Imaging amoeboid cancer cell motility in vivo

The availability of multi-photon intravital microscopy has recently allowed researchers to start to visualise the dynamic behaviour of cancer cells in vivo. This imaging has revealed that many cancer cells ranging from carcinoma to melanoma move in an amoeboid manner in order to invade surrounding tissue and escape from the primary tumour. This mode on cell motility is extremely rapid and does not require the activity of proteases to degrade the extra-cellular matrix (ECM). This review details the techniques that are available to study cell motility in vivo and discusses the current knowledge about the mechanisms of amoeboid cell motility.     

SPAG9 promotes prostate cancer growth and metastasis

Sperm-associated antigen 9 (SPAG9) expression is increased in prostate tissues of prostate cancerpatients. This experimental study aimed to investigate the role of SPAG9 in bone metastasis of prostate cancer.Immunohistochemical analysis showed that SPAG9 staining was positive in 81.67% of 240 cases of prostatic carcinomabut only in 6.67% of 120 cases of benign prostate hyperplasia. Strong PAG9 staining was positively correlated withGleason score and bone metastasis in 240 prostate cancer patients (p < 0.05), but not with the age or serum prostatespecific antigen level (p > 0.05). PC-3 cells were transfected with shRNA against SPAG9, and CCK-8 assay in triplicateshowed that PC-3 cell viability was inhibited by SPAG9 knockdown. In addition, transwell assay in triplicate showedthat PC-3 cell invasion was inhibited by SPAG9 knockdown. Furthermore, total 2 × 106 PC-3 cells were injectedsubcutaneously into the right flank of nude mice which were randomly divided into three groups (N = 8) and treatedby intratumoral injection of SPAG9 shRNA, control shRNA or PBS, respectively. SPAG9 shRNA inhibited thegrowth, invasion and angiogenesis while promoted apoptosis of xenografted PC-3 cells. SPAG9 knockdown led to theupregulation of E-cadherin and the downregulation of MMP2 and vimentin in xenografted tumors. In conclusion,this is the first study to provide evidence that SPAG9 promotes bone metastasis of prostate cancer, and SPAG9 is apromising target to prevent or treat bone metastasis of prostate cancer.     

Lymphatic function, lymphangiogenesis, and cancer metastasis

The lymphatic system serves as the primary route for the metastasis of many cancers and the extent of lymph node involvement is the most important indicator of tumor aggressiveness. Despite the apparent importance of the lymphatic vessels for tumor dissemination, it has remained unclear whether activation of lymphatic endothelial cells may affect tumor progression and metastasis and the molecular mechanisms of lymphangiogenesis are just beginning to be elucidated. This overview describes the unique structural and functional characteristics of the lymphatic vessels that render them particularly suitable for invasion by tumor cells and for their efficient transport to lymph nodes. Recent evidence indicates occurrence of tumor lymphangiogenesis and its correlation with metastasis. Molecular regulation of tumor lymphangiogenesis, its significance for tumor metastasis, and implications for cancer therapy are discussed.     

Use of multiphoton microscopy to diagnose liver cancer and lung metastasis in an orthotopic rat model

Liver or lung biopsy for suspicious lesions has several disadvantages such as bleeding, bile leak or pneumothorax, needle track seeding, and time-consuming histopathological procedure. The ability to directly observe cellular and subcellular details and then perform "optical biopsy" is a major goal in the development of new interventional techniques. Multiphoton microscopy (MPM) enables real-time noninvasive visualization of tissue architecture and cell morphology in live tissue. We performed a study to evaluate whether MPMcan make real-time optical diagnosis for liver cancer and lung metastasis using an orthotopic rat model with Morris hepatoma. We found that real-time high-resolution MPMimaging could clearly show tissue architecture and cell morphology. In the normal liver tissue, MPMimaging clearly revealed the blood-filled sinusoids and cords of hepatocytes. In the cancerous tissue, MPMimaging clearly illustrated that cancer cells displayed marked cellular and nuclear pleomorphism. MPMimages were comparable to golden standard hematoxylin-eosin staining images. Moreover, MPMimaging had deep penetration with the capability of optical sectioning. In short, MPMcan make real-time optical diagnosis for liver cancer and lung metastasis. This study provides the groundwork for further using multiphoton endoscopy to perform real-time noninvasive "optical biopsy" for liver cancer and lung metastasis in the near future.     

