk model had more advantages than other clinical factors in predicting the overall survival (OS) rate of patients with ccRCC. Using this model, it was demonstrated that the high-risk group had a more robust immune response. Furthermore, 61 potential drugs with half-maximal inhibitory concentration values that differed significantly between the two patient groups were screened to investigate potential drug treatment of ccRCC. In summary, the present study provided a novel index for predicting the survival probability of patients with ccRCC and may provide some insights into the mechanisms through which SE-related immune genes influence the diagnosis, prognosis and potential treatment drugs of ccRCC.

Blood Circulation Assessment by Steadily Fluorescent Near-Infrared-II Aggregation-Induced Emission Nano Contrast Agents.

The dysfunction of the blood circulation system typically induces acute or chronic ischemia in limbs and vital organs, with high disability and mortality. While conventional tomographic imaging modalities have shown good performance in the diagnosis of circulatory diseases, multiple limitations remain for real-time and precise hemodynamic evaluation. Recently, fluorescence imaging in the second region of the near-infrared (NIR-II, 1000-1700 nm) has garnered great attention in monitoring and tracing various biological processes in vivo due to its advantages of high spatial-temporal resolution and real-time feature. Herein, we employed NIR-II imaging to carry out a blood circulation assessment by aggregation-induced emission fluorescent aggregates (AIE nano contrast agent, AIE NPs). Thanks to the longer excited wavelength, enhanced absorptivity, higher brightness in the NIR-II region, and broader optimal imaging window of the AIE NPs, we have realized a multidirectional assessment for blood circulation in mice with a single NIR-II imaging modality. Thus, our work provides a fluorescence contrast agent platform for accurate hemodynamic assessment.

Polyetheretherketone (PEEK) as a Potential Material for the Repair of Maxillofacial Defect Compared with E-poly(tetrafluoroethylene) (e-PTFE) and Silicone.

Silicone and e-poly(tetrafluoroethylene) (e-PTFE) are the most commonly used artificial materials for repairing maxillofacial bone defects caused by facial trauma and tumors. However, their use is limited by poor histocompatibility, unsatisfactory support, and high infection rates. Polyetheretherketone (PEEK) has excellent mechanical strength and biocompatibility, but its application to the repair of maxillofacial bone defects lacks a theoretical basis. The microstructure and mechanical properties of e-PTFE, silicone, and PEEK were evaluated by electron microscopy, BOSE machine, and Fourier transformed infrared spectroscopy. Mouse fibroblast L929 cells were incubated on the surface of the three materials to assess cytotoxicity and adhesion. The three materials were implanted onto the left femoral surface of 90 male mice, and samples of the implants and surrounding soft tissues were evaluated histologically at 1, 2, 4, 8, and 12 weeks post-surgery. PEEK had a much higher Young's modulus than either e-PTFE or silicone (p < 0.05 each), and maintained high stiffness without degradation long after implantation. Both PEEK and e-PTFE facilitated L929 cell adhesion, with PEEK having lower cytotoxicity than e-PTFE and silicone (p < 0.05 each). All three materials similarly hindered the motor function of mice 12 weeks after implantation (p > 0.05 each). Connective tissue ingrowth was observed in PEEK and e-PTFE, whereas a fibrotic peri-prosthetic capsule was observed on the surface of silicone. The postoperative infection rate was significantly lower for both PEEK and silicone than for e-PTFE (p < 0.05 each). PEEK shows excellent biocompatibility and mechanical stability, suggesting that it can be effective as a novel implant to repair maxillofacial bone defects.

Fast Delivery of Multifunctional NIR-II Theranostic Nanoaggregates Enabled by the Photoinduced Thermoacoustic Process.

Multifunctional nanoaggregates are widely used in cancer phototheranostics. However, it is challenging to construct their multifunctionality with a single component, and deliver them rapidly and efficiently without complex modifications. Herein, a NIR-absorbing small molecule named TBT-2(TP-DPA) is designed and certify its theranostic potentials. Then, their nanoaggregates, which are simply encapsulated by DSPE-PEG, demonstrate a photothermal efficiency of 51% while keeping a high photoluminescence quantum yield in the NIR region. Moreover, the nanoaggregates can be excited and delivered by an 808 nm pulse laser to solid tumors within only 40 min. The delivery efficiency and theranostic efficacy are better than that of the traditional enhanced permeability and retention (EPR) effect (generally longer than 24 hours). This platform is first termed as the photoinduced thermoacoustic (PTA) process, and confirm its application requires both NIR-responsive materials and pulse laser irradiation. This study not only inspires the design of multifunctional nanoaggregates, but also offers a feasible approach to their fast delivery. The platform reported here provides a promising prospect to boost the development of multifunctional theranostic drugs and maximize the efficacy of used medicines for their clinical applications.

