See farther and more: a master-slave UAVs based synthetic optical aperture imaging system with wide and dynamic baseline.

An open challenge remained in designing an optical system to capture the aerial image with a wide field of view (FoV) and high resolution. The optical system of one camera from a single unmanned aerial vehicle (UAV) can hardly promise the FoV and resolution. The conventional swarm UAVs can form the camera array with a short or fixed baseline. They can capture the images with a wide FoV and high resolution, but the cost is the requirement of many UAVs. We aim to design a camera array with a wide and dynamic baseline to reduce the demand for UAVs to organize a synthetic optical aperture. In this thought, we propose a master-slave UAVs-based synthetic optical aperture imaging system with a wide and dynamic baseline. The system consists of one master UAV and multiple slave UAVs. Master and slave UAVs provide the global and local FoVs, respectively, and improve the efficiency of image acquisition. In such a system, fusing UAV images becomes a new challenge due to two factors: (i) the small FoV overlap of slave UAVs and (ii) the gap in resolution scale from slave to master UAV images. To deal with it, a coarse-to-fine stitching method is proposed to stitch up the multi-view images into one to obtain a wide FoV with high resolution. A video stabilization method has also been designed for the proposed imaging system. Challenges caused by wide and dynamic baselines can thus be solved by the above methods. Actual data experiments demonstrate that the proposed imaging system achieves high-quality imaging results.

Ag(I)-Mediated Annulation of 2-(2-Enynyl)pyridines and Propargyl Amines to Access 1-(2H-Pyrrol-3-yl)indolizines.

An effective Ag(I)-mediated annulation of 2-(2-enynyl)pyridines and propargyl amines was developed, unexpectedly affording a broad range of functionalized 1-(2H-pyrrol-3-yl)indolizines in moderate to excellent yields. The developed method is characterized by operational simplicity, ready availability of starting materials, high regioselectivity, and broad substrate scope under mild reaction conditions. The Ag(I)-promoted cyclization of 2-(2-enynyl)pyridines and propargyl amines possibly results in the formation of the spiroindolizine, the ring-opening rearrangement of which may give the 1-(2H-pyrrol-3-yl)indolizine. Furthermore, a gram-scale reaction and synthetic transformations are also studied.

Efficacy and safety of tafolecimab in Chinese patients with heterozygous familial hypercholesterolemia: a randomized, double-blind, placebo-controlled phase 3 trial (CREDIT-2).

BACKGROUND: Heterozygous familial hypercholesterolemia (HeFH) is largely underdiagnosed and undertreated in China where few patients achieved recommended target levels of low density lipoprotein cholesterol (LDL-C). We conducted the first randomized, placebo-controlled clinical trial in Chinese patients with HeFH to assess the efficacy and safety of tafolecimab, a novel fully human proprotein convertase subtilisin/kexin type 9 (PCSK9) monoclonal antibody. METHODS: Patients diagnosed with HeFH by Simon Broome criteria and on a stable lipid-lowering therapy for at least 4 weeks were randomized 2:2:1:1 to receive subcutaneous tafolecimab 150 mg every 2 weeks (Q2W), tafolecimab 450 mg every 4 weeks (Q4W), placebo Q2W or placebo Q4W in the 12-week double-blind treatment period. After that, participants received open-label tafolecimab 150 mg Q2W or 450 mg Q4W for 12 weeks. The primary endpoint was the percent change from baseline to week 12 in LDL-C levels. Secondary endpoints included proportion of participants achieving ≥50% LDL-C reductions and proportion of participants with LDL-C <1.8 mmol/L at week 12 and 24, the change from baseline to week 12 in non-high density lipoprotein cholesterol (non-HDL-C), apolipoprotein B and lipoprotein(a) levels, as well as the change from baseline to week 24 in lipid levels. RESULTS: In total, 149 participants were randomized and 148 received at least one dose of the study treatment. At week 12, tafolecimab treatment induced significant reductions in LDL-C levels (treatment difference versus placebo [on-treatment estimand]: -57.4% [97.5% CI, -69.2 to -45.5] for 150 mg Q2W; -61.9% [-73.4 to -50.4] for 450 mg Q4W; both P <0.0001). At both dose regimens, significantly more participants treated with tafolecimab achieved ≥50% LDL-C reductions or LDL-C <1.8 mmol/L at week 12 as compared with corresponding placebo groups (all P <0.0001). Meanwhile, non-HDL-C, apolipoprotein B and lipoprotein(a) levels were significantly reduced in the tafolecimab groups at week 12. The lipid-lowering effects of tafolecimab were maintained till week 24. During the double-blind treatment period, the most commonly-reported adverse events in the tafolecimab groups included upper respiratory tract infection, increased blood creatine phosphokinase, increased alanine aminotransferase, increased aspartate aminotransferase and hypertension. CONCLUSIONS: Tafolecimab administered either 150 mg Q2W or 450 mg Q4W yielded significant and persistent reductions in LDL-C levels and showed a favorable safety profile in Chinese patients with HeFH. TRIAL REGISTRATION: ClinicalTrials.gov, NCT04179669.