HAND2-AS1, PRKAA2 and VLDLR predict the risk of peritoneal metastasis in gastric cancer of different Lauren types based on STEPP analysis

The peritoneum is the most common site of recurrence of gastric cancer (GC). Early occult peritonealmetastasis is difficult to detect by imaging examination. Stratifying the risk of peritoneal metastasis in patients withdifferent Lauren subtypes is of great clinical value. We performed a univariate Cox regression to identify those geneswith prognostic value of overall survival (OS) and peritoneal-specified disease-free survival (psDFS) from the GeneExpression Omnibus database. The candidate genes were screened by the Subpopulation Treatment Effect PatternPlot (STEPP) method. Propensity score matching (PSM) analysis was used to reduce the interference of confounderson the results. Based on the optimal cut-off values determined by the STEPP method, we found overexpression ofthree genes (HAND2-AS1, PRKAA2, and VLDLR) was correlated with shorter 1-year psDFS among patients withdiffuse-type than that of patients with intestinal-type GC, and it is highly significant. Gene Set Enrichment Analysis(GSEA) potentially suggested that the three genes promote the early occurrence of peritoneal metastasis in patientswith diffuse-type GC through glucose metabolism-related pathways. These three genes may be potential biomarkers.They can be used to assess the risk of peritoneal metastases to guide treatment decisions and follow-up strategies.     

Video-rate confocal reflection microscopy of neoplastic cells: Rate of intracellular movement and peripheral motility characteristic of neoplastic cell line (RSK4) with high degree of growth independence in vitro

Video-rate laser confocal interference reflection microscopy was used to demonstrate rapid motion of intracellular organelles and features at the cell periphery in a fully transformed neoplastic cell line, RSK4, and in four other neoplastic cell populations. In the RSK4 cells, vibrational and trafficking movements of intracellular particles at a rate greater than 25 Hz and ranging down to 5 Hz were recorded. Rapidly moving processes changed to ruffles, then microspikes, and previously undetectable ephemeral intercellular contacts were seen. Dynamic cyclical changes were revealed in the sizes of the podosomal close contacts of the transformed cells. The visibility of such features and the temporal and spatial resolution are improved over earlier methods. The fact that fast cellular and intracellular movements can be detected with this microscopic technique offers new possibilities in attempting to recognise differences between unimpaired living cells, and it may prove useful in the identification of malignant cells.     

空间成像光谱仪主体支撑技术研究

为了改善某型号空间成像光谱仪主体支撑方式对系统光机结构稳定性的影响,从三个方面分析了系统光机结构稳定性的表达方式,介绍了常用空间光学遥感仪器的支撑方式。在传统支耳支撑方式不能满足离轴非球面三反射镜（TMA）系统热稳定性的情况下,提出了一种新的双耦合的支撑方案并对其进行了理论分析。优化设计过程中,采用有限元分析软件MSC.PATRAN对的三维样机进行工程分析,结果表明,在正弦振动载荷作用下,主体一阶频率为95Hz;在10℃均匀温降与自重耦合作用下,各反射镜面RMS最大值为12nm、PV最大值为60nm,主镜绕Z轴的转角由原来的55'降低为12'。热真空环境下整机成像试验获得的光谱数据验证工程分析的正确性,双耦合的支撑方案能满足整机刚度和光机系统的热稳定性的要求。     

Ketamine effects on the urogenital system--changes in the urinary bladder and sperm motility