Design of One-for-All Near-Infrared Aggregation-Induced Emission Nanoaggregates for Boosting Theranostic Efficacy.

Fluorescence-guided phototherapy, including photodynamic and photothermal therapy, is considered an emerging noninvasive strategy for cancer treatments. Organic molecules are promising theranostic agents because of their facile construction, simple modification, and good biocompatibility. Organic systems that integrated multifunctionalities in a single component and achieved high efficiency in both imaging and therapies are rarely reported as the inherently competitive energy relaxation pathways are hard to modulate, and fluorescence quenching occurs upon molecular aggregation. Herein, a versatile theranostic platform with near-infrared emission, high fluorescence quantum yield, robust reactive oxygen species production, and excellent photothermal conversion efficiency was developed based on an aggregation-induced emission luminogen, namely, TPA-TBT. In vivo studies revealed that the TPA-TBT nanoaggregates exhibit outstanding photodynamic and photothermal therapy efficacy to ablate tumors inoculated in a mouse model. This work offers a design strategy to develop one-for-all cancer theranostic agents by modulating and utilizing the relaxation energy of excitons in full.

Recent Advances in the Enzyme-Activatable Organic Fluorescent Probes for Tumor Imaging and Therapy.

The exploration of advanced probes for cancer diagnosis and treatment is of high importance in fundamental research and clinical practice. In comparison with the traditional "always-on" probes, the emerging activatable probes enjoy advantages in promoted accuracy for tumor theranostics by specifically releasing or activating fluorophores at the targeting sites. The main designing principle for these probes is to incorporate responsive groups that can specifically react with the biomarkers (e. g., enzymes) involved in tumorigenesis and progression, realizing the controlled activation in tumors. In this review, we summarize the latest advances in the molecular design and biomedical application of enzyme-responsive organic fluorescent probes. Particularly, the fluorophores can be endowed with ability of generating reactive oxygen species (ROS) to afford the photosensitizers, highlighting the potential of these probes in simultaneous tumor imaging and therapy with rational design. We hope that this review could inspire more research interests in the development of tumor-targeting theranostic probes for advanced biological studies.

Synthesis of aligned porous polyethylene glycol/silk fibroin/hydroxyapatite scaffolds for osteoinduction in bone tissue engineering.

BACKGROUND: The physical factors of the extracellular matrix have a profound influence on the differentiation behavior of mesenchymal stem cells. In this study, the effect of the biophysical microenvironment on rat bone marrow mesenchymal stem cell (BMSC) osteogenesis was studied both in vitro and in vivo. METHODS: To prepare cell culture scaffolds of varying stiffness, increasing amounts of hydroxyapatite (HAp) were mixed into a polyethylene glycol/silk fibroin (PEG/SF) solution. The amount of HAp ranged from 25 to 100 mg, which provided for different ratios between HAp and the PEG/SF composite. In vitro, the effect of stiffness on the osteogenic differentiation of rat BMSCs was studied. The outcome measures, which were verified in vivo, included the protein expression of runt-related transcription factor 2 and osteocalcin, alkaline phosphatase activity, and the mRNA expression of osteogenesis-related markers. RESULTS: Increasing amounts of HAp resulted in an increased elastic modulus of the cell culture scaffolds. The PEG/SF/HAp fabricated with HAp (50 mg) significantly increased cell adhesion and viability (p < 0.05) as well as the expression of all the osteogenesis-related markers (p < 0.05). CONCLUSIONS: We developed a novel cell culture scaffold and demonstrated that substrate stiffness influenced the osteogenic differentiation of rat BMSCs.

A Novel Anterior Transpedicular Screw Artificial Vertebral Body System for Lower Cervical Spine Fixation: A Finite Element Study.

