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Sample records for laser microdissected vascular

  1. Laser capture microdissection technology.

    PubMed

    Espina, Virginia; Heiby, Michael; Pierobon, Mariaelena; Liotta, Lance A

    2007-09-01

    Deciphering the cellular and molecular interactions that drive disease within the tissue microenvironment holds promise for discovering drug targets of the future. In order to recapitulate the in vivo interactions through molecular analysis, one must be able to analyze specific cell populations within the context of their heterogeneous tissue microecology. Laser capture microdissection is a method to procure subpopulations of tissue cells under direct microscopic visualization. Laser capture microdissection technology can harvest the cells of interest directly or can isolate specific cells by cutting away unwanted cells to give histologically pure enriched cell populations. A variety of downstream applications exist: DNA genotyping and loss-of-heterozygosity analysis, RNA transcript profiling, cDNA library generation, mass spectrometry proteomics discovery and signal pathway profiling.

  2. Laser-capture microdissection.

    PubMed

    Espina, Virginia; Wulfkuhle, Julia D; Calvert, Valerie S; VanMeter, Amy; Zhou, Weidong; Coukos, George; Geho, David H; Petricoin, Emanuel F; Liotta, Lance A

    2006-01-01

    Deciphering the cellular and molecular interactions that drive disease within the tissue microenvironment holds promise for discovering drug targets of the future. In order to recapitulate the in vivo interactions thorough molecular analysis, one must be able to analyze specific cell populations within the context of their heterogeneous tissue microecology. Laser-capture microdissection (LCM) is a method to procure subpopulations of tissue cells under direct microscopic visualization. LCM technology can harvest the cells of interest directly or can isolate specific cells by cutting away unwanted cells to give histologically pure enriched cell populations. A variety of downstream applications exist: DNA genotyping and loss-of-heterozygosity (LOH) analysis, RNA transcript profiling, cDNA library generation, proteomics discovery and signal-pathway profiling. Herein we provide a thorough description of LCM techniques, with an emphasis on tips and troubleshooting advice derived from LCM users. The total time required to carry out this protocol is typically 1-1.5 h.

  3. Laser capture microdissection for gene expression analysis.

    PubMed

    Bidarimath, Mallikarjun; Edwards, Andrew K; Tayade, Chandrakant

    2015-01-01

    Laser capture microdissection (LCM) is an excellent and perhaps the only platform to isolate homogeneous cell populations from specific microscopic regions of heterogeneous tissue section, under direct microscopic visualization. The basic operations of the LCM system are based on (a) microscopic visualization of phenotypically identified cells of interest, (b) selective adherence of cells to a melting thermolabile film/membrane using a low-energy infrared laser (IR system) or photovolatization of cells within a selected region (UV system), (c) capturing or catapulting of structurally intact cells from a stained tissue section. RNA/DNA or protein can be extracted from the cell or tissue fragments for downstream applications to quantitatively study gene expression. This method can be applied to many downstream analyses including but not limited to quantitative real-time polymerase chain reaction (PCR), microarray, DNA genotyping, RNA transcript profiling, generation of cDNA library, mass spectrometry analysis, and proteomic discovery.The application of LCM is described here to specifically and reliably obtain a homogeneous cell population in order to extract RNA to study microRNA expression by quantitative real-time PCR.

  4. Laser capture microdissection for gene expression analysis.

    PubMed

    Bidarimath, Mallikarjun; Edwards, Andrew K; Tayade, Chandrakant

    2015-01-01

    Laser capture microdissection (LCM) is an excellent and perhaps the only platform to isolate homogeneous cell populations from specific microscopic regions of heterogeneous tissue section, under direct microscopic visualization. The basic operations of the LCM system are based on (a) microscopic visualization of phenotypically identified cells of interest, (b) selective adherence of cells to a melting thermolabile film/membrane using a low-energy infrared laser (IR system) or photovolatization of cells within a selected region (UV system), (c) capturing or catapulting of structurally intact cells from a stained tissue section. RNA/DNA or protein can be extracted from the cell or tissue fragments for downstream applications to quantitatively study gene expression. This method can be applied to many downstream analyses including but not limited to quantitative real-time polymerase chain reaction (PCR), microarray, DNA genotyping, RNA transcript profiling, generation of cDNA library, mass spectrometry analysis, and proteomic discovery.The application of LCM is described here to specifically and reliably obtain a homogeneous cell population in order to extract RNA to study microRNA expression by quantitative real-time PCR. PMID:25308266

  5. Laser capture microdissection in the tissue biorepository.

    PubMed

    Liu, Angen

    2010-09-01

    An important need of many cancer research projects is the availability of high-quality, appropriately selected tissue. Tissue biorepositories are organized to collect, process, store, and distribute samples of tumor and normal tissue for further use in fundamental and translational cancer research. This, in turn, provides investigators with an invaluable resource of appropriately examined and characterized tissue specimens and linked patient information. Human tissues, in particular, tumor tissues, are complex structures composed of heterogeneous mixtures of morphologically and functionally distinct cell types. It is essential to analyze specific cell types to identify and define accurately the biologically important processes in pathologic lesions. Laser capture microdissection (LCM) is state-of-the-art technology that provides the scientific community with a rapid and reliable method to isolate a homogeneous population of cells from heterogeneous tissue specimens, thus providing investigators with the ability to analyze DNA, RNA, and protein accurately from pure populations of cells. This is particularly well-suited for tumor cell isolation, which can be captured from complex tissue samples. The combination of LCM and a tissue biorepository offers a comprehensive means by which researchers can use valuable human biospecimens and cutting-edge technology to facilitate basic, translational, and clinical research. This review provides an overview of LCM technology with an emphasis on the applications of LCM in the setting of a tissue biorepository, based on the author's extensive experience in LCM procedures acquired at Fox Chase Cancer Center and Hollings Cancer Center.

  6. Live cell isolation by laser microdissection with gravity transfer

    NASA Astrophysics Data System (ADS)

    Podgorny, Oleg V.

    2013-05-01

    Laser microdissection by pulsing ultraviolet laser allows the isolation and recultivation of live cells based on morphological features or/and fluorescent labelling from adherent cell cultures. Previous investigations described only the use of the laser microdissection and pressure catapulting (LMPC) for live cell isolation. But LMPC requires complex manipulations and some skill. Furthermore, single-cell cloning using laser microdissection has not yet been demonstrated. The first evidence of successful application of laser microdissection with gravity transfer (LMDGT) for capturing and recultivation of live cells is presented. A new strategy for LMDGT is presented because of the failure to reproduce the manufacturer's protocol. Using the new strategy, successful capturing and recultivation of circle-shaped samples from confluent monolayer of HeLa cells was demonstrated. It was found that LMDGT is easier than LMPC because it doesn't require personal participation of investigator in transferring of isolated samples to final culture dishes. Moreover, for the first time, the generation of clonal colonies from single live cells isolated by laser microdissection was demonstrated. Data obtained in this study confirm that LMDGT is a reliable and high-yield method allowing isolation and expansion of both cell clusters and single cells from adherent cell cultures.

  7. Isolation of single Chlamydia-infected cells using laser microdissection.

    PubMed

    Podgorny, Oleg V; Polina, Nadezhda F; Babenko, Vladislav V; Karpova, Irina Y; Kostryukova, Elena S; Govorun, Vadim M; Lazarev, Vassili N

    2015-02-01

    Chlamydia are obligate intracellular parasites of humans and animals that cause a wide range of acute and chronic infections. To elucidate the genetic basis of chlamydial parasitism, several approaches for making genetic modifications to Chlamydia have recently been reported. However, the lack of the available methods for the fast and effective selection of genetically modified bacteria restricts the application of genetic tools. We suggest the use of laser microdissection to isolate of single live Chlamydia-infected cells for the re-cultivation and whole-genome sequencing of single inclusion-derived Chlamydia. To visualise individual infected cells, we made use of the vital labelling of inclusions with the fluorescent Golgi-specific dye BODIPY® FL C5-ceramide. We demonstrated that single Chlamydia-infected cells isolated by laser microdissection and placed onto a host cell monolayer resulted in new cycles of infection. We also demonstrated the successful use of whole-genome sequencing to study the genomic variability of Chlamydia derived from a single inclusion. Our work provides the first evidence of the successful use of laser microdissection for the isolation of single live Chlamydia-infected cells, thus demonstrating that this method can help overcome the barriers to the fast and effective selection of Chlamydia.

  8. [Laser microdissection in the molecular oncology of prostate cancer].

    PubMed

    Wernert, N

    2004-06-01

    Nearly all diseases, including prostate cancer (PCA), occur in mixed tissues with different cell types interconnected by multiple interactions. Laser microdissection permits a separate analysis of specific cell types necessary to understand tumorigenesis. Microdissection can be combined with different molecular methods for analyses at the levels of the genome, the transcriptome or the proteome. With respect to the molecular pathogenesis of PCA, normal glands can be compared to preneoplasias, and these in turn to the carcinoma. Different malignancy grades, as well as intra- and extraprostatic tumor parts, can be specifically analysed and molecular markers of aggressiveness can be identified. The molecular signatures obtained provide the basis for functional studies. New prognostic markers and therapeutic targets can be expected from such approaches in the near future. A far reaching goal is the computer representation of multiple molecular components and their interactions, "E-cell in cyberspace", in which prognostic behaviour and therapeutic responsiveness can be approximately predicted. PMID:15098090

  9. Laser Capture Microdissection as a Tool to Study Tumor Stroma.

    PubMed

    Bertos, Nicholas R; Park, Morag

    2016-01-01

    Laser capture microdissection (or LCM) allows for isolation of cells from specific tissue compartments, which can then be followed by DNA, RNA, and/or protein isolation and downstream characterization. Unlike other methods for cell isolation, LCM can be directed towards cells situated in specific anatomical contexts, and is therefore of significant value when investigating the tumor microenvironment, where localization is often key to function. Here, we present a summary of ways in which LCM can be utilized, as well as protocols for the isolation of tumor and tumor-associated stromal elements from frozen breast cancer samples, with a focus on preparation of samples for RNA characterization. PMID:27581011

  10. Laser capture microdissection of gonads from juvenile zebrafish

    PubMed Central

    2009-01-01

    Background Investigating gonadal gene expression is important in attempting to elucidate the molecular mechanism of sex determination and differentiation in the model species zebrafish. However, the small size of juvenile zebrafish and correspondingly their gonads complicates this type of investigation. Furthermore, the lack of a genetic sex marker in juvenile zebrafish prevents pooling gonads from several individuals. The aim of this study was to establish a method to isolate the gonads from individual juvenile zebrafish allowing future investigations of gonadal gene expression during sex determination and differentiation. Methods The laser capture microdissection technique enables isolation of specific cells and tissues and thereby removes the noise of gene expression from other cells or tissues in the gene expression profile. A protocol developed for laser microdissection of human gonocytes was adjusted and optimised to isolate juvenile zebrafish gonads. Results The juvenile zebrafish gonad is not morphologically distinguishable when using dehydrated cryosections on membrane slides and a specific staining method is necessary to identify the gonads. The protocol setup in this study allows staining, identification, isolation and subsequent RNA purification and amplification of gonads from individual juvenile zebrafish thereby enabling gonadal gene expression profiling. Conclusion The study presents a protocol for isolation of individual juvenile zebrafish gonads, which will enable future investigations of gonadal gene expression during the critical period of sex differentiation. Furthermore, the presented staining method is applicable to other species as it is directed towards alkaline phosphatase that is expressed in gonocytes and embryonic stem cells, which is conserved among vertebrate species. PMID:19747405

  11. Advances in isolation and characterization of homogeneous cell populations using laser microdissection.

    PubMed

    Mizuarai, S; Takahashi, K; Kobayashi, T; Kotani, H

    2005-01-01

    The isolation and characterization of homogeneous cell populations are of great importance for the analysis of gene expression, because normal tissues contain various types of cells, and the differences in the populations of isolated cells exert significant effects on gene expression analysis. Researchers have attempted to develop methods for the isolation of homogeneous cell populations, such as flow cytometry and mechanical dissection. However, the recent emergence of laser-assisted microdissection has revolutionized the isolation of single-cell populations from solid tissues. With the help of a cutting laser, laser microdissection can isolate tissues (cells) of interest without contamination from surrounding tissues with the microscopic visualization field. By combining laser microdissection and subsequent microarray technology, several studies have resulted in the identification of disease-related genes. In this review, we summarize the principle of laser microdissection and provide several successful examples of target-gene identification using the conventional method combining laser microdissection and microarray. Next, we discuss the practical drawbacks of the combinational method, such as the need for a large number of cells and the disturbance of the relative abundance of transcripts during RNA amplification. We introduce our modifications to combined laser microdissection and microarray for detection of disease-related genes; the technique is simple, yet practical and accurate. Finally, versatile applications of laser microdissection, not only to transcript expression analysis, but also to other genomics and proteomics analyses are, also presented.