Different doses of ketamine (10 mg/kg, 20 mg/kg, 30 mg/kg, 40 mg/kg, 50 mg/kg, and 60 mg/kg) were injected i.p. (I.P.), respectively, to male ICR mice to determine the optimal dosage for chronic administration. At and above 40 mg/kg I.P. injection, mice had almost no hindlimb movement during swimming test. Subsequently, 30 mg/kg was used as the dose for the study in the toxicity of long-term ketamine administration on urinary bladder and sperm motility. The treatment group were subdivided into two (n = 10 each group); one received daily ketamine treatment i.p. for 3 months and another group for 6 months. Corresponding number of mice in control groups (n = 5 each group) received saline injection instead of ketamine. Terminal dUTP nick and labeling (TUNEL) study and Sirius red staining were carried out on the sectioned slides of the urinary bladders to study the degree of apoptosis in both epithelium and muscular layers of the urinary bladder and the relative thickness of the muscular layers in this organ was also computed. Apoptosis in the bladder epithelium was observed initially in the 3-month ketamine treated mice and the number of apoptotic cells was significantly different (P < 0.05) between the 3-month and 6-month ketamine treated mice and the control. The relative thickness of muscular layers in the bladder wall also decreased significantly (P < 0.05) when the 6-month treated mice and the control were compared. Sirius red staining revealed increase of collagen in the urinary bladder of the treated mice, most evidently 6 months after ketamine treatment. In addition, the sperm motility was studied and there was a statistically significant difference between the control and ketamine treated groups in the percentages of sperms which were motile (P < 0.05). This suggested that the chronic administration of ketamine affected the genital system as well.     

Oncolytic adenovirus targeting LASP-1 inhibited renal cell cancer progression

Recent studies suggested that LIM and SH3 protein 1 (LASP-1) is a promising therapeutic target for renal cellcancer (RCC). This study aimed to explore the role of LASP-1 in RCC. For this purpose, LASP-1 expression in RCCtissues was analyzed by immunohistochemistry and Western blot analysis. Cell proliferation, migration, invasion, andgene expression were detected by CCK-8 assay, Transwell assay, and Western blot analysis. The results showed thatLASP-1 was highly expressed in RCC, and its expression level,t was positively correlated with lymph node metastasisand tumor, nodes, and metastases (TNM) stage. The knockdown of LASP-1 expression significantly inhibited theproliferation of RCC cells, increased the apoptosis rate, and inhibited RCC cell invasion and migration by inhibitingepithelial–mesenchymal transition. We conclude that LASP-1 promotes RCC progression and metastasis and is apromising therapeutic target for RCC.     

Intravital dual-colored visualization of colorectal liver metastasis in living mice using two photon laser scanning microscopy

A major challenge of cancer biology is to visualize the dynamics of the metastatic process in secondary organs at high optical resolution in vivo real-time. Here, we presented intravital, dual-colored imaging of liver metastasis formation from a single cancer cell to metastatic colonies in the living liver of living mice using two photon laser scanning microscopy (TPLSM). Red fluorescent protein expressing murine (SL4) or human (HT29) colorectal cancer cell lines were inoculated to the spleen of green fluorescent protein expressing mice. Intravital TPLSM was performed by exteriorizing and fixing the liver lobe of living mice. This was repeated several times for the long-term imaging of the same mouse. Viable cancer cells in the living liver of living mice were visualized intravitally at a magnification of over 600×. Single cancer cells were arrested within hepatic sinusoids 2 h after injection. Platelet aggregation surrounding a cancer cell was observed, indicating a phenomenon of tumor-cell induced platelet aggregation. Cancer cells were extravasated from hepatic sinusoids to the space of Disse. Protrusions of Kupffer cells surrounding a cancer cell were observed, indicating that Kupffer cells appear to phagocytose cancer cells. SL4 cells formed liver metastatic colonies with extensive stromal reaction. Liver metastases by HT29 cells were observed as a cluster of micrometastatic nodules. High-resolution, dual-colored, real-time visualization of cancer metastasis using intravital TLPSM can help to understand spatiotemporal tumor-host interactions during metastatic processes in the living organs of living animals.     

The complementary use of electron backscatter diffraction and ion channelling imaging for the characterization of nanotwins

On the example of electrodeposited nickel films, it is shown that unique information on twins with dimensions on the nanoscale can be obtained by suitable combination of ion channelling imaging and electron backscatter diffraction analysis, whereas both (routine) single techniques cannot meet the requirements for analysis of these films. High‐resolution electron backscatter diffraction is inadequate for full characterization of nanotwins, but image quality maps obtained from electron backscatter diffraction at least yield a qualitative estimation of the location and number of nanotwins. Complementing this information with ion channelling imaging provides more representative insights into the microstructure, because it supplements the quantitative investigation of the number and width of twin lamellae with additional crystallographic orientation analysis provided by EBSD. To this end, two methods for adjusting EBSD data based on ion channelling images are proposed. Thorough selection of the complementary techniques opens future perspectives for the investigation of other challenging samples with nanoscale features in the microstructure.     