A finite element model was used to compare the biomechanical properties of a novel anterior transpedicular screw artificial vertebral body system (AVBS) with a conventional anterior screw plate system (ASPS) for fixation in the lower cervical spine. A model of the intact cervical spine (C3-C7) was established. AVBS or ASPS constructs were implanted between C4 and C6. The models were loaded in three-dimensional (3D) motion. The Von Mises stress distribution in the internal fixators was evaluated, as well as the range of motion (ROM) and facet joint force. The models were generated and analyzed by mimics, geomagic studio, and ansys software. The intact model of the lower cervical spine consisted of 286,382 elements. The model was validated against previously reported cadaveric experimental data. In the ASPS model, stress was concentrated at the connection between the screw and plate and the connection between the titanium mesh and adjacent vertebral body. In the AVBS model, stress was evenly distributed. Compared to the intact cervical spine model, the ROM of the whole specimen after fixation with both constructs is decreased by approximately 3 deg. ROM of adjacent segments is increased by approximately 5 deg. Facet joint force of the ASPS and AVBS models was higher than those of the intact cervical spine model, especially in extension and lateral bending. AVBS fixation represents a novel reconstruction approach for the lower cervical spine. AVBS provides better stability and lower risk for internal fixator failure compared with traditional ASPS fixation.

Pioglitazone inhibits growth of human retinoblastoma cells via regulation of NF-κB inflammation signals.

OBJECTIVES: We aimed to study the antitumor effects of the PPARγ agonist pioglitazone on human retinoblastoma. METHODS: The effects of pioglitazone on cell proliferation and apoptosis of the human retinoblastoma Y79 cells were investigated by MTT assay and Hoechst 33258 staining assay. The apoptosis related protein levels were detected by western blot. Inflammationary factors analysis was evaluated by western blot and ELISA. The effect of pioglitazone on nuclear factor-kappa B (NF-κB)-dependent reporter gene transcription induced by LPS was analyzed by NF-κB-luciferase assay. Then human retinoblastoma Y79 cells were subcutaneously transplanted in BALB/c nude mice and the animals were treated with pioglitazone to verify its antitumor effect in vivo. RESULTS: Our data revealed that pioglitazone suppressed the viability of Y79 cells dose- and time-dependently and induced apoptosis in Y79 cells in vitro. Molecular biology analysis found that pioglitazone could affect the apoptosis and inflammation related signal via modulating the activity of NF-κB signal. Also we found that pioglitazone could markedly reduce the growth of Y79 cells transplanted into the mice without causing significant side effects. CONCLUSIONS: Our results suggested that pioglitazone demonstrated antitumor activity against the human retinoblastoma Y79 cells by inhibiting cell growth, inducing apoptosis and modulating NF-κB pathway, and thus delayed tumor growth in vivo.

[The biomechanical study to evaluate tightening condition for AO lag screw depending on pull-out strength and interfragmentary compressive force].

The aim of this experimental study focused on the relationship between pull-out strength (POS) and interfragmentary compressive force (IFCF) of AO cancellous lag screw during tightening procedure. The 6.5 mm AO cancellous lag screw and synthetic cancellous bone were used for this research. The test contains rotation tests and the subsequent pull-out tests, to record the IFCF and POS under different tightening angle groups. The results of this study demonstrated the specific relationship between IFCF and POS and showed that they didn't reach the peak at the very same time. This study revealed the change of mechanical environment surrounding AO lag screw during tightening procedure and found the effective method to determine the optimum terminating time of AO lag screw inserting.

Biomechanical analysis of differential pull-out strengths of bone screws using cervical anterior transpedicular technique in normal and osteoporotic cervical cadaveric spines.