  12. Analysis of neuronal gene expression with laser capture microdissection.

    PubMed

    Vincent, Valerie A M; DeVoss, Jason J; Ryan, Heather S; Murphy, Greer M

    2002-09-01

    The brain is a heterogeneous tissue in which the numbers of neurons, glia, and other cell types vary among anatomic regions. Gene expression studies performed on brain homogenates yield results reflecting mRNA abundance in a mixture of cell types. Therefore, a method for quantifying gene expression in individual cell populations would be useful. Laser capture microdissection (LCM) is a new technique for obtaining pure populations of cells from heterogeneous tissues. Most studies thus far have used LCM to detect DNA sequences. We developed a method to quantify gene expression in hippocampal neurons from mouse brain using LCM and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). This method was optimized to permit histochemical or immunocytochemical visualization of nerve cells during LCM while minimizing RNA degradation. As an example, gene expression was quantified in hippocampal neurons from the Tg2576 mouse model for Alzheimer's disease.

  13. Automatic detection of spermatozoa for laser capture microdissection.

    PubMed

    Vandewoestyne, Mado; Van Hoofstat, David; Van Nieuwerburgh, Filip; Deforce, Dieter

    2009-03-01

    In sexual assault crimes, differential extraction of spermatozoa from vaginal swab smears is often ineffective, especially when only a few spermatozoa are present in an overwhelming amount of epithelial cells. Laser capture microdissection (LCM) enables the precise separation of spermatozoa and epithelial cells. However, standard sperm-staining techniques are non-specific and rely on sperm morphology for identification. Moreover, manual screening of the microscope slides is time-consuming and labor-intensive. Here, we describe an automated screening method to detect spermatozoa stained with Sperm HY-LITER. Different ratios of spermatozoa and epithelial cells were used to assess the automatic detection method. In addition, real postcoital samples were also screened. Detected spermatozoa were isolated using LCM and DNA analysis was performed. Robust DNA profiles without allelic dropout could be obtained from as little as 30 spermatozoa recovered from postcoital samples, showing that the staining had no significant influence on DNA recovery.

  14. Targeting pancreatic islets with phage display assisted by laser pressure catapult microdissection.

    PubMed

    Yao, Virginia J; Ozawa, Michael G; Trepel, Martin; Arap, Wadih; McDonald, Donald M; Pasqualini, Renata

    2005-02-01

    Heterogeneity of the microvasculature in different organs has been well documented by multiple methods including in vivo phage display. However, less is known about the diversity of blood vessels within functionally distinct regions of organs. Here, we combined in vivo phage display with laser pressure catapult microdissection to identify peptide ligands for vascular receptors in the islets of Langerhans in the murine pancreas. Protein database analyses of the peptides, CVSNPRWKC and CHVLWSTRC, showed sequence identity to two ephrin A-type ligand homologues, A2 and A4. Confocal microscopy confirmed that most immunoreactivity of CVSNPRWKC and CHVLWSTRC phage was associated with blood vessels in pancreatic islets. Antibodies recognizing EphA4, a receptor for ephrin-A ligands, were similarly associated with islet blood vessels. Importantly, binding of both islet-homing phage and anti-EphA4 antibody was strikingly increased in blood vessels of pancreatic islet tumors in RIP-Tag2 transgenic mice. These results indicate that endothelial cells of blood vessels in pancreatic islets preferentially express EphA4 receptors, and this expression is increased in tumors. Our findings show in vivo phage display and laser pressure catapult microdissection can be combined to reveal endothelial cell specialization within focal regions of the microvasculature.

  15. Laser microdissection-based analysis of plant sex chromosomes.

    PubMed

    Hobza, Roman; Vyskot, Boris

    2007-01-01

    A recent progress in plant molecular biology has led to enormous available data of DNA sequences, including complete nuclear genomes of Arabidopsis, rice, and poplar. On the other hand, in plant species with more complex genomes, containing widespread repetitive sequences, it is important to establish genomic resources that help us to focus on particular part of genomes. Laser technology enables to handle with specific subcellular structures or even individual chromosomes. Here we present a comprehensive protocol to isolate and characterize DNA sequences derived from the sex chromosomes of white campion (Silene latifolia). This dioecious plant has become the most favorite model to study the structure, function, and evolution of plant sex chromosomes due to a large and distinguishable size of both the X and Y chromosomes. The protocol includes a versatile technique to prepare metaphase chromosomes from either germinating seeds or in vitro cultured hairy roots. Such slides can be used for laser chromosome microdissection, fluorescence in situ-hybridization mapping, and immunostaining. Here we also demonstrate some applications of the laser-dissected chromosome template, especially a modified FAST-FISH technique to paint individual chromosomes, and construction and screening of chromosome-specific DNA libraries.

  16. Epigenetic Analysis of Laser Capture Microdissected Fetal Epithelia1

    PubMed Central

    Seelan, Ratnam S.; Warner, Dennis R.; Mukhopadhyay, Partha M.; Andres, Sarah A.; Smolenkova, Irina A.; Wittliff, James L.; Pisano, M. Michele; Greene, Robert M.

    2013-01-01

    Laser capture microdissection (LCM) is a superior method for non-destructive collection of specific cell populations from tissue sections. While DNA, RNA and protein have been analyzed from LCM-procured samples, epigenetic analyses, particularly of fetal, highly hydrated tissue, have not been attempted. A standardized protocol with quality assurance measures was established to procure cells by LCM of the medial edge epithelia (MEE) of the fetal palatal processes for isolation of intact microRNA for expression analyses and genomic DNA for CpG methylation analyses. MicroRNA preparations, obtained using the RNAqueous® Micro kit (Life Technologies), exhibited better yields and higher quality than those obtained using the Arcturus® PicoPure® RNA Isolation kit (Life Technologies). The approach was validated using real-time PCR to determine expression of selected microRNAs (miR-99a and miR-200b) and pyrosequencing to determine CpG methylation status of selected genes (Aph1a and Dkk4) in the MEE. These studies describe an optimized approach for employing LCM of epithelial cells from fresh frozen fetal tissue that enables quantitative analyses of miRNA expression levels and CpG methylation. PMID:23911529

  17. Laser capture microdissection: Big data from small samples

    PubMed Central

    Datta, Soma; Malhotra, Lavina; Dickerson, Ryan; Chaffee, Scott; Sen, Chandan K.; Roy, Sashwati

    2015-01-01

    Any tissue is made up of a heterogeneous mix of spatially distributed cell types. In response to any (patho) physiological cue, responses of each cell type in any given tissue may be unique and cannot be homogenized across cell-types and spatial co-ordinates. For example, in response to myocardial infarction, on one hand myocytes and fibroblasts of the heart tissue respond differently. On the other hand, myocytes in the infarct core respond differently compared to those in the peri-infarct zone. Therefore, isolation of pure targeted cells is an important and essential step for the molecular analysis of cells involved say in the progression of disease. Laser capture microdissection (LCM) is powerful to obtain a pure targeted cell subgroup, or even a single cell, quickly and precisely under the microscope, successfully tackling the problem of tissue heterogeneity in molecular analysis. This review presents an overview of LCM technology, the principles, advantages and limitations and its down-stream applications in the fields of proteomics, genomics and transcriptomics. With powerful technologies and appropriate applications, this technique provides unprecedented insights into cell biology from cells grown in their natural tissue habitat as opposed to those cultured in artificial petri dish conditions. PMID:25892148

  18. [Fixation of cells for analysis by laser microdissection--comparative studies in forensic trace material].

    PubMed

    Fischer, Elisabeth J; Laberke, Patrick J; Kübler, Eric; Balitzki, Beate

    2012-01-01

    This paper is focused on the preparation of samples for laser microdissection (LM) in forensic casework. In forensic genetics, it is essential to preserve and separate cellular traces during sample preparation, as they are usually gathered in very small amounts and are often contaminated with undesired cells. This is made possible by laser microdissection, a technique developed to cut cells or tissue of a certain type from a microscopical specimen by UV laser and catapult them directly into a PCR reactor. This method minimizes the risk of getting inconclusive, mixed DNA profiles due to contamination by foreign DNA and also supplies information about the cellular origin of a DNA profile. A method for optimized fixation and staining of spermatozoa for laser microdissection was established. Four different fixation methods combined with two staining methods were tested on two different microscope slides. Moreover, the effect of a blocker pen to contain the specimen on the slide was investigated.

  19. Laser capture microdissection for protein and NanoString RNA analysis.

    PubMed

    Golubeva, Yelena; Salcedo, Rosalba; Mueller, Claudius; Liotta, Lance A; Espina, Virginia

    2013-01-01

    Laser capture microdissection (LCM) allows the precise procurement of enriched cell populations from a heterogeneous tissue, or live cell culture, under direct microscopic visualization. Histologically enriched cell populations can be procured by harvesting cells of interest directly or isolating specific cells by ablating unwanted cells. The basic components of laser microdissection technology are (a) visualization of cells via light microscopy, (b) transfer of laser energy to a thermolabile polymer with either the formation of a polymer-cell composite (capture method) or transfer of laser energy via an ultraviolet laser to photovolatize a region of tissue (cutting method), and (c) removal of cells of interest from the heterogeneous tissue section. The capture and cutting methods (instruments) for laser microdissection differ in the manner by which cells of interest are removed from the heterogeneous sample. Laser energy in the capture method is infrared (810 nm), while in the cutting mode the laser is ultraviolet (355 nm). Infrared lasers melt a thermolabile polymer that adheres to the cells of interest, whereas ultraviolet lasers ablate cells for either removal of unwanted cells or excision of a defined area of cells. LCM technology is applicable to an array of applications including mass spectrometry, DNA genotyping and loss-of-heterozygosity analysis, RNA transcript profiling, cDNA library generation, proteomics discovery, and signal kinase pathway profiling. This chapter describes LCM using an Arcturus(XT) instrument for downstream protein sample analysis and using an mmi CellCut Plus® instrument for RNA analysis via NanoString technology.

  20. Optimizing Frozen Sample Preparation for Laser Microdissection: Assessment of CryoJane Tape-Transfer System®.

    PubMed

    Golubeva, Yelena G; Smith, Roberta M; Sternberg, Lawrence R

    2013-01-01

    Laser microdissection is an invaluable tool in medical research that facilitates collecting specific cell populations for molecular analysis. Diversity of research targets (e.g., cancerous and precancerous lesions in clinical and animal research, cell pellets, rodent embryos, etc.) and varied scientific objectives, however, present challenges toward establishing standard laser microdissection protocols. Sample preparation is crucial for quality RNA, DNA and protein retrieval, where it often determines the feasibility of a laser microdissection project. The majority of microdissection studies in clinical and animal model research are conducted on frozen tissues containing native nucleic acids, unmodified by fixation. However, the variable morphological quality of frozen sections from tissues containing fat, collagen or delicate cell structures can limit or prevent successful harvest of the desired cell population via laser dissection. The CryoJane Tape-Transfer System®, a commercial device that improves cryosectioning outcomes on glass slides has been reported superior for slide preparation and isolation of high quality osteocyte RNA (frozen bone) during laser dissection. Considering the reported advantages of CryoJane for laser dissection on glass slides, we asked whether the system could also work with the plastic membrane slides used by UV laser based microdissection instruments, as these are better suited for collection of larger target areas. In an attempt to optimize laser microdissection slide preparation for tissues of different RNA stability and cryosectioning difficulty, we evaluated the CryoJane system for use with both glass (laser capture microdissection) and membrane (laser cutting microdissection) slides. We have established a sample preparation protocol for glass and membrane slides including manual coating of membrane slides with CryoJane solutions, cryosectioning, slide staining and dissection procedure, lysis and RNA extraction that facilitated

  1. Laser capture microdissection: Arcturus(XT) infrared capture and UV cutting methods.

    PubMed

    Gallagher, Rosa I; Blakely, Steven R; Liotta, Lance A; Espina, Virginia

    2012-01-01

    Laser capture microdissection (LCM) is a technique that allows the precise procurement of enriched cell populations from a heterogeneous tissue under direct microscopic visualization. LCM can be used to harvest the cells of interest directly or can be used to isolate specific cells by ablating the unwanted cells, resulting in histologically enriched cell populations. The fundamental components of laser microdissection technology are (a) visualization of the cells of interest via microscopy, (b) transfer of laser energy to a thermolabile polymer with either the formation of a polymer-cell composite (capture method) or transfer of laser energy via an ultraviolet laser to photovolatize a region of tissue (cutting method), and (c) removal of cells of interest from the heterogeneous tissue section. Laser energy supplied by LCM instruments can be infrared (810 nm) or ultraviolet (355 nm). Infrared lasers melt thermolabile polymers for cell capture, whereas ultraviolet lasers ablate cells for either removal of unwanted cells or excision of a defined area of cells. LCM technology is applicable to an array of applications including mass spectrometry, DNA genotyping and loss-of-heterozygosity analysis, RNA transcript profiling, cDNA library generation, proteomics discovery, and signal kinase pathway profiling. This chapter describes the unique features of the Arcturus(XT) laser capture microdissection instrument, which incorporates both infrared capture and ultraviolet cutting technology in one instrument, using a proteomic downstream assay as a model.