A non-genetic engineering platform for rapidly generating and expanding cancer-specific armed T cells.

BACKGROUND: Cancer-specific adoptive T cell therapy has achieved successful milestones in multiple clinical treatments. However, the commercial production of cancer-specific T cells is often hampered by laborious cell culture procedures, the concern of retrovirus-based gene transfection, or insufficient T cell purity. METHODS: In this study, we developed a non-genetic engineering technology for rapidly manufacturing a large amount of cancer-specific T cells by utilizing a unique anti-cancer/anti-CD3 bispecific antibody (BsAb) to directly culture human peripheral blood mononuclear cells (PBMCs). The anti-CD3 moiety of the BsAb bound to the T cell surface and stimulated the differentiation and proliferation of T cells in PBMCs. The anti-cancer moiety of the BsAb provided these BsAb-armed T cells with the cancer-targeting ability, which transformed the naïve T cells into cancer-specific BsAb-armed T cells. RESULTS: With this technology, a large amount of cancer-specific BsAb-armed T cells can be rapidly generated with a purity of over 90% in 7 days. These BsAb-armed T cells efficiently accumulated at the tumor site both in vitro and in vivo. Cytotoxins (perforin and granzyme) and cytokines (TNF-α and IFN-γ) were dramatically released from the BsAb-armed T cells after engaging cancer cells, resulting in a remarkable anti-cancer efficacy. Notably, the BsAb-armed T cells did not cause obvious cytokine release syndrome or tissue toxicity in SCID mice bearing human tumors. CONCLUSIONS: Collectively, the BsAb-armed T cell technology represents a simple, time-saving, and highly safe method to generate highly pure cancer-specific effector T cells, thereby providing an affordable T cell immunotherapy to patients.

UAV-YOLOv8: A Small-Object-Detection Model Based on Improved YOLOv8 for UAV Aerial Photography Scenarios.

Unmanned aerial vehicle (UAV) object detection plays a crucial role in civil, commercial, and military domains. However, the high proportion of small objects in UAV images and the limited platform resources lead to the low accuracy of most of the existing detection models embedded in UAVs, and it is difficult to strike a good balance between detection performance and resource consumption. To alleviate the above problems, we optimize YOLOv8 and propose an object detection model based on UAV aerial photography scenarios, called UAV-YOLOv8. Firstly, Wise-IoU (WIoU) v3 is used as a bounding box regression loss, and a wise gradient allocation strategy makes the model focus more on common-quality samples, thus improving the localization ability of the model. Secondly, an attention mechanism called BiFormer is introduced to optimize the backbone network, which improves the model's attention to critical information. Finally, we design a feature processing module named Focal FasterNet block (FFNB) and propose two new detection scales based on this module, which makes the shallow features and deep features fully integrated. The proposed multiscale feature fusion network substantially increased the detection performance of the model and reduces the missed detection rate of small objects. The experimental results show that our model has fewer parameters compared to the baseline model and has a mean detection accuracy higher than the baseline model by 7.7%. Compared with other mainstream models, the overall performance of our model is much better. The proposed method effectively improves the ability to detect small objects. There is room to optimize the detection effectiveness of our model for small and feature-less objects (such as bicycle-type vehicles), as we will address in subsequent research.