Transmission electron microscopy method providing high resolution in the cell section without radiation damage

Several new features of mitochondrial nucleoid and its surroundings in mammalian cells were described previously (Pracha?, 2016). Very small details were observed using the improved transmission electron microscopy method, as described in the article. In the meantime, the method has again been improved to 2 Å resolutions in the cell section. The method described in detail in the present work is documented on the same records that were published in lower resolution in the work Pracha? (2016), enabling comparison of the achieved resolution with the previous one. New records are also presented, showing extremely high resolution and thus implying the importance of the method. Potential use of this method in different fields is suggested.     

A high-resolution and large force-range load cell by means of nonlinear cantilever beams

In the present work, we describe a nonlinear stiffening load cell with high resolution (the ability to detect 1% changes in the force) that can function over a large force range (5 orders of magnitude), and exhibit minimal hysteresis and intrinsic geometric protection from force overload. The stiffening nature of the load cell causes its deflection and strain to be very sensitive to small forces and less sensitive to large forces. High stiffness at high forces prevents the load cell from over-straining. We physically implement the nonlinear springs with cantilever beams that increasingly contact rigid surfaces with carefully chosen curvatures as more force is applied. We analytically describe the performance of the load cell as a function of its geometric and material parameters. We also describe a method for manufacturing the mechanical component of the load cell out of one monolithic part, which decreases hysteresis and assembly costs. We experimentally verify the theory for two load cells with two different sets of parameters.     

Incidence of sperm-tail tyrosine phosphorylation and hyperactivated motility in normozoospermic and asthenozoospermic human sperm samples

Our objective was to study the incidence of sperm-tail phosphotyrosine immunoreactivity in normozoospermic and asthenozoospermic human sperm samples, its association with sperm motion parameters, particularly hyperactivated motility, and its potential involvement in the pathogenesis of asthenozoospermia. The work was conducted as a prospective experimental study in the Sperm Biology and Andrology laboratories of the Jones Institute, a medical school-based fertility center. The study subjects were healthy fertile male donors (normozoospermic samples) and infertile patients (asthenozoospermic samples) attending the center. Recently ejaculated semen samples were washed twice to eliminate seminal plasma and a swim-up was performed to select the motile population which, in turn, was incubated up to 18 h at 37°C in 3.5% human serum albumin-supplemented Ham's F10 to allow for capacitation. For evaluation, sperm aliquots were taken pre-swim-up (T_o), immediately post swim-up (T₁), at 6 h (T₆), and 18 h (T₁₈) of incubation. The main outcome measures were computer-analyzed sperm motion parameters and hyperactivated motility, and immunodetection of phosphotyrosine (PY)-containing proteins. During the capacitating incubation, normozoospermic samples displayed maximum motility, velocity, and hyperactivation at T₆, significantly decreasing their values at T₁₈. PY-proteins were located both at the tail and head of spermatozoa. Their expression increased progressively during the incubation, being present in about 70% of the sperm tails at T₁₈. Asthenozoospermic samples showed an inability to respond to capacitation with an increase in motion parameters and PY-phosphorylation. At T₆, both hyperactivation and PY-phosphorylation were significantly lower than in normal samples. Our results suggest that PY-phosphorylation of tail proteins is highly conspicuous in human spermatozoa, and increases its incidence in a time-dependent manner, as more sperm become capacitated. Asthenozoospermic samples displaying low percentages of motile sperm and altered motion characteristics showed a decreased incidence of PY-phosphorelated sperm. Tail protein PY-phosphorylation may be related to sperm movement, especially to hyperactivated motility and its deficiency may be associated to asthenozoospermia.     