  2. ADAM12 and ADAM17 gene expression in laser-capture microdissected and non-microdissected breast tumors.

    PubMed

    Narita, Diana; Seclaman, Edward; Ilina, Razvan; Cireap, Natalia; Ursoniu, Sorin; Anghel, Andrei

    2011-06-01

    ADAM (a disintegrin and metalloprotease)12 and ADAM17 are multidomain transmembrane proteins involved in ectodomain shedding of cytokines, growth factors and adhesion molecules, with pivotal activities in the tumor microenvironment. The aim of this study was to confirm the up-regulation of ADAM17 and ADAM12 gene splicing variants in breast tumors and to delineate their expression between laser-capture microdissected (LCM) and non-microdissected breast tumors. The gene expression was analyzed by quantitative-reverse transcription-PCR in a total sample of 109 breast tumors paired with corresponding non-neoplastic breast tissues. ADAM12 and 17 proteins expression for corresponding tissue samples was confirmed by immunohistochemistry. ADAM12S, 12L and 17 genes were significantly up-regulated in either malign or benign LCM samples when compared to non-tumor controls. For non-LCM samples, it was obtained also an increased expression for ADAM12 and 17 genes in cancers, while in benign tumors only ADAM12 variants were significantly up-regulated compared to controls. When benign versus malignant tumors were compared, in LCM samples all investigated genes displayed a higher expression in cancers, whereas in non-LCM, ADAM12 variants were overexpressed in benign samples. The increased expression of ADAM12 protein in the tumor cells and stroma of benign breast diseases was immunohistochemically confirmed. These differences between LCM and non-LCM samples were explained by the contribution of the stroma to the expression of this marker. This study underlines the accuracy conferred by homogenous LCM samples on gene expression profiles and confers further evidence regarding the role of ADAM12 and 17 in the breast tumorigenesis and progression.

  3. Laser Capture Microdissection for Protein and NanoString RNA analysis

    PubMed Central

    Golubeva, Yelena; Salcedo, Rosalba; Mueller, Claudius; Liotta, Lance A.; Espina, Virginia

    2013-01-01

    Laser capture microdissection (LCM) allows the precise procurement of enriched cell populations from a heterogeneous tissue, or live cell culture, under direct microscopic visualization. Histologically enriched cell populations can be procured by harvesting cells of interest directly, or isolating specific cells by ablating unwanted cells. The basic components of laser microdissection technology are a) visualization of cells via light microscopy, b) transfer of laser energy to a thermolabile polymer with either the formation of a polymer-cell composite (capture method) or transfer of laser energy via an ultraviolet laser to photovolatize a region of tissue (cutting method), and c) removal of cells of interest from the heterogeneous tissue section. The capture and cutting methods (instruments) for laser microdissection differ in the manner by which cells of interest are removed from the heterogeneous sample. Laser energy in the capture method is infrared (810nm), while in the cutting mode the laser is ultraviolet (355nm). Infrared lasers melt a thermolabile polymer that adheres to the cells of interest, whereas ultraviolet lasers ablate cells for either removal of unwanted cells or excision of a defined area of cells. LCM technology is applicable to an array of applications including mass spectrometry, DNA genotyping and loss-of-heterozygosity analysis, RNA transcript profiling, cDNA library generation, proteomics discovery, and signal kinase pathway profiling. This chapter describes laser capture microdissection using an ArcturusXT instrument for protein LCM sample analysis, and using a mmi CellCut Plus® instrument for RNA analysis via NanoString technology. PMID:23027006

  4. Beyond laser microdissection technology: follow the yellow brick road for cancer research

    PubMed Central

    Legres, Luc G; Janin, Anne; Masselon, Christophe; Bertheau, Philippe

    2014-01-01

    Normal biological tissues harbour different populations of cells with intricate spacial distribution patterns resulting in heterogeneity of their overall cellular composition. Laser microdissection involving direct viewing and expertise by a pathologist, enables access to defined cell populations or specific region on any type of tissue sample, thus selecting near-pure populations of targeted cells. It opens the way for molecular methods directed towards well-defined populations, and provides also a powerful tool in studies focused on a limited number of cells. Laser microdissection has wide applications in oncology (diagnosis and research), cellular and molecular biology, biochemistry and forensics for tissue selection, but other areas have been gradually opened up to these new methodological approaches, such as cell cultures and cytogenetics. In clinical oncology trials, molecular profiling of microdissected samples can yield global “omics” information which, together, with the morphological analysis of cells, can provide the basis for diagnosis, prognosis and patient-tailored treatments. This remarkable technology has brought new insights in the understanding of DNA, RNA, and the biological functions and regulation of proteins to identify molecular disease signatures. We review herein the different applications of laser microdissection in a variety of fields, and we particularly focus attention on the pre-analytical steps that are crucial to successfully perform molecular-level investigations. PMID:24482735

  5. Research Techniques Made Simple: Laser Capture Microdissection in Cutaneous Research.

    PubMed

    Chen Gonzalez, Estela; McGee, Jean Suh

    2016-10-01

    In cutaneous research, we aim to study the molecular signature of a diseased tissue. However, such a study is met with obstacles due to the inherent heterogeneous nature of tissues because multiple cell types reside within a tissue. Furthermore, there is cellular communication between the tissue and the neighboring extracellular matrix. Laser capture microdissection is a powerful technique that allows researchers to isolate cells of interest from any tissue using a laser source under microscopic visualization, thereby circumventing the issue of tissue heterogeneity. Target cells from fixed preparations can be extracted and examined without disturbing the tissue structure. In live cultures, a subpopulation of cells can be extracted in real time with minimal disturbance of cellular communication and molecular signatures. Here we describe the basic principles of the technique, the different types of laser capture microdissection, and the subsequent downstream analyses. This article will also discuss how the technique has been employed in cutaneous research, as well as future directions. PMID:27664715

  6. mRNA expression profiling of laser microbeam microdissected cells from slender embryonic structures.

    PubMed

    Scheidl, Stefan J; Nilsson, Sven; Kalén, Mattias; Hellström, Mats; Takemoto, Minoru; Håkansson, Joakim; Lindahl, Per

    2002-03-01

    Microarray hybridization has rapidly evolved as an important tool for genomic studies and studies of gene regulation at the transcriptome level. Expression profiles from homogenous samples such as yeast and mammalian cell cultures are currently extending our understanding of biology, whereas analyses of multicellular organisms are more difficult because of tissue complexity. The combination of laser microdissection, RNA amplification, and microarray hybridization has the potential to provide expression profiles from selected populations of cells in vivo. In this article, we present and evaluate an experimental procedure for global gene expression analysis of slender embryonic structures using laser microbeam microdissection and laser pressure catapulting. As a proof of principle, expression profiles from 1000 cells in the mouse embryonic (E9.5) dorsal aorta were generated and compared with profiles for captured mesenchymal cells located one cell diameter further away from the aortic lumen. A number of genes were overexpressed in the aorta, including 11 previously known markers for blood vessels. Among the blood vessel markers were endoglin, tie-2, PDGFB, and integrin-beta1, that are important regulators of blood vessel formation. This demonstrates that microarray analysis of laser microbeam micro-dissected cells is sufficiently sensitive for identifying genes with regulative functions.

  7. Amplification of multiple genomic loci from single cells isolated by laser micro-dissection of tissues

    PubMed Central

    Frumkin, Dan; Wasserstrom, Adam; Itzkovitz, Shalev; Harmelin, Alon; Rechavi, Gideon; Shapiro, Ehud

    2008-01-01

    Background Whole genome amplification (WGA) and laser assisted micro-dissection represent two recently developed technologies that can greatly advance biological and medical research. WGA allows the analysis of multiple genomic loci from a single genome and has been performed on single cells from cell suspensions and from enzymatically-digested tissues. Laser micro-dissection makes it possible to isolate specific single cells from heterogeneous tissues. Results Here we applied for the first time WGA on laser micro-dissected single cells from stained tissue sections, and developed a protocol for sequentially performing the two procedures. The combined procedure allows correlating the cell's genome with its natural morphology and precise anatomical position. From each cell we amplified 122 genomic and mitochondrial loci. In cells obtained from fresh tissue sections, 64.5% of alleles successfully amplified to ~700000 copies each, and mitochondrial DNA was amplified successfully in all cells. Multiplex PCR amplification and analysis of cells from pre-stored sections yielded significantly poorer results. Sequencing and capillary electrophoresis of WGA products allowed detection of slippage mutations in microsatellites (MS), and point mutations in P53. Conclusion Comprehensive genomic analysis of single cells from stained tissue sections opens new research opportunities for cell lineage and depth analyses, genome-wide mutation surveys, and other single cell assays. PMID:18284708

  8. Gene-expression analysis of single cells-nested polymerase chain reaction after laser microdissection

    PubMed Central

    Shi, Xin; Kleeff, JÖrg; Zhu, Zhao-Wen; Schmied, Bruno; Tang, Wen-Hao; Zimmermann, Arthur; BÜchler, Markus W.; Friess, Helmut

    2003-01-01

    AIM: The structural and functional characteristics of cells are dependent on the specific gene expression profile. The ability to study and compare gene expression at the cellular level will therefore provide valuable insights into cell physiology and pathophysiology. METHODS: Individual cells were isolated from frozen colon tissue sections using laser microdissection. DNA as well as RNA were extracted, and total RNA was reversely transcribed to complementary DNA (cDNA). Both DNA and cDNA were analyzed by nested polymerase chain reaction (PCR). The quality of isolated DNA and RNA was satisfactory. RESULTS: Single cells were successfully microdissected using an ultraviolet laser micromanipulator. Nested PCR amplification products of DNA and cDNA of single cells could clearly be visualized by agarose gel electrophoresis. CONCLUSION: The combined use of laser microdissection and nested-PCR provides an opportunity to analyze gene expression in single cells. This method allows the analysis and identification of specific genes which are involved in physiological and pathophysiological processes in a complex of variable cell phenotypes. PMID:12800252

  9. Laser microdissection and its application to analyze gene expression in arbuscular mycorrhizal symbiosis.

    PubMed

    Gomez, S Karen; Harrison, Maria J

    2009-05-01

    Phosphorus is essential for plant growth, and in many soils phosphorus availability limits crop production. Most plants in natural ecosystems obtain phosphorus via a symbiotic partnership with arbuscular mycorrhizal (AM) fungi. While the significance of these associations is apparent, their molecular basis is poorly understood. Consequently, the potential to harness the mycorrhizal symbiosis to improve phosphorus nutrition in agriculture is not realized. Transcript profiling has recently been used to investigate gene expression changes that accompany development of the AM symbiosis. While these approaches have enabled the identification of AM-symbiosis-associated genes, they have generally involved the use of RNA from whole mycorrhizal roots. Laser microdissection techniques allow the dissection and capture of individual cells from a tissue. RNA can then be isolated from these samples and cell-type specific gene expression information can be obtained. This technology has been applied to obtain cells from plants and more recently to study plant-microbe interactions. The latter techniques, particularly those developed for root-microbe interactions, are of relevance to plant-parasitic weed research. Here, laser microdissection, its use in plant biology and in particular plant-microbe interactions are discussed. An overview of the AM symbiosis is then provided, with a focus on recent advances in understanding development of the arbuscule-cortical cell interface. Finally, the recent applications of laser microdissection for analyses of AM symbiosis are discussed.

  10. Characterization of Glomerular Diseases Using Proteomic Analysis of Laser Capture Microdissected Glomeruli

    PubMed Central

    Satoskar, AA; Shapiro, JP; Bott, C; Song, H; Nadasdy, GM; Brodsky, SV; Hebert, L; Birmingham, DJ; Nadasdy, T; Freitas, M; Rovin, BH

    2012-01-01

    The application of molecular techniques to characterize clinical kidney biopsies has the potential to provide insights into glomerular diseases that cannot be revealed by traditional renal pathology. The present work is a proof-of-concept approach to test whether proteomic analysis of glomeruli isolated from clinical biopsies by laser capture microdissection can provide unique information regarding differentially-expressed proteins relevant to disease pathogenesis. The proteomes of glomeruli isolated by laser capture microdissection from biopsies of normal kidneys (living-related donor kidneys) were compared to those from patients with diabetic nephropathy, lupus nephritis, and fibronectin glomerulopathy. Glomerular proteins were extracted, trypsin digested and subjected to liquid chromatography-tandem mass spectrometry for identification and quantitation. Relative to normal glomeruli, all disease-associated glomeruli showed an increased presence of complement components, a marked decline in podocyte-associated proteins, and a decrease in proteins associated with cellular metabolism. Additionally, fibronectin glomerulopathy glomeruli differed from all the other glomeruli because of a significant accumulation of fibronectin and fibulin. This study demonstrates that our method acquires reproducible and quantitative proteomic information from laser capture microdissection isolates that can be used to characterize the molecular features of glomerular diseases. PMID:22282304

  11. Laser capture microdissection for analysis of macrophage gene expression from atherosclerotic lesions.

    PubMed

    Feig, Jonathan E; Fisher, Edward A

    2013-01-01

    Coronary artery disease, resulting from atherosclerosis, is the leading cause of death in the Western world. Most previous studies have subjected atherosclerotic arteries, a tissue of mixed cellular composition, to homogenization in order to identify the factors in plaque development, thereby obscuring information relevant to specific cell types. Because macrophage foam cells are critical mediators in atherosclerotic plaque advancement, we reasoned that performing gene analysis on those cells would provide specific insight in novel regulatory factors and potential therapeutic targets. We demonstrated for the first time in vascular biology that foam cell-specific RNA can be isolated by laser capture microdissection (LCM) of plaques. As expected, compared to whole tissue, a significant enrichment in foam cell-specific RNA transcripts was observed. Furthermore, because regression of atherosclerosis is a tantalizing clinical goal, we developed and reported a transplantation-based mouse model. This involved allowing plaques to form in apoE-/- mice and then changing the plaque's plasma environment from hyperlipidemia to normolipidemia. Under those conditions, rapid regression ensued in a process involving emigration of plaque foam cells to regional and systemic lymph nodes. Using LCM, we were able to show that under regression conditions, there was decreased expression in foam cells of inflammatory genes, but an up-regulation of cholesterol efflux genes. Interestingly, we also found that increased expression of chemokine receptor CCR7, a known factor in dendritic cell migration, was required for regression. In conclusion, the LCM methods described in this chapter, which have already lead to a number of striking findings, will likely further facilitate the study of cell type-specific gene expression in animal and human plaques during various stages of atherosclerosis, and after genetic, pharmacologic, and environmental perturbations.