Diterpenoids from Sauropus spatulifolius Leaves with Antimicrobial Activities.

As a plant used in both food and medicine, Sauropus spatulifolius is consumed widely as a natural herbal tea, food source, and Chinese medicine. Inspired by its extensive applications, we conducted a systematic phytochemical study of the leaves of S. spatulifolius. Thirteen new diterpenoids, sauspatulifols A-M (1-13), including four ent-cleistanthane-type diterpenoids (1-4), eight 15,16-di-nor-ent-cleistanthane-type diterpenoids (5-12), and one 17-nor-ent-pimarane-type diterpenoid (13) as well as one known diterpenoid, cleistanthol (14), were isolated. All of these diterpenoids feature a 2α,3α-dihydroxy unit within the A ring, and their structures were elucidated by spectroscopic data analysis, electronic circular dichroism calculations, and single-crystal X-ray diffraction analysis. Compound 14 displayed moderate inhibitory activity against Staphylococcus aureus, Staphylococcus epidermidis, Bacillus subtilis, and Shigella flexneri with the same minimum inhibitory concentration value of 12 µg/mL as well as activity against vesicular stomatitis virus and influenza A virus.

LiDAR-camera-system-based 3D object detection with proposal selection and grid attention pooling.

3D object detection is an important module for autonomous driving. A LiDAR camera optical system is suitable for accurate object detection, for it provides both 3D structure and 2D texture features. However, as LiDAR and a camera have different sensor properties, it is challenging to generate effective fusion features. Motivated by this, we propose, to the best of our knowledge, a novel LiDAR-camera based 3D object detection method. First, proposal selection is presented to utilize accurate 2D proposals predicted from RGB images to improve the quality of 3D proposals. It contains a (i) proposal addition and (ii) proposal filter. To increase the recall rate, the proposal addition generates extra 3D proposals via back-projecting 2D proposals on LiDAR depth. The proposal filter removes unrelated 3D proposals by matching 2D proposals with intersection-over-union thresholds. Then, considering the LiDAR mechanism, grid attention pooling is employed to estimate weights of grid points from LiDAR and image features to generate salient pooling features. Comparisons and ablation studies demonstrate that the proposed method achieves better performance and benefits the advanced application of a LiDAR camera system.

Downregulation of gap junctional intercellular communication and connexin 43 expression by bisphenol A in human granulosa cells.

Gap junctional intercellular communication (GJIC) is the transfer of ions, metabolites, and second messengers between neighboring cells through intercellular junctions. Connexin 43 (Cx43) was found to be the type of gap junction protein responsible for human granulosa cells (GCs) and oocyte communication, which is required for folliculogenesis and oocyte maturation. Bisphenol A (BPA), an estrogenic-like endocrine-disrupting chemical, is one of the most widely produced chemicals around the world. There are reports that the chemical might cause endometrial tumorigenesis and several female reproductive disorders. This study demonstrated that cell culture medium, containing antioxidants (N-acetyl-l-cysteine and l-ascorbic acid-2-phosphate), was able to enhance the survival and self-renewal of GCs. In addition, we found that BPA at environmentally relevant concentration (10-7  M) reduced Cx43 expression and GJIC in GCs through estrogen receptor and mitogen-activated protein kinase pathways. The results of this study not only reveal the reproductive toxicity of BPA but also provide possible mechanisms by which BPA inhibited GJIC in GCs.

Two-point calibration method for a zoom camera with an approximate focal-invariant radial distortion model.

A zoom camera can change its focal length and track moving objects with an adjustable resolution. To extract precise geometric information for the tracked objects, a zoom camera requires an accurate calibration method. High-precision camera calibration methods, however, usually require a number of control points that are not guaranteed in some practical situations. Most zoom cameras suffer radial distortion. Athough a traditional method can recover an undistorted image with known intrinsic parameters, it fails to work for a zoom camera with an unknown focal length. Motivated by these problems, we propose a two-point calibration method (TPCM). In this scheme, we first propose an approximate focal-invariant radial distortion (AFRD) model. With the AFRD model, an RGB image can be undistorted with an unknown focal length. After that, the TPCM method is presented to estimate the focal length and rotation matrix with only two control points for one image. Synthetic experiments demonstrate that the AFRD model is efficient. In the real data experiment, the mean reprojection error of the TPCM method is less than one pixel, which is smaller than current state-of-the-art methods, and we believe meets the demand for high-precision calibration.