STUDY DESIGN: Biomechanical in vitro study. OBJECTIVE: To determine whether the peak pull-out force (PPF) of cervical anterior transpedicular screw (ATPS) fixed in osteoporotic vertebrae positively influence screw stability or not before and after fatigue. SUMMARY OF BACKGROUND DATA: Multilevel cervical spine procedures with osteoporosis can challenge the stability of current screw-and-plate systems. A second surgical posterior approach is coupled with potential risks of increased morbidity and complications. Hence, anterior cervical instrumentation that increases primary construct stability, while avoiding the need for posterior augmentation, would be valuable. METHODS: Sixty formalin-fixed vertebrae at different levels were randomly selected. The vertebrae were divided into healthy controls (groups A1, A2), osteoporotic controls (B1, B2), healthy ATPS groups (C1, C2), osteoporotic ATPS groups (D1, D2), and osteoporotic restoration controls (E1, E2). The procedure of ATPS insertion was simulated with 2 pilot holes being drilled on each side of 20 vertebral bodies that were implanted with either vertebral screw or polymethylmethacrylate. Each side randomly received either instant PPF or PPF beyond fatigue (2.5 Hz; 20,000 times). RESULTS: The prefatigue PPFs were significantly higher than the postfatigue PPFs in all groups (group A: 366.06 ± 58.78 vs. 248.93 ± 57.21 N; group B: 275.58 ± 23.18 vs. 142.79 ± 44.78 N; group C: 635.99 ± 185.28 vs. 542.57 ± 136.58 N; group D: 519.22 ± 122.12 vs. 393.16 ± 192.07 N, and group E: 431.78 ± 75.77 vs. 325.74 ± 95.10 N). The postfatigue PPFs were reduced by 32.00% (group A), 48.19% (group B), 14.69% (group C), 24.28% (group D), and 24.72% (group E). The acute and postfatigue PPFs of both control groups were significantly lower than that of ATPS groups (P < 0.05). The cyclic osteoporosis ATPS group achieved the same PPF compared with the vertebral restoration screw group. CONCLUSION: The findings of this study suggest that instant PPF and fatigue resistance capability of an ATPS fixation were significantly better than other control groups, especially in the osteoporotic vertebrae.

Load-bearing evaluation of spinal posterior column by measuring surface strain from lumbar pedicles. An in vitro study.

An understanding of the load transfer within spinal posterior column of lumbar spine is necessary to determine the influence of mechanical factors on potential mechanisms of the motion-sparing implant such as artificial intervertebral disc and the dynamic spine stabilization systems. In this study, a new method has been developed for evaluating the load bearing of spinal posterior column by the surface strain of spinal pedicle response to the loading of spinal segment. Six cadaveric lumbar spine segments were biomechanically evaluated between levels L1 and L5 in intact condition and the strain gauges were pasted to an inferior surface of L2 pedicles. Multidirectional flexibility testing used the Panjabi testing protocol; pure moments for the intact condition with overall spinal motion and unconstrained intact moments of ±8 Nm were used for flexion-extension and lateral bending testing. High correlation coefficient (0.967-0.998) indicated a good agreement between the load of spinal segment and the surface strain of pedicle in all loading directions. Principal compressive strain could be observed in flexion direction and tensile strain in extension direction, respectively. In conclusion, the new method seems to be effective for evaluating posterior spinal column loads using pedicles' surface strain data collected during biomechanical testing of spine segments.

[Six degree-of-freedom acquisition and analysis of jaw opening and closing with motion capture system].

OBJECTIVE: To explore the six degrees of freedom of jaw opening and closing movement with motion capture and analysis system to establish a quantitative method for studying mandibular movement and a digital basis for virtual reality study of mandibular movement. METHODS: In a male adult with normal dentition without temporomandibular joint disorders, 3 fluorescent markers were pasted in the upper dentition and 4 in the lower dentition. Six cameras of the motion capture system were arranged in a semi-circular fashion. The subject sat in front of the camera at an 80-cm distance with the Frankfort plane kept parallel to the horizontal plane. The degree-of-freedom (3 linear displacement and 3 angular displacement) of jaw opening and closing movement was obtained by collecting the marker motion. RESULTS: Six degrees of freedom of jaw opening and closing were obtained using the motion capture system. The maximum linear displacements of X, Y and Z axes were 5.888 089 cm, 0.782 269 cm, and 0.138 931 cm, and the minimum linear displacements were -3.649 83 cm, -35.961 2 cm, -5.818 63 cm, respectively. The maximum angular displacements of X, Y and Z axes were 0.760 088°, 2.803 753°, and 0.786 493°, with the minimum angular displacements of -2.526 18°, -0.625 94°, and -25.429 8°, respectively. Variations of linear displacements during jaw opening and closing occurred mainly in the Y axis, and those of angular displacement occurred mainly in the Z axis. CONCLUSION: The six degree-of-freedom of mandibular movement can be accurately obtained with the motion capture system to allow quantitative examination of the mandibular movement.