Micro‐CT measurements of tumoral vessels supplied by portal circulation in hepatic colorectal metastasis mouse model

The purpose of this study was to elucidate the micro CT findings of tumoral vessels supplied by portal circulation during establishment of hepatic metastasis of colorectal cancer in a mouse model. Hepatic metastases were induced in 15 BALB/c mice through the injection of murine colonic adenocarcinoma tumor cells into the mesenteric vein. Micro‐CT imaging of the tumoral vessels was obtained to clarify the microvascular architecture. We evaluated the sinusoidal structure, diameter of the tumoral vessels (DTV) and blood vessel density (BVD) according to tumor sizes ranging from 201 to 3,000 µm in diameter. A total of 116 tumors were observed on day 15 after cell injection. The mean diameter of a normal hepatic sinusoid was 11.7 ± 2.0 µm on micro CT. The DTV supplied by the portal vein of tumors measuring 1,001–1,500 µm in diameter was greater than that of tumors 200–1,000 µm in diameter. The mean BVD from the portal vein gradually decrease according to size of tumor from 201 to 3,000 µm in diameter (r² = ?0.584, P < 0.01). The characteristics of tumoral vessels supplied by portal circulation during establishment of hepatic colorectal metastases were well visualized with micro‐CT imaging. Microsc. Res. Tech. 77:415–421, 2014. © 2014 Wiley Periodicals, Inc.     

基于加权时域弯折的Lamb波自适应温度补偿方法

针对航空结构Lamb波损伤监测中急需解决的环境温度影响问题,本文提出了加权时域弯折(WTDW)的温度补偿方法,该方法无需先验参数或模型便可自适应补偿Lamb波信号中的温度影响,而且克服了传统动态时间弯折(DTW)容易抑制被补偿信号损伤信息的温度过补偿问题,从而更便于在Lamb波损伤监测中的实际应用.首先在DTW基本理论...     

Computer automated cell size and shape analysis in cryomicroscopy

Computer vision techniques have been developed for quantitative analysis of size and shape changes in cells frozen on a cryomicroscope. The analysis is based on implementation of standard serial edge detection algorithms in conjunction with a shape transform to isolate individual cells in complex scenes which may include adjacent and overlying ice crystals. In the present study the sensitivity of the automated analysis procedure is evaluated for images obtained by various microscope optical systems for progressive degrees of subject blurring by defocusing. Size measurements in calibration trials for freezing latex spheres with extracellular ice in the field of view were least sensitive for bright field images, although the most consistent data was obtained by differential interference contrast microscopy. In all cases phase contrast images produced the least accurate data. An example analysis is presented for the freezing of pancreas β-cells.     

轻小型高分辨率星载高光谱成像光谱仪

在小型化成像光谱仪的研制和应用中,如何同时实现轻量化、高地面分辨率和高信噪比是目前亟待突破的技术难题.本文通过将线性渐变滤光片分光技术和数字域时间延迟积分技术相结合,并对镜头进行紧凑化处理,设计了一款工作波段为403～988 nm、平均光谱分辨率为8.9 nm、系统总质量为7 kg的轻小型星载高光谱成像光谱仪.仿真和实...     

Cyclic nucleotide phosphodiesterase inhibition increases tyrosine phosphorylation and hyper motility in normal and pathological human spermatozoa.

Our objective was to determine the effect of phosphodiesterase (PDE) inhibition on: 1) tyrosine phosphorylation of human spermatozoa at the tail level; and 2) sperm motion parameters and hyperactivated motility. The study was conducted with normozoospermic and asthenozoospermic samples incubated under in vitro capacitating conditions. The main outcome measures were computer-assisted sperm motion analysis and fluorescent immunodetection of phosphotyrosine-containing proteins. Pentoxifylline (PTX) was used as PDE inhibitor because of its wide use in the clinic. PTX-treatment significantly increased sperm velocity, hyperactivated motility and tyrosine-phosphorylation, both in normo and asthenozoospermic samples. Tyrosine-phosphorylation of tail proteins was highly conspicuous in both types of samples, showing no differential pattern after PTX-treatment. Normozoospermic samples treated with pentoxifylline showed an increase in the number of spermatozoa displaying hyperactivated movement and tyrosine-phosphorylation at the tail level. Preliminary data on asthenozoospermic samples exhibiting altered motion characteristics and defective phosphorylation of sperm-tail proteins showed that both defects can be concomitantly overcome by pentoxifylline treatment. Tyrosine-phosphorylation of sperm-tail proteins is underlying the enhancement of hyperactivated motility resulting from PDE inhibition by pentoxifylline.