Multi-level height maps-based registration method for sparse LiDAR point clouds in an urban scene.

The LiDAR sensor has been widely used for reconstruction in urban scenes. However, the current registration method makes it difficult to find stable 3D point correspondences from sparse and low overlapping LiDAR point clouds. In the urban situation, most of the LiDAR point clouds have a common flat ground. Therefore, we propose a novel, to the best of our knowledge, multi-level height (MH) maps-based coarse registration method. It requires that source and target point clouds have a common flat ground, which is easily satisfied for LiDAR point clouds in urban scenes. With MH maps, 3D registration is simplified as 2D registration, increasing the speed of registration. Robust correspondences are extracted in MH maps with different height intervals and statistic height information, improving the registration accuracy. The solid-state LiDAR Livox Mid-100 and mechanical LiDAR Velodyne HDL-64E are used in real-data and dataset experiments, respectively. Verification results demonstrate that our method is stable and outperforms state-of-the-art coarse registration methods for the sparse case. Runtime analysis shows that our method is faster than these methods, for it is non-iterative. Furthermore, our method can be extended for the unordered multi-view point clouds.

LiDAR-camera system extrinsic calibration by establishing virtual point correspondences from pseudo calibration objects.

Extrinsic calibration on LiDAR-camera system without specific calibration objects is a challenging task, for it is difficult to find point correspondences from RGB image and sparse LiDAR point cloud. In a natural scene, some objects if satisfying three conditions can be regarded as pseudo calibration objects. In this paper, we propose the virtual point correspondence at the first time. It is established from the 2D box of one pseudo calibration object in RGB image and its corresponding 3D frustum box in point cloud. Based on virtual point correspondence, we present a novel LiDAR-camera extrinsic calibration method without specific calibration objects. It requires two calibration conditions that easily satisfied in the practical application. A normal guided foreground detection method is proposed to automatically extract 3D frustum box. After that, a geometrical optimization scheme is presented to estimate the extrinsic parameters with the virtual point correspondences. Simulations and real data experiments demonstrate that our method is accurate, robust, and outperforms state-of-the-art calibration object based method.

Geometric calibration for LiDAR-camera system fusing 3D-2D and 3D-3D point correspondences.

Calibrating the extrinsic parameters on a system of 3D Light Detection And Ranging (LiDAR) and the monocular camera is a challenging task, because accurate 3D-2D or 3D-3D point correspondences are hard to establish from the sparse LiDAR point clouds in the calibration procedure. In this paper, we propose a geometric calibration method for estimating the extrinsic parameters of the LiDAR-camera system. In this method, a novel combination of planar boards with chessboard patterns and auxiliary calibration objects are proposed. The planar chessboard provides 3D-2D and 3D-3D point correspondences. Auxiliary calibration objects provide extra constraints for stable calibration results. After that, a novel geometric optimization framework is proposed to utilize these point correspondences, thus leading calibration results robust to LiDAR sensor noise. Besides, we contribute an automatic approach to extract point clouds of calibration objects. In the experiments, our method has a superior performance over state-of-the-art calibration methods. Furthermore, we verify our method by computing depth map and improvements can also be found. These results demonstrate that our method performance on the LiDAR-camera system is applicable for future advanced visual applications.

Effectiveness of modified complex decongestive physiotherapy for preventing lower extremity lymphedema after radical surgery for cervical cancer: a randomized controlled trial.