[Application of three-dimensional laser scanning-based maxillofacial soft tissue reconstruction in orthodontic treatment].

OBJECTIVE: To establish a convenient and rapid method for constructing a digital model of the maxillofacial soft tissue based on three-dimensional laser surface scanning to allow direct and accurate observation of the soft tissue changes in the course of orthodontic treatment. METHODS: The point cloud data of three-dimensional laser scanning of the maxillofacial region were acquired from a healthy woman with Angle Class I occlusion, who maintained a horizontal Frankfort plane during scanning with the scanner placed at a distance of 80 cm. The scanning was repeated twice after wearing the dental cast for an Angle Class I occlusion. The three-dimensional digital model of the maxillofacial soft tissue was constructed based on the point cloud using GeoMagic10.0 software. RESULTS: The high-resolution three-dimensional model of the maxillofacial soft tissue reconstructed allowed accurate observation of the distinct facial anatomical landmarks and represented directly the soft tissue changes in the process of orthodontic treatment by merging the models. Using the analytic tool provided by the software, this model also allowed direct quantitative measurement of the nasolabial angle and the distances from the esthetic plane to the upper lip, labral inferior, and mentolabial sulcus, which were 111.86°, -3.57 mm, -2.54 mm, and 3.95 mm before orthodontic treatment as compared to 114.31°, -2.73 mm, -1.06 mm, and 3.46 mm during treatment, and 116.53°, -0.15 mm, 0.64 mm, and 3.11 mm after the treatment, respectively. CONCLUSION: Three-dimensional laser surface scanning enables accurate and rapid construction of the digital model of the facial soft tissues, which may provide valuable assistance in orthodontic treatment.

[Three-dimensional reconstruction and anatomic variation of the portal vein based on 64-slice spiral CT data].

OBJECTIVE: To investigate the three-dimensional reconstruction methods of the portal vein using 64-slice spiral CT data and the anatomical variation of the portal vein. METHODS: Three-dimensional reconstruction of the portal vein was performed using Mimics software based on the 64-slice spiral CT data of 64 cases. Each model of the portal vein and its branches was evaluated according to the presentation rate, depiction quality and anatomic variation. RESULTS: The reconstructed model showed a depiction rates of 100% for the 4-grade branches of the portal vein. The stem of the portal vein and the left and right branches of the level III or above were all displayed, but in 2 cases the superior mesenteric vein and in 1 case the spleen vein was displayed only to the level IV. Of the 64 cases, 50 (78.1%) had normal portal vein and 14 (21.9%) showed anatomical variations. CONCLUSION: The 3D model vividly mimics the anatomic variations of the portal vein to provide valuable information for surgical plans.

[Construction of the digital models of masseter and temporal muscles with three-dimensional laser scanning].

OBJECTIVE: To explore a new method for establishing digital models of the masseter and temporal muscles and superficial soft tissue using three-dimensional laser scanning technique. METHODS: One adult male cadaveric head without malformation was dissected to expose the superficial portion of the masseter and temporalis. Multiple aspects of the sample were scanned with three-dimensional laser scanning system, and the point clouds of the masseter and temporal muscles were generated. The specimen was scanned again after the masseter and temporal muscles had been removed. The digital model of the muscles was reconstructed with the point clouds using Geomagic software, and the morphology of the muscle model was observed and measured. RESULTS: The 3-D digital models of the masseter and temporal muscles with the anatomical characteristics were reconstructed based on the point clouds using Geomagic 8.0 software. CONCLUSION: The digital model of the muscles can vividly demonstrate the muscle contours, which provides a basis for morphological study and biomechanical analysis of the muscles.

[Three-dimensional design of surgery for mandibular retrusion].

OBJECTIVE: To explore the feasibility of surgical design for mandibular retrusion using three-dimensional software. METHODS: Three-dimensional reconstruction was performed by Mimics software based on the preoperative CT data. The model of the maxillofacial region was imported into Rapidform software for measuring the associated parameters and Geomagic software for simulation of osteotomy. The reliability of the virtual operation was validated during the surgery. RESULTS: The model of mandibular retrusion was reconstructed and successfully used to simulate the surgery. The simulation result was applied in subsequent actual surgery and good surgical outcomes were achieved. CONCLUSION: The three-dimensional software can be used to simulate the surgery for mandibular retrusion and improve the predictability and accuracy of the surgery.