  12. Laser microdissection of small tissue samples--application to chronic pancreatitis tissues.

    PubMed

    Heinmöller, Ernst; Bockholt, Anke; Werther, Meike; Ziemer, Maria; Müller, Annegret; Ghadimi, B Michael; Rüschoff, Josef

    2003-01-01

    Laser microdissection is considered to be the gold standard of tissue sampling, especially if a defined small tissue area consisting of single or few cells within a heterogeneous tissue compartment is of interest. This sophisticated technique offers the opportunity of rapid and contamination-free tissue sampling for RNA- or DNA-based molecular genetic studies. We have applied laser microdissection to a molecular genetic study of pancreatic intraductal lesions (PanINs) in tissues of chronic pancreatitis, where an exact microdissection of small ducts within a dense fibrous tissue is of paramount importance for following analysis. From nine patients suffering from chronic pancreatitis, formalin-fixed, paraffin-embedded tissue specimens were laser microdissected, and a total of 202 normal ducts and PanINs of grade PanIN-1A to grade PanIN-2 were harvested. After whole genome amplification by improved primer extension and preamplification PCR (I-PEP-PCR), microsatellite-PCR based loss of heterozygosity analysis (LOH) of the tumor suppressor gene loci TP53, p16INK4, and DPC4 was performed. One of 85 informative duct lesions (1.2%) had LOH of TP53, 1 of 76 duct lesions (1.3%) had LOH of DPC4, and 2/29 duct lesions (6.9%) showed LOH of p16INK4. Microsatellite instability (MSI) was seen in 2 of 178 duct lesions (1.1%). Immunohistochemical staining of p53 protein and DPC4 protein revealed no aberrant expression. These preliminary data indicate that LOH of tumor suppressor genes, important in pancreatic cancer genesis or MSI, can be found in chronic pancreatitis tissues, but their incidence is low. PMID:12924436

  13. Principles of laser microdissection and catapulting of histologic specimens and live cells.

    PubMed

    Vogel, Alfred; Horneffer, Verena; Lorenz, Kathrin; Linz, Norbert; Hüttmann, Gereon; Gebert, Andreas

    2007-01-01

    Rapid contact- and contamination-free procurement of specific samples of histologic material for proteomic and genomic analysis as well as separation and transport of living cells can be achieved by laser microdissection (LMD) of the sample of interest followed by a laser-induced forward transport process [laser pressure "catapulting," (LPC)] of the dissected material. We investigated the dynamics of LMD and LPC with focused and defocused laser pulses by means of time-resolved photography. The working mechanism of microdissection was found to be plasma-mediated ablation. Catapulting is driven by plasma formation, when tightly focused pulses are used, and by ablation at the bottom of the sample for moderate and strong defocusing. Driving pressures of several hundred megapascals accelerate the specimen to initial velocities of 100-300 m/s before it is rapidly slowed down by air friction. With strong defocusing, driving pressure and initial flight velocity decrease considerably. On the basis of a characterization of the thermal and optical properties of the histologic specimens and supporting materials used, we calculated the temporal evolution of the heat distribution in the sample. After laser microdissection and laser pressure catapulting (LMPC), the samples were inspected by scanning electron microscopy. Catapulting with tightly focused or strongly defocused pulses results in very little collateral damage, while slight defocusing involves significant heat and UV exposure of up to about 10% of the specimen volume, especially if samples are catapulted directly from a glass slide. Time-resolved photography of live-cell catapulting revealed that in defocused catapulting strong shear forces originate from the flow of the thin layer of culture medium covering the cells. By contrast, pulses focused at the periphery of the specimen cause a fast rotational movement that makes the specimen wind its way out of the culture medium, thereby undergoing much less shear stresses

  14. Laser capture microdissection to identify septum-associated proteins in Aspergillus nidulans.

    PubMed

    Zhang, Ying; Fischer, Reinhard; Teichert, Ines; Kück, Ulrich

    2016-01-01

    To spatially resolve genetic differences at the cellular level, the laser-capture microdissection technique was developed. With this method cells can be cut from tissues with a laser beam and analyzed for DNA, RNA or protein composition. Here we adapted the technique to isolate septal microtubule-organizing center (MTOC)-associated proteins in Aspergillus nidulans About 3000 septa were collected and subjected to peptide fingerprinting by mass-spectrometric analysis. We identified the microtubule polymerase AlpA and found it interacts with ApsB specifically at sMTOCs, suggesting that AlpA might be involved in the assembly or the functioning of this protein complex. PMID:26951366

  15. Laser Microdissection and Atmospheric Pressure Chemical Ionization Mass Spectrometry Coupled for Multimodal Imaging

    SciTech Connect

    Lorenz, Matthias; Ovchinnikova, Olga S; Kertesz, Vilmos; Van Berkel, Gary J

    2013-01-01

    This paper describes the coupling of ambient laser ablation surface sampling, accomplished using a laser capture microdissection system, with atmospheric pressure chemical ionization mass spectrometry for high spatial resolution multimodal imaging. A commercial laser capture microdissection system was placed in close proximity to a modified ion source of a mass spectrometer designed to allow for sampling of laser ablated material via a transfer tube directly into the ionization region. Rhodamine 6G dye of red sharpie ink in a laser etched pattern as well as cholesterol and phosphatidylcholine in a cerebellum mouse brain thin tissue section were identified and imaged from full scan mass spectra. A minimal spot diameter of 8 m was achieved using the 10X microscope cutting objective with a lateral oversampling pixel resolution of about 3.7 m. Distinguishing between features approximately 13 m apart in a cerebellum mouse brain thin tissue section was demonstrated in a multimodal fashion including co-registered optical and mass spectral chemical images.

  16. Laser Capture Microdissection of Archival Kidney Tissue for qRT-PCR.

    PubMed

    Hewitson, Tim D; Christie, Michael; Smith, Edward R

    2016-01-01

    Whole-organ molecular analysis of the kidney potentially misses important factors involved in the pathogenesis of disease in glomeruli and tubules. Organ wide analysis can however be augmented by using laser capture microdissection (LCM) to isolate morphologically similar cells and nephron structures from a heterogeneous tissue section via direct visualization of the cells. The protocol here provides a practical approach utilizing LCM in combination with RNA isolation techniques for downstream analysis. This technique is readily applicable to study mRNA expression in isolated glomeruli and tubules in both experimental animal models and human kidney biopsy material.

  17. Optimised laser microdissection of the human ocular surface epithelial regions for microarray studies

    PubMed Central

    2013-01-01

    Background The most important challenge of performing insitu transcriptional profiling of the human ocular surface epithelial regions is obtaining samples in sufficient amounts, without contamination from adjacent tissue, as the region of interest is microscopic and closely apposed to other tissues regions. We have effectively collected ocular surface (OS) epithelial tissue samples from the Limbal Epithelial Crypt (LEC), limbus, cornea and conjunctiva of post-mortem cadaver eyes with laser microdissection (LMD) technique for gene expression studies with spotted oligonucleotide microarrays and Gene 1.0 ST arrays. Methods Human donor eyes (4 pairs for spotted oligonucleotide microarrays, 3 pairs for Gene 1.0 ST arrays) consented for research were included in this study with due ethical approval of the Nottingham Research Ethics Committee. Eye retrieval was performed within 36 hours of post-mortem period. The dissected corneoscleral buttons were immersed in OCT media and frozen in liquid nitrogen and stored at −80°C till further use. Microscopic tissue sections of interest were taken on PALM slides and stained with Toluidine Blue for laser microdissection with PALM microbeam systems. Optimisation of the laser microdissection technique was crucial for efficient and cost effective sample collection. Results The starting concentration of RNA as stipulated by the protocol of microarray platforms was taken as the cut-off concentration of RNA samples in our studies. The area of LMD tissue processed for spotted oligonucleotide microarray study ranged from 86,253 μm2 in LEC to 392,887 μm2 in LEC stroma. The RNA concentration of the LMD samples ranged from 22 to 92 pg/μl. The recommended starting concentration of the RNA samples used for Gene 1.0 ST arrays was 6 ng/5 μl. To achieve the desired RNA concentration the area of ocular surface epithelial tissue sample processed for the Gene 1.0 ST array experiments was approximately 100,0000 μm2 to 130,0000 μm2. RNA

  18. Tissue-specific laser microdissection of the Brassica napus funiculus improves gene discovery and spatial identification of biological processes.

    PubMed

    Chan, Ainsley C; Khan, Deirdre; Girard, Ian J; Becker, Michael G; Millar, Jenna L; Sytnik, David; Belmonte, Mark F

    2016-05-01

    The three primary tissue systems of the funiculus each undergo unique developmental programs to support the growth and development of the filial seed. To understand the underlying transcriptional mechanisms that orchestrate development of the funiculus at the globular embryonic stage of seed development, we used laser microdissection coupled with RNA-sequencing to produce a high-resolution dataset of the mRNAs present in the epidermis, cortex, and vasculature of the Brassica napus (canola) funiculus. We identified 7761 additional genes in these tissues compared with the whole funiculus organ alone using this technology. Differential expression and enrichment analyses were used to identify several biological processes associated with each tissue system. Our data show that cell wall modification and lipid metabolism are prominent in the epidermis, cell growth and modification occur in the cortex, and vascular tissue proliferation and differentiation occur in the central vascular strand. We provide further evidence that each of the three tissue systems of the globular stage funiculus are involved in specific biological processes that all co-ordinate to support seed development. The identification of genes and gene regulators responsible for tissue-specific developmental processes of the canola funiculus now serves as a valuable resource for seed improvement research. PMID:27194740

  19. Tissue-specific laser microdissection of the Brassica napus funiculus improves gene discovery and spatial identification of biological processes

    PubMed Central

    Chan, Ainsley C.; Khan, Deirdre; Girard, Ian J.; Becker, Michael G.; Millar, Jenna L.; Sytnik, David; Belmonte, Mark F.

    2016-01-01

    The three primary tissue systems of the funiculus each undergo unique developmental programs to support the growth and development of the filial seed. To understand the underlying transcriptional mechanisms that orchestrate development of the funiculus at the globular embryonic stage of seed development, we used laser microdissection coupled with RNA-sequencing to produce a high-resolution dataset of the mRNAs present in the epidermis, cortex, and vasculature of the Brassica napus (canola) funiculus. We identified 7761 additional genes in these tissues compared with the whole funiculus organ alone using this technology. Differential expression and enrichment analyses were used to identify several biological processes associated with each tissue system. Our data show that cell wall modification and lipid metabolism are prominent in the epidermis, cell growth and modification occur in the cortex, and vascular tissue proliferation and differentiation occur in the central vascular strand. We provide further evidence that each of the three tissue systems of the globular stage funiculus are involved in specific biological processes that all co-ordinate to support seed development. The identification of genes and gene regulators responsible for tissue-specific developmental processes of the canola funiculus now serves as a valuable resource for seed improvement research. PMID:27194740

  20. Targeting hepatocytes from liver tissue by laser capture microdissection and proteomics expression profiling.

    PubMed

    Marko-Varga, György; Berglund, Magnus; Malmström, Johan; Lindberg, Henrik; Fehniger, Thomas E

    2003-11-01

    A tissue proteomics process is presented where hepatocyte cell isolation in combination with two-dimensional (2-D) gel electrophoresis and mass spectrometric identification were used to annotate the liver proteome. Laser microdissection of 8 microm liver tissue sections was performed and protein expression profiling was compared using a variety of quantities of input cells, and gel separation conditions. The 30 microm diameter laser generated the highest protein yields from the polymer coated caps following microsolubilization. We found that 6000 laser pulses (approximately 7200 hepatocytes) were required in order to generate high-resolution gel maps. Within homogeneous tissue samples, this could be accomplished in a total cycle time of 20 min using an automated dissection procedure. Close to 1000 high-quality gel annotations were generated from the corresponding 2-D gel expression profiles which matched closely the corresponding patterns of analytical-scale liver preparations detected by silver staining.