BACKGROUND: There is a lack of evidence in support of any prophylactic measure to prevent secondary lower extremity lymphedema after radical hysterectomy among patients with cervical cancer. This study aimed to determine the effectiveness of modified complex decongestive physiotherapy in reducing the risk of secondary lower extremity lymphedema after radical surgery. METHODS: A randomized single-blind clinical trial was conducted in 120 patients with cervical cancer who underwent laparoscopic radical hysterectomy with pelvic lymphadenectomy between January 2016 and April 2017 in the Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China. Participants were randomly assigned to a modified complex decongestive physiotherapy intervention group (n=60) or control group (n=60). The intervention group received a modified complex decongestive physiotherapy program, which included manual lymph drainage, compression hosiery, regular exercise, and health education. The control group only received a health education program. The main outcome was the incidence of secondary lower extremity lymphedema. RESULTS: A total of 117 patients with cervical cancer completed a 1-year follow-up. Twenty-eight (23.9%) patients developed secondary lower extremity lymphedema (20 (34.5%) patients in the control group and 8 (13.6%) in the intervention group). The incidence of secondary lower extremity lymphedema was significantly higher in the control group than in the intervention group (p=0.008; OR 0.30 (95% CI 0.12 to 0.75). The median percentage of excess volume was significantly less in the intervention group (2.1%, IQR 0.5-3.4%) than in the control group (2.96%, IQR 1.1-4.98%); (p=0.042). The mean (SD) onset time of lymphedema was 8 (2.00) months vs 4.6 (2.82) months in the intervention and control groups, respectively (p=0.004). CONCLUSIONS: This randomized trial showed that modified complex decongestive physiotherapy is effective for preventing lower extremity lymphedema in patients with cervical cancer after laparoscopic radical hysterectomy with pelvic lymphadenectomy.

Simply Mixing Poly Protein G with Detection Antibodies Enhances the Detection Limit and Sensitivity of Immunoassays.

An insufficient amount of detection antibodies bound to their antigens usually limits the sensitivity of immunoassays. Here, we describe a simple method to improve the detection limit and sensitivity of various immunoassays by mixing detection antibodies with a soluble poly protein G (named 8pG). 8pG was developed by fusing eight repeated fragment crystallizable (Fc) binding domains of streptococcal protein G to a linear polymer. Simply mixing detection antibodies with 8pG to form an antibody/8pG complex largely increased the accumulation of detection antibody to target molecules, which dramatically enhanced the sensitivity in direct ELISA, sandwich ELISA, Western blot, and flow cytometry systems, separately. The detection limit of Western blot for low-abundance PEGylated interferon (Pegasys) and recombinant human CTLA4 (rhCTLA4) improved by at least 13-fold and 31-fold, respectively, upon mixing detection antibodies with 8pG. Moreover, the nanoscale size of the antibody/8pG complex did not influence the granularity and dimension of target cells in the flow cytometry system. Collectively, we provide a quick and easy-to-operate method to make various immunoassays to sensitively detect low-abundance target molecules by just mixing their detection antibodies with 8pG.

Predictive Value of Quantitative Uterine Fibroid Perfusion Parameters From Contrast-Enhanced Ultrasound for the Therapeutic Effect of High-Intensity Focused Ultrasound Ablation.

OBJECTIVES: To assess the predictive significance of quantitative perfusion parameters from contrast-enhanced ultrasound (CEUS) for the therapeutic response to high-intensity focused ultrasound (HIFU) ablation in patients with uterine fibroids. METHODS: A total of 263 patients with single uterine fibroids were treated with HIFU ablation under ultrasound guidance. The arrival time, peak time, enhancement time, enhancement intensity, and enhancement rate were evaluated with pretreatment CEUS. According to a nonperfused volume ratio evaluation by posttreatment magnetic resonance imaging, all patients were assigned to groups with volume ratios of 70% or higher and lower than 70%. Then the predictive performances of different parameters for ablation efficacy were studied. RESULTS: The arrival time, peak time, and enhancement time in the group with a nonperfused volume ratio of 70% or higher were longer than those in the group with a volume ratio lower than 70% (mean ± SD, 16.7 ± 3.5, 26.5 ± 4.9, and 10.2 ± 2.6 seconds, respectively, versus 13.3 ± 4.2, 20.8 ± 5.4, and 7.6 ± 2.3 seconds), whereas patients with a volume ratio of 70% or higher had a lower mean enhancement intensity and enhancement rate than those with a volume ratio lower than 70% (29.7 ± 16.7 dB and 3.2 ± 1.5 dB/s versus 63.2 ± 26.3 dB and 8.6 ± 4.3 dB/s; P < .05). The nonperfused volume ratio was negatively correlated with the enhancement intensity and enhancement rate (r = -0.631 and -0.712) but positively correlated with the arrival time, peak time, and enhancement time (r = 0.322, 0.456, and 0.477; P < .05). The areas under the receiver operating characteristic curve for the enhancement time, enhancement intensity, and enhancement rate were 0.73, 0.79, and 0.81 (P < .05). CONCLUSIONS: Quantitative parameters from CEUS are potentially useful for evaluating the therapeutic effect of HIFU ablation for uterine fibroids.

Bone marrow mesenchymal stem cells repair Cr (VI)- injured kidney by regulating mitochondria-mediated apoptosis and mitophagy mediated via the MAPK signaling pathway.

The present study aimed to explore the repair effect and mechanism of bone marrow mesenchymal stem cells (BMSCs) transplantation on injured kidneys caused by hexavalent chromium (Cr (VI)). Wistar rats were intraperitoneally injected with 0.4 mg/kg•bw Cr (VI) ion solution. After 30 days, 1 × 107 BMSCs were transplanted into rats. After cell transplantation for 2 weeks, there was no significant difference in the chromium content between the model and BMSCs-therapy group by atomic absorption spectrometry. In BMSCs-therapy group, the renal organ index, the serum levels of blood urea nitrogen (BUN) and creatinine (CRE), malonaldehyde (MDA) content were significantly decreased, superoxide dismutase (SOD) activity was significantly elevated, and the pathological changes were improved compared with the model group. The results of immunohistochemical and western blot assays showed that the expressions of apoptosis-related proteins Bax, Cytochrome c, and Caspase-3, as well as autophagy-associated proteins Beclin 1, PINK1, Parkin, p-Parkin, LC3B, and the MAPK signaling pathway, including the ratio of p-p38 to p38 and p-JNK to JNK were all significantly decreased, Bcl-2 and p62 expressions, and the ratio of p-ERK to ERK were significantly elevated in BMSCs-therapy group compared with the model group. These results suggested that BMSCs repaired Cr (VI)-injured kidney through decreasing mitochondria-mediated apoptosis and mitophagy mediated by downregulating phosphorylation of p38 and JNK, upregulating phosphorylation of ERK.

Genome wide analysis of narcolepsy in China implicates novel immune loci and reveals changes in association prior to versus after the 2009 H1N1 influenza pandemic.

Previous studies in narcolepsy, an autoimmune disorder affecting hypocretin (orexin) neurons and recently associated with H1N1 influenza, have demonstrated significant associations with five loci. Using a well-characterized Chinese cohort, we refined known associations in TRA@ and P2RY11-DNMT1 and identified new associations in the TCR beta (TRB@; rs9648789 max P = 3.7 × 10(-9) OR 0.77), ZNF365 (rs10995245 max P = 1.2 × 10(-11) OR 1.23), and IL10RB-IFNAR1 loci (rs2252931 max P = 2.2 × 10(-9) OR 0.75). Variants in the Human Leukocyte Antigen (HLA)- DQ region were associated with age of onset (rs7744020 P = 7.9×10(-9) beta -1.9 years) and varied significantly among cases with onset after the 2009 H1N1 influenza pandemic compared to previous years (rs9271117 P = 7.8 × 10(-10) OR 0.57). These reflected an association of DQB1*03:01 with earlier onset and decreased DQB1*06:02 homozygosity following 2009. Our results illustrate how genetic association can change in the presence of new environmental challenges and suggest that the monitoring of genetic architecture over time may help reveal the appearance of novel triggers for autoimmune diseases.

Myosin light chain kinase (MLCK) regulates cell migration in a myosin regulatory light chain phosphorylation-independent mechanism.