  1. Improved Protocol For Laser Microdissection Of Human Pancreatic Islets From Surgical Specimens

    PubMed Central

    Sturm, Dorothée; Marselli, Lorella; Ehehalt, Florian; Richter, Daniela; Distler, Marius; Kersting, Stephan; Grützmann, Robert; Bokvist, Krister; Froguel, Philippe; Liechti, Robin; Jörns, Anne; Meda, Paolo; Baretton, Gustavo Bruno; Saeger, Hans-Detlev; Schulte, Anke M.; Marchetti, Piero; Solimena, Michele

    2013-01-01

    Laser microdissection (LMD) is a technique that allows the recovery of selected cells and tissues from minute amounts of parenchyma 1,2. The dissected cells can be used for a variety of investigations, such as transcriptomic or proteomic studies, DNA assessment or chromosomal analysis 2,3. An especially challenging application of LMD is transcriptome analysis, which, due to the lability of RNA 4, can be particularly prominent when cells are dissected from tissues that are rich of RNases, such as the pancreas. A microdissection protocol that enables fast identification and collection of target cells is essential in this setting in order to shorten the tissue handling time and, consequently, to ensure RNA preservation. Here we describe a protocol for acquiring human pancreatic beta cells from surgical specimens to be used for transcriptomic studies 5. Small pieces of pancreas of about 0.5-1 cm3 were cut from the healthy appearing margins of resected pancreas specimens, embedded in Tissue-Tek O.C.T. Compound, immediately frozen in chilled 2-Methylbutane, and stored at -80 °C until sectioning. Forty serial sections of 10 μm thickness were cut on a cryostat under a -20 °C setting, transferred individually to glass slides, dried inside the cryostat for 1-2 min, and stored at -80 °C. Immediately before the laser microdissection procedure, sections were fixed in ice cold, freshly prepared 70% ethanol for 30 sec, washed by 5-6 dips in ice cold DEPC-treated water, and dehydrated by two one-minute incubations in ice cold 100% ethanol followed by xylene (which is used for tissue dehydration) for 4 min; tissue sections were then air-dried afterwards for 3-5 min. Importantly, all steps, except the incubation in xylene, were performed using ice-cold reagents - a modification over a previously described protocol 6. utilization of ice cold reagents resulted in a pronounced increase of the intrinsic autofluorescence of beta cells, and facilitated their recognition. For

  2. Cell-Type-Specific Genome-wide Expression Profiling after Laser Capture Microdissection of Living Tissue

    SciTech Connect

    Marchetti, F; Manohar, C F

    2005-02-09

    The purpose of this technical feasibility study was to develop and evaluate robust microgenomic tools for investigations of genome-wide expression of very small numbers of cells isolated from whole tissue sections. Tissues contain large numbers of cell-types that play varied roles in organ function and responses to endogenous and exogenous toxicants whether bacterial, viral, chemical or radiation. Expression studies of whole tissue biopsy are severely limited because heterogeneous cell-types result in an averaging of molecular signals masking subtle but important changes in gene expression in any one cell type(s) or group of cells. Accurate gene expression analysis requires the study of specific cell types in their tissue environment but without contamination from surrounding cells. Laser capture microdissection (LCM) is a new technology to isolate morphologically distinct cells from tissue sections. Alternative methods are available for isolating single cells but not yet for their reliable genome-wide expression analyses. The tasks of this feasibility project were to: (1) Develop efficient protocols for laser capture microdissection of cells from tissues identified by antibody label, or morphological stain. (2) Develop reproducible gene-transcript analyses techniques for single cell-types and determine the numbers of cells needed for reliable genome-wide analyses. (3) Validate the technology for epithelial and endothelial cells isolated from the gastrointestinal tract of mice.

  3. Laser Microdissection of Narrow Sheath Mutant Maize Uncovers Novel Gene Expression in the Shoot Apical Meristem

    PubMed Central

    Zhang, Xiaolan; Madi, Shahinez; Borsuk, Lisa; Nettleton, Dan; Elshire, Robert J; Buckner, Brent; Janick-Buckner, Diane; Beck, Jon; Timmermans, Marja; Schnable, Patrick S; Scanlon, Michael J

    2007-01-01

    Microarrays enable comparative analyses of gene expression on a genomic scale, however these experiments frequently identify an abundance of differentially expressed genes such that it may be difficult to identify discrete functional networks that are hidden within large microarray datasets. Microarray analyses in which mutant organisms are compared to nonmutant siblings can be especially problematic when the gene of interest is expressed in relatively few cells. Here, we describe the use of laser microdissection microarray to perform transcriptional profiling of the maize shoot apical meristem (SAM), a ~100-μm pillar of organogenic cells that is required for leaf initiation. Microarray analyses compared differential gene expression within the SAM and incipient leaf primordium of nonmutant and narrow sheath mutant plants, which harbored mutations in the duplicate genes narrow sheath1 (ns1) and narrow sheath2 (ns2). Expressed in eight to ten cells within the SAM, ns1 and ns2 encode paralogous WUSCHEL1-like homeobox (WOX) transcription factors required for recruitment of leaf initials that give rise to a large lateral domain within maize leaves. The data illustrate the utility of laser microdissection-microarray analyses to identify a relatively small number of genes that are differentially expressed within the SAM. Moreover, these analyses reveal potentially conserved WOX gene functions and implicate specific hormonal and signaling pathways during early events in maize leaf development. PMID:17571927

  4. Using laser micro-dissection and qRT-PCR to analyze cell type-specific gene expression in Norway spruce phloem.

    PubMed

    Nagy, Nina E; Sikora, Katarzyna; Krokene, Paal; Hietala, Ari M; Solheim, Halvor; Fossdal, Carl Gunnar

    2014-01-01

    The tangentially oriented polyphenolic parenchyma (PP) and radially organized ray parenchyma in the phloem are central in the defense of conifer stems against insects and pathogens. Laser micro-dissection enables examination of cell-specific defense responses. To examine induced defense responses in Norway spruce stems inoculated with the necrotrophic blue-stain fungus Ceratocystis polonica, RNA extracted from laser micro-dissected phloem parenchyma and vascular cambium was analyzed using real-time RT-PCR (qRT-PCR) to profile transcript levels of selected resistance marker genes. The monitored transcripts included three pathogenesis-related proteins (class IV chitinase (CHI4), defensin (SPI1), peroxidase (PX3), two terpene synthesis related proteins (DXPS and LAS), one ethylene biosynthesis related protein (ACS), and a phenylalanine ammonia-lyase (PAL). Three days following inoculation, four genes (CHI4, PAL, PX3, SPI1) were differentially induced in individual cell and tissue types, both close to the inoculation site (5 mm above) and, to a lesser degree, further away (10 mm above). These resistance marker genes were all highly induced in ray parenchyma, supporting the important role of the rays in spruce defense propagation. CHI4 and PAL were also induced in PP cells and in conducting secondary phloem tissues. Our data suggests that different cell types in the secondary phloem of Norway spruce have overlapping but not fully redundant roles in active host defense. Furthermore, the study demonstrates the usefulness of laser micro-dissection coupled with qRT-PCR to characterize gene expression in different cell types of conifer bark.

  5. Immunoguided Laser Assisted Microdissection Techniques for DNA Methylation Analysis of Archival Tissue Specimens

    PubMed Central

    Eberle, Franziska C.; Hanson, Jeffrey C.; Killian, J. Keith; Wei, Lai; Ylaya, Kris; Hewitt, Stephen M.; Jaffe, Elaine S.; Emmert-Buck, Michael R.; Rodriguez-Canales, Jaime

    2010-01-01

    Altered DNA methylation is a fundamental characteristic of carcinogenesis. The analysis of DNA methylation in tumor cells may help to better understand tumor pathogenesis and more importantly may be used as diagnostic tool with therapeutic consequences. To detect targets relevant in tumorigenesis, it is essential to separate neoplastic cells from nonneoplastic cells. An excellent method for isolating specific cells is laser-assisted microdissection (LAM). Target cell identification for immunoguided LAM (ILAM) requires immunohistochemistry (IHC). Yet, it is unclear whether IHC for ILAM influences DNA methylation. The goals of this study were to establish an optimized protocol for antigen retrieval and IHC of formalin-fixed paraffin-embedded (FFPE) specimens suitable for ILAM and to evaluate its effect on the DNA methylome using a high throughput array. Using ten archival FFPE specimens, we showed specific staining suitable for ILAM. Extracted DNA from microdissected cells of immunohistochemically or H&E-stained tissue sections showed identical DNA quality and a strong correlation (r = 0.94 to 0.98) for CpG target methylation of 1505 analyzed sites in a series of five paired samples. No differential methylation between H&E and IHC was detected in 1501 of 1505 CpG targets (99.7%; P < 0.05). These results demonstrate the validity and utility of the herein described protocol, which allows the application of ILAM for large-scale genomic and epigenetic analyses of archival tissue specimens. PMID:20413681

  6. Laser microdissection-based expression analysis of key genes involved in muscle regeneration in mdx mice.

    PubMed

    Marotta, Mario; Sarria, Yaris; Ruiz-Roig, Claudia; Munell, Francina; Roig-Quilis, Manuel

    2007-10-01

    We have used the mdx mice strain (C57BL/10ScSn-mdx) as an experimental subject for the study of reiterative skeletal muscle necrosis-regeneration with basement membrane preservation. In young mdx muscle, by means of Hematoxylin-Eosin staining, different types of degenerative-regenerative groups (DRG) can be recognized and assigned to a defined muscle regeneration phase. To evaluate the expression of known key-regulatory genes in muscle regeneration, we have applied Laser Capture Microdissection technique to obtain tissue from different DRGs encompassing the complete skeletal muscle regenerative process. The expression of MyoD, Myf-5 and Myogenin showed a rapid increase in the first two days post-necrosis, which were followed by MRF4 expression, when newly regenerating fibers started to appear (3-5days post-necrosis). MHCd mRNA levels, undetectable in mature non-injured fibers, increased progressively from the first day post-necrosis and reached its maximum level of expression in DRGs showing basophilic regenerating fibers. TGFbeta-1 mRNA expression showed a prompt and strong increase following fiber necrosis that persisted during the inflammatory phase, and progressively decreased when new regenerating fibers began to appear. In contrast, IGF-2 mRNA expression decreased during the first days post-necrosis but was followed by a progressive rise in its expression coinciding with the appearance of the newly formed myofibers, reaching the maximum expression levels in DRGs composed of medium caliber basophilic regenerating myofibers (5-7 days post-necrosis). mdx degenerative-regenerative group typing, in conjunction with laser microdissection-based gene expression analysis, opens up a new approach to the molecular study of skeletal muscle regeneration.

  7. Contact-free isolation of sperm and epithelial cells by laser microdissection and pressure catapulting.

    PubMed

    Seidl, Stephan; Burgemeister, Renate; Hausmann, Roland; Betz, Peter; Lederer, Thomas

    2005-06-01

    With the PALM MicroBeam system, precise laser microdissection of single cells from cell smears or tissue preparations is possible. Furthermore, this system uses a contact-free and therefore contamination-free laser pressure catapulting technique in which high energy generated by a focused laser pulse catapults single dissected cells into a collecting vessel. In this study, this technique was tested for forensic purposes with smear preparations from postcoital vaginal swabs, sperm swabs, and buccal cell swabs on different types of microscopic slides. Apart from super-frosted slides, cutting and catapulting of selected cells was possible in all cases. Subsequent polymerase chain reaction was performed using the genRES MPX-2 Amplification Kit. In the case of sperm cells stained with hematoxylin and eosin, fragments larger than approximately 200 bp could not be detected. Partial genetic profiles were obtained for DNA amounts originating from only two cell equivalents. Complete profiles, however, were observed with all preparations of a minimum of 10 epithelial cells, demonstrating a potential benefit of this technique for the contamination-free forensic analysis of extremely small specimens or mixed stains. PMID:25869953

  8. Contact-free isolation of sperm and epithelial cells by laser microdissection and pressure catapulting.

    PubMed

    Seidl, Stephan; Burgemeister, Renate; Hausmann, Roland; Betz, Peter; Lederer, Thomas

    2005-06-01

    With the PALM MicroBeam system, precise laser microdissection of single cells from cell smears or tissue preparations is possible. Furthermore, this system uses a contact-free and therefore contamination-free laser pressure catapulting technique in which high energy generated by a focused laser pulse catapults single dissected cells into a collecting vessel. In this study, this technique was tested for forensic purposes with smear preparations from postcoital vaginal swabs, sperm swabs, and buccal cell swabs on different types of microscopic slides. Apart from super-frosted slides, cutting and catapulting of selected cells was possible in all cases. Subsequent polymerase chain reaction was performed using the genRES MPX-2 Amplification Kit. In the case of sperm cells stained with hematoxylin and eosin, fragments larger than approximately 200 bp could not be detected. Partial genetic profiles were obtained for DNA amounts originating from only two cell equivalents. Complete profiles, however, were observed with all preparations of a minimum of 10 epithelial cells, demonstrating a potential benefit of this technique for the contamination-free forensic analysis of extremely small specimens or mixed stains.