Myosin light chain kinase (MLCK) has long been implicated in the myosin phosphorylation and force generation required for cell migration. Here, we surprisingly found that the deletion of MLCK resulted in fast cell migration, enhanced protrusion formation, and no alteration of myosin light chain phosphorylation. The mutant cells showed reduced membrane tether force and fewer membrane F-actin filaments. This phenotype was rescued by either kinase-dead MLCK or five-DFRXXL motif, a MLCK fragment with potent F-actin-binding activity. Pull-down and co-immunoprecipitation assays showed that the absence of MLCK led to attenuated formation of transmembrane complexes, including myosin II, integrins and fibronectin. We suggest that MLCK is not required for myosin phosphorylation in a migrating cell. A critical role of MLCK in cell migration involves regulating the cell membrane tension and protrusion necessary for migration, thereby stabilizing the membrane skeleton through F-actin-binding activity. This finding sheds light on a novel regulatory mechanism of protrusion during cell migration.

Sanguinarine inhibits Rac1b-rendered cell survival enhancement by promoting apoptosis and blocking proliferation.

AIM: Small GTPase Rac1 is a member of the Ras superfamily, which plays important roles in regulation of cytoskeleton reorganization, cell growth, proliferation, migration, etc. The aim of this study was to determine how a constitutively active Rac1b regulated cell proliferation and to investigate the effects of the Rac1b inhibitor sanguinarine. METHODS: Three HEK293T cell lines stably overexpressing GFP, Rac1-GFP or Rac1b-GFP were constructed by lentiviral infection. The cells were treated with sanguinarine (1 µmol/L) or its analogue berberine (1 µmol/L) for 4 d. Cell proliferation was evaluated by counting cell numbers and with a BrdU incorporation assay. The levels of cleaved PARP-89 (an apoptosis marker) and cyclin-D1 (a proliferative index) were measured using Western blotting. RESULTS: In 10% serum-containing media, overexpressing either Rac1 or Rac1b did not significantly change the cell proliferation. In the serum-starved media, however, the survival rate of Rac1b cells was significantly increased, whereas that of Rac1 cells was moderately increased. The level of cleaved PARP-89 was significantly increased in serum-starved Rac1 cells, but markedly reduced in serum-starved Rac1b cells. The level of cyclin-D1 was significantly increased in both serum-starved Rac1 and Rac1b cells. Treatment with sanguinarine, but not berberine, inhibited the proliferation of Rac1b cells, which was accompanied by significantly increased the level of PARP-89, and decreased both the level of cyclin-D1 and the percentage of BrdU positive cells. CONCLUSION: Rac1b enhances the cell proliferation under a growth-limiting condition via both anti-apoptotic and pro-proliferative mechanisms. Sanguinarine, as the specific inhibitor of Rac1b, is a potential therapeutic agent for malignant tumors with up-regulated Rac1b.

Upregulation of Nanog and Sox-2 genes following ectopic expression of Oct-4 in amniotic fluid mesenchymal stem cells.

Octamer-binding transcription factor 4 (Oct-4), an important gene regulating stem cell pluripotency, is well-known for its ability to reprogram somatic cells in vitro, either alone or in concert with other factors. The aim of this study was to assess the effect of ectopic expression of Oct human amniotic fluid stem cells. We developed a novel method for isolation of putative human amniotic fluid-derived multipotent stem cells. These cells showing mesenchymal stem cell phenotypes (human amniotic fluid-derived mesenchymal stem cells, hAFMSCs) were transfected with a plasmid carrying genes for Oct-4 and the green fluorescent protein (GFP). The stably transfected cells, hAFMSCs-Oct4/GFP, were selected by using G418 and found to express the GFP reporter gene under the control of Oct-4 promoter. We found that hAFMSCs developed by our method possess very high self-renewal ability (about 78 cumulative population doublings) and multilineage differentiation potency. Significantly, the hAFMSCs-Oct4/GFP cells showed enhanced expression of the three major pluripotency genes Oct-4, Nanog, and Sox-2, and increased colony-forming ability and growth rate compared with the parental hAFMSCs. We demonstrated that the ectopic expression of Oct-4 gene in hAFMSCs with high self-renewal ability could upregulate Nanog and Sox-2 gene expression and enhance cell growth rate and colony-forming efficiency. Therefore, the ectopic expression of Oct-4 could be a strategy to develop pluripotency in hAFMSCs for clinical applications.