  9. Subtissue-Specific Evaluation of Promoter Efficiency by Quantitative Fluorometric Assay in Laser Microdissected Tissues of Rapeseed[W

    PubMed Central

    Jasik, Jan; Schiebold, Silke; Rolletschek, Hardy; Denolf, Peter; Van Adenhove, Katrien; Altmann, Thomas; Borisjuk, Ljudmilla

    2011-01-01

    β-Glucuronidase (GUS) is a useful reporter for the evaluation of promoter characteristics in transgenic plants. Here, we introduce an original technique to quantify the strength of promoters at subtissue resolution of cell clusters. The method combines cryotomy, laser microdissection, and improved fluorometric analysis of GUS activity using 6-chloro-4-methylumbelliferyl-β-d-glucuronide as an efficient fluorogenic substrate for kinetic studies in plants. The laser microdissection/6-chloro-4-methylumbelliferyl-β-d-glucuronide method is robust and reliable in a wide range of GUS expression levels and requires extremely low (few cells) tissue amounts. Suitability of the assay was demonstrated on rapeseed (Brassica napus) plants transformed with a P35S2::GUS construct. GUS expression patterns were visualized and quantified in approximately 30 tissues of vegetative and generative organs. Considerable differences in promoter activity within the tissues are discussed in relation to the cell type and developmental state. PMID:21825109

  10. Quantitative Proteomics in Laser Capture Microdissected Sleep Nuclei From Rat Brain

    PubMed Central

    Miller, Ronald A.; Winrow, Christopher J.; Spellman, Daniel S.; Song, Qinghua; Reiss, Duane R.; Conway, James P.; Taylor, Rhonda R.; Coleman, Paul J.; Hendrickson, Ronald C.

    2014-01-01

    The combination of stable isotope labeling of amino acids in mammals (SILAM) and laser capture microdissection (LCM) for selective proteomic analysis of the targeted tissues holds tremendous potential for refined characterization of proteome changes within complex tissues such as the brain. The authors have applied this approach to measure changes in relative protein abundance in ventral tegmental area (VTA) of the rat brain that correlate to pharmacological perturbations. Enriched 13C6 15N2-lysine was introduced in vivo via diet. These animals were sacrificed during the middle of the 12-hour light period to extract isotopically “heavy” proteins, which were then used as a reference for extracts from dosed, unlabeled rats. Animals were administered an orexin peptide (Ox-B), an orexin receptor antagonist (ORA), or a mixture of both (Ox-B + ORA). All samples were obtained at same phase of the sleep cycle. Labeled-pair identification and differential quantitation provided protein identification and expression ratio data. Five proteins were found to exhibit decreased relative abundance after administration of an ORA, including α-synuclein and rat myelin basic protein. Conversely, six proteins showed increased relative abundance upon antagonist treatment, including 2’,3’-cyclic nucleotide 3’-phosphodiesterase. PMID:24579665

  11. Gene expression profiling of reproductive meristem types in early rice inflorescences by laser microdissection.

    PubMed

    Harrop, Thomas W R; Ud Din, Israr; Gregis, Veronica; Osnato, Michela; Jouannic, Stefan; Adam, Hélène; Kater, Martin M

    2016-04-01

    In rice, inflorescence architecture is established at early stages of reproductive development and contributes directly to grain yield potential. After induction of flowering, the complexity of branching, and therefore the number of seeds on the panicle, is determined by the activity of different meristem types and the timing of transitions between them. Although some of the genes involved in these transitions have been identified, an understanding of the network of transcriptional regulators controlling this process is lacking. To address this we used a precise laser microdissection and RNA-sequencing approach in Oryza sativa ssp. japonica cv. Nipponbare to produce quantitative data that describe the landscape of gene expression in four different meristem types: the rachis meristem, the primary branch meristem, the elongating primary branch meristem (including axillary meristems), and the spikelet meristem. A switch in expression profile between apical and axillary meristem types followed by more gradual changes during transitions in axillary meristem identity was observed, and several genes potentially involved in branching were identified. This resource will be vital for a mechanistic understanding of the link between inflorescence development and grain yield. PMID:26932536

  12. Selective capture of endothelial and perivascular cells from brain microvessels using laser capture microdissection.

    PubMed

    Kinnecom, Katie; Pachter, Joel S

    2005-12-01

    Laser capture microdissection (LCM) of the major cell types comprising brain microvessels offers a powerful technology to explore the molecular basis of the blood-brain barrier in health and disease. However, the ability to selectively retrieve endothelial or perivascular cells, without cross-contamination from the other, has proven difficult. Additionally, histochemical methods previously described for use with LCM have not allowed for identification of all the different size branches of the microvascular tree. Here, we describe a double immunostaining method, combining bright-field and fluorescence microscopy, and using an extensive dehydration with xylene, to clearly identify and spatially resolve endothelial from perivascular cells within all size microvascular branches in frozen brain sections. LCM of these sections, coupled with RNA analysis by reverse-transcription polymerase chain reaction, revealed that captured endothelial cells show endothelial markers but no detectable markers for astrocytes or smooth muscle cells/pericytes. Conversely, captured astrocytes or smooth muscle cells/pericytes demonstrate their respective markers, but not those of endothelial cells. This approach has applicability to microarray analysis, thereby enabling global gene profiling of the different cell types along the entirety of the brain microvascular tree.

  13. Application of laser microdissection to identify the mycorrhizal fungi that establish arbuscules inside root cells.

    PubMed

    Berruti, Andrea; Borriello, Roberto; Lumini, Erica; Scariot, Valentina; Bianciotto, Valeria; Balestrini, Raffaella

    2013-01-01

    Obligate symbiotic fungi that form arbuscular mycorrhizae (AMF; belonging to the Glomeromycota phylum) are some of the most important soil microorganisms. AMFs facilitate mineral nutrient uptake from the soil, in exchange for plant-assimilated carbon, and promote water-stress tolerance and resistance to certain diseases. AMFs colonize the root by producing inter- and intra-cellular hyphae. When the fungus penetrates the inner cortical cells, it produces a complex ramified structure called arbuscule, which is considered the preferential site for nutrient exchange. Direct DNA extraction from the whole root and sequencing of ribosomal gene regions are commonly carried out to investigate intraradical AMF communities. Nevertheless, this protocol cannot discriminate between the AMFs that actively produce arbuscules and those that do not. To solve this issue, the authors have characterized the AMF community of arbusculated cells (AC) through a laser microdissection (LMD) approach, combined with sequencing-based taxa identification. The results were then compared with the AMF community that was found from whole root DNA extraction. The AMF communities originating from the LMD samples and the whole root samples differed remarkably. Five taxa were involved in the production of arbuscules, while two taxa were retrieved inside the root but not in the AC. Unexpectedly, one taxon was found in the AC, but its detection was not possible when extracting from the whole root. Thus, the LMD technique can be considered a powerful tool to obtain more precise knowledge on the symbiotically active intraradical AMF community.

  14. Laser Capture Microdissection of Feline Streptomyces spp Pyogranulomatous Dermatitis and Cellulitis.

    PubMed

    Traslavina, R P; Reilly, C M; Vasireddy, R; Samitz, E M; Stepnik, C T; Outerbridge, C; Affolter, V K; Byrne, B A; Lowenstine, L J; White, S D; Murphy, B

    2015-11-01

    Suspected Streptomyces spp infections were identified in 4 cats at UC Davis Veterinary Medical Teaching Hospital between 1982 and 2011. Three had ulcerated, dark red mycetomas involving the dermis, subcutis, and fascia with fistulous tracts and/or regional lymphadenopathy. One cat had pyogranulomatous mesenteric lymphadenitis. Granulomatous inflammation in all cats contained colonies of Gram-positive, non-acid-fast organisms. All 4 cats failed to respond to aggressive medical and surgical treatment and were euthanized. Laser capture microdissection (LCM) was used to selectively harvest DNA from the affected formalin-fixed, paraffin-embedded (FFPE) tissues. Cloned amplicons from LCM-derived tissue confirmed the presence of Streptomyces spp in the dermatitis cases. Amplicons from the remaining cat with peritoneal involvement aligned with the 16S ribosomal RNA gene for Actinomycetales. Usually considered a contaminant, Streptomyces spp can be associated with refractory pyogranulomatous dermatitis and cellulitis in cats with outdoor access. LCM is useful in the diagnosis of bacterial diseases where contamination may be an issue.

  15. Gene expression profiling of reproductive meristem types in early rice inflorescences by laser microdissection.

    PubMed

    Harrop, Thomas W R; Ud Din, Israr; Gregis, Veronica; Osnato, Michela; Jouannic, Stefan; Adam, Hélène; Kater, Martin M

    2016-04-01

    In rice, inflorescence architecture is established at early stages of reproductive development and contributes directly to grain yield potential. After induction of flowering, the complexity of branching, and therefore the number of seeds on the panicle, is determined by the activity of different meristem types and the timing of transitions between them. Although some of the genes involved in these transitions have been identified, an understanding of the network of transcriptional regulators controlling this process is lacking. To address this we used a precise laser microdissection and RNA-sequencing approach in Oryza sativa ssp. japonica cv. Nipponbare to produce quantitative data that describe the landscape of gene expression in four different meristem types: the rachis meristem, the primary branch meristem, the elongating primary branch meristem (including axillary meristems), and the spikelet meristem. A switch in expression profile between apical and axillary meristem types followed by more gradual changes during transitions in axillary meristem identity was observed, and several genes potentially involved in branching were identified. This resource will be vital for a mechanistic understanding of the link between inflorescence development and grain yield.

  16. Proteomic Analysis of Laser Microdissected Melanoma Cells from Skin Organ Cultures

    PubMed Central

    Hood, Brian L.; Grahovac, Jelena; Flint, Melanie S.; Sun, Mai; Charro, Nuno; Becker, Dorothea; Wells, Alan; Conrads, Thomas P

    2010-01-01

    Gaining insights into the molecular events that govern the progression from melanoma in situ to advanced melanoma, and understanding how the local microenvironment at the melanoma site influences this progression, are two clinically pivotal aspects that to date are largely unexplored. In an effort to identify key regulators of the crosstalk between melanoma cells and the melanoma-skin microenvironment, primary and metastatic human melanoma cells were seeded into skin organ cultures (SOCs), and grown for two weeks. Melanoma cells were recovered from SOCs by laser microdissection and whole-cell tryptic digests analyzed by nanoflow liquid chromatography-tandem mass spectrometry with an LTQ-Orbitrap. The differential protein abundances were calculated by spectral counting, the results of which provides evidence that cell-matrix and cell-adhesion molecules that are upregulated in the presence of these melanoma cells recapitulate proteomic data obtained from comparative analysis of human biopsies of invasive melanoma and a tissue sample of adjacent, non-involved skin. This concordance demonstrates the value of SOCs for conducting proteomic investigations of the melanoma microenvironment. PMID:20459140

  17. Recovery of high-quality RNA from laser capture microdissected human and rodent pancreas

    PubMed Central

    Butler, Alexandra E.; Matveyenko, Aleksey V.; Kirakossian, David; Park, Johanna; Gurlo, Tatyana; Butler, Peter C.

    2016-01-01

    Laser capture microdissection (LCM) is a powerful method to isolate specific populations of cells for subsequent analysis such as gene expression profiling, for example, microarrays or ribonucleic (RNA)-Seq. This technique has been applied to frozen as well as formalin-fixed, paraffin-embedded (FFPE) specimens with variable outcomes regarding quality and quantity of extracted RNA. The goal of the study was to develop the methods to isolate high-quality RNA from islets of Langerhans and pancreatic duct glands (PDG) isolated by LCM. We report an optimized protocol for frozen sections to minimize RNA degradation and maximize recovery of expected transcripts from the samples using quantitative real-time polymerase chain reaction (RT-PCR) by adding RNase inhibitors at multiple steps during the experiment. This technique reproducibly delivered intact RNA (RIN values 6–7). Using quantitative RT-PCR, the expected profiles of insulin, glucagon, mucin6 (Muc6), and cytokeratin-19 (CK-19) mRNA in PDGs and pancreatic islets were detected. The described experimental protocol for frozen pancreas tissue might also be useful for other tissues with moderate to high levels of intrinsic ribonuclease (RNase) activity. PMID:27231405

  18. Laser Capture Microdissection of Feline Streptomyces spp Pyogranulomatous Dermatitis and Cellulitis.

    PubMed

    Traslavina, R P; Reilly, C M; Vasireddy, R; Samitz, E M; Stepnik, C T; Outerbridge, C; Affolter, V K; Byrne, B A; Lowenstine, L J; White, S D; Murphy, B

    2015-11-01

    Suspected Streptomyces spp infections were identified in 4 cats at UC Davis Veterinary Medical Teaching Hospital between 1982 and 2011. Three had ulcerated, dark red mycetomas involving the dermis, subcutis, and fascia with fistulous tracts and/or regional lymphadenopathy. One cat had pyogranulomatous mesenteric lymphadenitis. Granulomatous inflammation in all cats contained colonies of Gram-positive, non-acid-fast organisms. All 4 cats failed to respond to aggressive medical and surgical treatment and were euthanized. Laser capture microdissection (LCM) was used to selectively harvest DNA from the affected formalin-fixed, paraffin-embedded (FFPE) tissues. Cloned amplicons from LCM-derived tissue confirmed the presence of Streptomyces spp in the dermatitis cases. Amplicons from the remaining cat with peritoneal involvement aligned with the 16S ribosomal RNA gene for Actinomycetales. Usually considered a contaminant, Streptomyces spp can be associated with refractory pyogranulomatous dermatitis and cellulitis in cats with outdoor access. LCM is useful in the diagnosis of bacterial diseases where contamination may be an issue. PMID:25516065

  19. Laser Capture Microdissection of Embryonic Cells and Preparation of RNA for Microarray Assays

    PubMed Central

    Redmond, Latasha C.; Pang, Christopher J.; Dumur, Catherine; Haar, Jack L.; Lloyd, Joyce A.

    2014-01-01

    In order to compare the global gene expression profiles of different embryonic cell types, it is first necessary to isolate the specific cells of interest. The purpose of this chapter is to provide a step-by-step protocol to perform laser capture microdissection (LCM) on embryo samples and obtain sufficient amounts of high-quality RNA for microarray hybridizations. Using the LCM/microarray strategy on mouse embryo samples has some challenges, because the cells of interest are available in limited quantities. The first step in the protocol is to obtain embryonic tissue, and immediately cryoprotect and freeze it in a cryomold containing Optimal Cutting Temperature freezing media (Sakura Finetek), using a dry ice–isopentane bath. The tissue is then cryosectioned, and the microscope slides are processed to fix, stain, and dehydrate the cells. LCM is employed to isolate specific cell types from the slides, identified under the microscope by virtue of their morphology. Detailed protocols are provided for using the currently available ArcturusXT LCM instrument and CapSure® LCM Caps, to which the selected cells adhere upon laser capture. To maintain RNA integrity, upon removing a slide from the final processing step, or attaching the first cells on the LCM cap, LCM is completed within 20 min. The cells are then immediately recovered from the LCM cap using a denaturing solution that stabilizes RNA integrity. RNA is prepared using standard methods, modified for working with small samples. To ensure the validity of the microarray data, the quality of the RNA is assessed using the Agilent bioanalyzer. Only RNA that is of sufficient integrity and quantity is used to perform microarray assays. This chapter provides guidance regarding troubleshooting and optimization to obtain high-quality RNA from cells of limited availability, obtained from embryo samples by LCM. PMID:24318813

  20. Laser-assisted Microdissection (LAM) as a Tool for Transcriptional Profiling of Individual Cell Types.

    PubMed

    Florez Rueda, Ana Marcela; Grossniklaus, Ueli; Schmidt, Anja

    2016-01-01

    The understanding of developmental processes at the molecular level requires insights into transcriptional regulation, and thus the transcriptome, at the level of individual cell types. While the methods described here are generally applicable to a wide range of species and cell types, our research focuses on plant reproduction. Plant cultivation and seed production is of crucial importance for human and animal nutrition. A detailed understanding of the regulatory networks that govern the formation of the reproductive lineage (germline) and ultimately of seeds is a precondition for the targeted manipulation of plant reproduction. In particular, the engineering of apomixis (asexual reproduction through seeds) into crop plants promises great improvements, as it leads to the formation of clonal seeds that are genetically identical to the mother plant. Consequently, the cell types of the female germline are of major importance for the understanding and engineering of apomixis. However, as the corresponding cells are deeply embedded within the floral tissues, they are very difficult to access for experimental analyses, including cell-type specific transcriptomics. To overcome this limitation, sections of individual cells can be isolated by laser-assisted microdissection (LAM). While LAM in combination with transcriptional profiling allows the identification of genes and pathways active in any cell type with high specificity, establishing a suitable protocol can be challenging. Specifically, the quality of RNA obtained after LAM can be compromised, especially when small, single cells are targeted. To circumvent this problem, we have established a workflow for LAM that reproducibly results in high RNA quality that is well suitable for transcriptomics, as exemplified here by the isolation of cells of the female germline in apomictic Boechera. In this protocol, procedures are described for tissue preparation and LAM, also with regard to RNA extraction and quality control

  1. Gene expression profiling of gastric cancer by microarray combined with laser capture microdissection

    PubMed Central

    Wu, Ming-Shiang; Lin, Yi-Shing; Chang, Yu-Ting; Shun, Chia-Tung; Lin, Ming-Tsan; Lin, Jaw-Town

    2005-01-01

    AIM: To examine the gene expression profile of gastric cancer (GC) by combination of laser capture microdissection (LCM) and microarray and to correlate the profiling with histological subtypes. METHODS: Using LCM, pure cancer cells were procured from 45 cancerous tissues. After procurement of about 5 000 cells, total RNA was extracted and the quality of RNA was determined before further amplification and hybridization. One microgram of amplified RNA was converted to cDNA and hybridized to cDNA microarray. RESULTS: Among 45 cases, only 21 were qualified for their RNAs. A total of 62 arrays were performed. These included 42 arrays for cancer (21 cases with dye-swab duplication) and 20 arrays for non-tumorous cells (10 cases with dye-swab duplication) with universal reference. Analyzed data showed 504 genes were differentially expressed and could distinguish cancerous and non-cancerous groups with more than 99% accuracy. Of the 504 genes, trefoil factors 1, 2, and 3 were in the list and their expression patterns were consistent with previous reports. Immunohistochemical staining of trefoil factor 1 was also consistent with the array data. Analyses of the tumor group with these 504 genes showed that there were 3 subgroups of GC that did not correspond to any current classification system, including Lauren’s classification. CONCLUSION: By using LCM, linear amplification of RNA, and cDNA microarray, we have identified a panel of genes that have the power to discriminate between GC and non-cancer groups. The new molecular classification and the identified novel genes in gastric carcinogenesis deserve further investigations to elucidate their clinicopathological significance. PMID:16437709

  2. Microarray Cluster Analysis of Irradiated Growth Plate Zones Following Laser Microdissection

    SciTech Connect

    Damron, Timothy A. Zhang Mingliang; Pritchard, Meredith R.; Middleton, Frank A.; Horton, Jason A.; Margulies, Bryan M.; Strauss, Judith A.; Farnum, Cornelia E.; Spadaro, Joseph A.

    2009-07-01

    Purpose: Genes and pathways involved in early growth plate chondrocyte recovery after fractionated irradiation were sought as potential targets for selective radiorecovery modulation. Materials and Methods: Three groups of six 5-week male Sprague-Dawley rats underwent fractionated irradiation to the right tibiae over 5 days, totaling 17.5 Gy, and then were killed at 7, 11, and 16 days after the first radiotherapy fraction. The growth plates were collected from the proximal tibiae bilaterally and subsequently underwent laser microdissection to separate reserve, perichondral, proliferative, and hypertrophic zones. Differential gene expression was analyzed between irradiated right and nonirradiated left tibia using RAE230 2.0 GeneChip microarray, compared between zones and time points and subjected to functional pathway cluster analysis with real-time polymerase chain reaction to confirm selected results. Results: Each zone had a number of pathways showing enrichment after the pattern of hypothesized importance to growth plate recovery, yet few met the strictest criteria. The proliferative and hypertrophic zones showed both the greatest number of genes with a 10-fold right/left change at 7 days after initiation of irradiation and enrichment of the most functional pathways involved in bone, cartilage, matrix, or skeletal development. Six genes confirmed by real-time polymerase chain reaction to have early upregulation included insulin-like growth factor 2, procollagen type I alpha 2, matrix metallopeptidase 9, parathyroid hormone receptor 1, fibromodulin, and aggrecan 1. Conclusions: Nine overlapping pathways in the proliferative and hypertrophic zones (skeletal development, ossification, bone remodeling, cartilage development, extracellular matrix structural constituent, proteinaceous extracellular matrix, collagen, extracellular matrix, and extracellular matrix part) may play key roles in early growth plate radiorecovery.

  3. Combining laser microdissection and RNA-seq to chart the transcriptional landscape of fungal development

    PubMed Central

    2012-01-01

    Background During sexual development, filamentous ascomycetes form complex, three-dimensional fruiting bodies for the protection and dispersal of sexual spores. Fruiting bodies contain a number of cell types not found in vegetative mycelium, and these morphological differences are thought to be mediated by changes in gene expression. However, little is known about the spatial distribution of gene expression in fungal development. Here, we used laser microdissection (LM) and RNA-seq to determine gene expression patterns in young fruiting bodies (protoperithecia) and non-reproductive mycelia of the ascomycete Sordaria macrospora. Results Quantitative analysis showed major differences in the gene expression patterns between protoperithecia and total mycelium. Among the genes strongly up-regulated in protoperithecia were the pheromone precursor genes ppg1 and ppg2. The up-regulation was confirmed by fluorescence microscopy of egfp expression under the control of ppg1 regulatory sequences. RNA-seq analysis of protoperithecia from the sterile mutant pro1 showed that many genes that are differentially regulated in these structures are under the genetic control of transcription factor PRO1. Conclusions We have generated transcriptional profiles of young fungal sexual structures using a combination of LM and RNA-seq. This allowed a high spatial resolution and sensitivity, and yielded a detailed picture of gene expression during development. Our data revealed significant differences in gene expression between protoperithecia and non-reproductive mycelia, and showed that the transcription factor PRO1 is involved in the regulation of many genes expressed specifically in sexual structures. The LM/RNA-seq approach will also be relevant to other eukaryotic systems in which multicellular development is investigated. PMID:23016559

  4. Evaluation of two-dimensional electrophoresis and liquid chromatography – tandem mass spectrometry for tissue-specific protein profiling of laser-microdissected plant samples

    SciTech Connect

    Schad, Martina; Lipton, Mary S.; Giavalisco, Patrick; Smith, Richard D.; Kehr, Julia

    2005-07-14

    Laser microdissection (LM) allows the collection of homogeneous tissue- and cell specific plant samples. The employment of this technique with subsequent protein analysis has thus far not been reported for plant tissues, probably due to the difficulties associated with defining a reasonable cellular morphology and, in parallel, allowing efficient protein extraction from tissue samples. The relatively large sample amount needed for successful proteome analysis is an additional issue that complicates protein profiling on a tissue- or even cell-specific level. In contrast to transcript profiling that can be performed from very small sample amounts due to efficient amplification strategies, there is as yet no amplification procedure for proteins available. In the current study, we compared different tissue preparation techniques prior to LM/laser pressure catapulting (LMPC) with respect to their suitability for protein retrieval. Cryosectioning was identified as the best compromise between tissue morphology and effective protein extraction. After collection of vascular bundles from Arabidopsis thaliana stem tissue by LMPC, proteins were extracted and subjected to protein analysis, either by classical two-dimensional gel electrophoresis (2-DE), or by high-efficiency liquid chromatography (LC) in conjunction with tandem mass spectrometry (MS/MS). Our results demonstrate that both methods can be used with LMPC collected plant material. But because of the significantly lower sample amount required for LC-MS/MS than for 2-DE, the combination of LMPC and LC-MS/MS has a higher potential to promote comprehensive proteome analysis of specific plant tissues.

  5. Laser-capture microdissection of developing barley seeds and cDNA array analysis of selected tissues.

    PubMed

    Thiel, Johannes; Weier, Diana; Weschke, Winfriede

    2011-01-01

    Laser microdissection provides a useful method for isolating specific cell types from complex biological samples for downstream applications. In contrast to the texture of mammalian cells, most plant tissues exhibit a cell organization with hard, cellulose-containing cell walls, large vacuoles, and air spaces, thus complicating tissue preparation and extraction of macromolecules such as DNA and RNA. Especially, barley seeds show cell types with enormous differences in osmolarity (degenerating and differentiating tissues) and contain high amounts of the main storage product starch, thus requiring specific procedures for morphological preservation and RNA extraction. In this study, we report about methods allowing tissue-specific gene expression profiling of developing barley seeds. Details on aspects of tissue preparation, including fixation and embedding procedures, laser-capture microdissection, RNA isolation, and linear mRNA amplification to produce high-quality labelled probes for large-scale expression analysis are provided. Particular emphasis is placed on the fidelity of transcript data obtained by the developed methods in relation to the in vivo transcriptome.

  6. Laser-assisted vascular anastomosis

    NASA Astrophysics Data System (ADS)

    Kao, Race L.; Tsao-Wu, George; Magovern, George J.

    1990-06-01

    The milliwatt CO2 laser and a thermal activated binding compound (20% serum albumin) were used for microvascular anastomoses. Under general anesthesia, the femoral arteries (0.7 to 1.0 mm diameter) of 6 rats were isolated. After the left femoral artery in each rat was clamped and transected, the vessel was held together with 3 equidistant 10-0 Xomed sutures. The cut edges were coated 3 to 4 times with the albumin solution and sealed with the CO2 laser (power density = 120 W/cm2). The binding compound solidified to a translucent tensile substance which supported the anastomosis until self healing and repair were achieved. The right femoral artery was used as sham operated control. Complete hemostasis and patency were observed in every case immediately and at 1, 3, and 6 months following surgery. The binding compound absorbed most of the laser energy thus minimizing thermal injury to the underlying tissue. Mongrel dogs weighing 28 to 33 kg were anesthetized and prepared for sterile surgical procedures. In 5 dogs, the femoral and jugular veins were exposed, transected, and anastomosed using a CO2 laser (Sharplan 1040) with the binding compound. In another 12 dogs, cephalic veins were isolated and used for aortocoronary artery bypass procedures. The Sharplan 1040 CO2 laser and 20% albumin solution were utilized to complete the coronary anastomoses in 6 dogs, and 6 dogs were used as controls by suturing the vessels. Again, hemostasis, patency, and minimal tissue damage were observed immediately and 6 weeks after the procedures. Improved surgical results, reduced operating time, minimized tissue damage, and enhanced anastomotic integrity are the advantages of laser assisted vascular anastomosis with a thermal activated binding compound.

  7. Direct polymerase chain reaction amplification of formalin-fixed, paraffin-wax-embedded tissue after automated sequential laser microdissection and pressure catapulting.

    PubMed

    O'Kane, S L; Garimella, V; Sivarajasingham, N; Drew, P J; Cawkwell, L

    2007-02-01

    A robust method to facilitate rapid laser microdissection and pressure catapulting (LMPC) coupled with direct polymerase chain reaction (dPCR) to eliminate the need for extraction of DNA before a PCR-based assay is described. This sequential LMPC-dPCR method is rapid and decreases the number of processing steps, reducing the chance of tissue loss and contamination.

  8. DETECTION OF K-RAS AND P53 MUTATIONS IN SPUTUM SAMPLES OF LUNG CANCER PATIENTS USING LASER CAPTURE MICRODISSECTION MICROSCOPE AND MUTATION ANALYSIS

    EPA Science Inventory

    Detection of K-ras and p53 Mutations in Sputum Samples of Lung Cancer Patients Using Laser Capture Microdissection Microscope and Mutation Analysis

    Phouthone Keohavong a,*, Wei-Min Gao a, Kui-Cheng Zheng a, Hussam Mady b, Qing Lan c, Mona Melhem b, and Judy Mumford d.
    <...

  9. Laser microdissection coupled with RNA-seq analysis of porcine enterocytes infected with an obligate intracellular pathogen (Lawsonia intracellularis)

    PubMed Central

    2013-01-01

    Background Lawsonia intracellularis is an obligate intracellular bacterium and the etiologic agent of proliferative enteropathy. The disease is endemic in pigs, emerging in horses and has been described in various other species including nonhuman primates. Cell proliferation is associated with bacterial replication in enterocyte cytoplasm, but the molecular basis of the host-pathogen interaction is unknown. We used laser capture microdissection coupled with RNA-seq technology to characterize the transcriptional responses of infected enterocytes and the host-pathogen interaction. Results Proliferative enterocytes was associated with activation of transcription, protein biosynthesis and genes acting on the G1 phase of the host cell cycle (Rho family). The lack of differentiation in infected enterocytes was demonstrated by the repression of membrane transporters related to nutrient acquisition. The activation of the copper uptake transporter by infected enterocytes was associated with high expression of the Zn/Cu superoxide dismutase by L. intracellularis. This suggests that the intracellular bacteria incorporate intracytoplasmic copper and express a sophisticated mechanism to cope with oxidative stress. Conclusions The feasibility of coupling microdissection and RNA-seq was demonstrated by characterizing the host-bacterial interactions from a specific cell type in a heterogeneous tissue. High expression of L. intracellularis genes encoding hypothetical proteins and activation of host Rho genes infers the role of unrecognized bacterial cyclomodulins in the pathogenesis of proliferative enteropathy. PMID:23800029

  10. Fertilization of C57BL/6 mouse sperm collected from cauda epididymides after preservation or transportation at 4 degrees C using laser-microdissected oocytes.

    PubMed

    Kaneko, Takehito; Fukumoto, Kiyoko; Haruguchi, Yukie; Kondo, Tomoko; Machida, Hiromi; Koga, Mika; Nakagawa, Yoshiko; Tsuchiyama, Shuuji; Saiki, Kiyora; Noshiba, Shiho; Nakagata, Naomi

    2009-08-01

    The C57BL/6 mouse is commonly used to produce transgenic and knockout strains for biomedical research. However, the motility and fertility of its sperm decrease markedly with freezing. Short-term preservation of sperm without freezing can avoid this. Furthermore, such samples can be transported safety without the special skills or equipment needed for the transportation of live animals or frozen products. We evaluated the motility and fertility of sperm collected from cauda epididymides after preservation or transportation at 4 degrees C. Oocytes with the zona pellucida subjected to laser-microdissection were used to assist fertilization in vitro. Although the motility of sperm gradually decreased with storage (P<0.05), no disruption of the sperm plasma membrane was seen. The proportion of zona-intact oocytes fertilized with sperm preserved for 0, 24, 48 and 72h were 70, 14, 5 and 1%, respectively. On the other hand, 45, 20 and 14% of laser-microdissected oocytes were fertilized by sperm preserved for 24, 48 and 72h, respectively (P<0.05). The fertility of sperm collected from cauda epididymides of two transgenic strains after transportation at 4 degrees C were also significantly increased using laser-microdissected oocytes rather than zona-intact oocytes (57 and 68% vs. 5%, P<0.05). Efficient production of offspring from sperm preserved or transported at 4 degrees C was achieved using laser-microdissected oocytes. Thus the fertility of sperm preserved or transported at 4 degrees C could be maintained, although motility gradually decreased with storage. Laser-microdissected oocytes will contribute to the efficient production of embryos and offspring using such preserved sperm samples.

  11. Oral vascular malformations: laser treatment and management

    NASA Astrophysics Data System (ADS)

    Romeo, U.; Rocchetti, F.; Gaimari, G.; Tenore, G.; Palaia, G.; Lo Giudice, G.

    2016-03-01

    Vascular malformations are a very heterogeneous group of circulatory system's diseases that can involve different kind of vessels: arterial, venous or lymphatic ones. Many treatments, such as conventional surgery, embolization, steroid therapy and laser therapy, are available for vascular lesions. The laser approach relies more therapeutic techniques: the transmucosal thermophotocoagulation, intralesional photocoagulation, the excisional biopsy. Today laser is demonstrated to be the gold standard technique to treat vascular lesions that allows a safe and efficient treatment and a lower post-operative healing time. The only disadvantage is the risk of carbonization that could be avoided by using the multiple-spot single pulsed wave technique.

  12. Transcriptomic Analysis of Trout Gill Ionocytes in Fresh Water and Sea Water Using Laser Capture Microdissection Combined with Microarray Analysis

    PubMed Central

    Leguen, Isabelle; Le Cam, Aurélie; Montfort, Jérôme; Peron, Sandrine; Fautrel, Alain

    2015-01-01

    Fish gills represent a complex organ composed of several cell types that perform multiple physiological functions. Among these cells, ionocytes are implicated in the maintenance of ion homeostasis. However, because the ionocyte represents only a small percent of whole gill tissue, its specific transcriptome can be overlooked among the numerous cell types included in the gill. The objective of this study is to better understand ionocyte functions by comparing the RNA expression of this cell type in freshwater and seawater acclimated rainbow trout. To realize this objective, ionocytes were captured from gill cryosections using laser capture microdissection after immunohistochemistry. Then, transcriptome analyses were performed on an Agilent trout oligonucleotide microarray. Gene expression analysis identified 108 unique annotated genes differentially expressed between freshwater and seawater ionocytes, with a fold change higher than 3. Most of these genes were up-regulated in freshwater cells. Interestingly, several genes implicated in ion transport, extracellular matrix and structural cellular proteins appeared up-regulated in freshwater ionocytes. Among them, several ion transporters, such as CIC2, SLC26A6, and NBC, were validated by qPCR and/or in situ hybridization. The latter technique allowed us to localize the transcripts of these ion transporters in only ionocytes and more particularly in the freshwater cells. Genes involved in metabolism and also several genes implicated in transcriptional regulation, cell signaling and the cell cycle were also enhanced in freshwater ionocytes. In conclusion, laser capture microdissection combined with microarray analysis allowed for the determination of the transcriptional signature of scarce cells in fish gills, such as ionocytes, and aided characterization of the transcriptome of these cells in freshwater and seawater acclimated trout. PMID:26439495

  13. Pathway-Focused PCR Array Profiling of Enriched Populations of Laser Capture Microdissected Hippocampal Cells after Traumatic Brain Injury

    PubMed Central

    Boone, Deborah R.; Micci, Maria-Adelaide; Taglialatela, Isabella G.; Hellmich, Judy L.; Weisz, Harris A.; Bi, Min; Prough, Donald S.; DeWitt, Douglas S.; Hellmich, Helen L.

    2015-01-01

    Cognitive deficits in survivors of traumatic brain injury (TBI) are associated with irreversible neurodegeneration in brain regions such as the hippocampus. Comparative gene expression analysis of dying and surviving neurons could provide insight into potential therapeutic targets. We used two pathway-specific PCR arrays (RT2 Profiler Apoptosis and Neurotrophins & Receptors PCR arrays) to identify and validate TBI-induced gene expression in dying (Fluoro-Jade-positive) or surviving (Fluoro-Jade- negative) pyramidal neurons obtained by laser capture microdissection (LCM). In the Apoptosis PCR array, dying neurons showed significant increases in expression of genes associated with cell death, inflammation, and endoplasmic reticulum (ER) stress compared with adjacent, surviving neurons. Pro-survival genes with pleiotropic functions were also significantly increased in dying neurons compared to surviving neurons, suggesting that even irreversibly injured neurons are able to mount a protective response. In the Neurotrophins & Receptors PCR array, which consists of genes that are normally expected to be expressed in both groups of hippocampal neurons, only a few genes were expressed at significantly different levels between dying and surviving neurons. Immunohistochemical analysis of selected, differentially expressed proteins supported the gene expression data. This is the first demonstration of pathway-focused PCR array profiling of identified populations of dying and surviving neurons in the brain after TBI. Combining precise laser microdissection of identifiable cells with pathway-focused PCR array analysis is a practical, low-cost alternative to microarrays that provided insight into neuroprotective signals that could be therapeutically targeted to ameliorate TBI-induced neurodegeneration. PMID:26016641

  14. Laser capture microdissection and multiplex-tandem PCR analysis of proximal tubular epithelial cell signaling in human kidney disease.

    PubMed

    Wilkinson, Ray; Wang, Xiangju; Kassianos, Andrew J; Zuryn, Steven; Roper, Kathrein E; Osborne, Andrew; Sampangi, Sandeep; Francis, Leo; Raghunath, Vishwas; Healy, Helen

    2014-01-01

    Interstitial fibrosis, a histological process common to many kidney diseases, is the precursor state to end stage kidney disease, a devastating and costly outcome for the patient and the health system. Fibrosis is historically associated with chronic kidney disease (CKD) but emerging evidence is now linking many forms of acute kidney disease (AKD) with the development of CKD. Indeed, we and others have observed at least some degree of fibrosis in up to 50% of clinically defined cases of AKD. Epithelial cells of the proximal tubule (PTEC) are central in the development of kidney interstitial fibrosis. We combine the novel techniques of laser capture microdissection and multiplex-tandem PCR to identify and quantitate "real time" gene transcription profiles of purified PTEC isolated from human kidney biopsies that describe signaling pathways associated with this pathological fibrotic process. Our results: (i) confirm previous in-vitro and animal model studies; kidney injury molecule-1 is up-regulated in patients with acute tubular injury, inflammation, neutrophil infiltration and a range of chronic disease diagnoses, (ii) provide data to inform treatment; complement component 3 expression correlates with inflammation and acute tubular injury, (iii) identify potential new biomarkers; proline 4-hydroxylase transcription is down-regulated and vimentin is up-regulated across kidney diseases, (iv) describe previously unrecognized feedback mechanisms within PTEC; Smad-3 is down-regulated in many kidney diseases suggesting a possible negative feedback loop for TGF-β in the disease state, whilst tight junction protein-1 is up-regulated in many kidney diseases, suggesting feedback interactions with vimentin expression. These data demonstrate that the combined techniques of laser capture microdissection and multiplex-tandem PCR have the power to study molecular signaling within single cell populations derived from clinically sourced tissue.

  15. Transcriptomic Analysis of Trout Gill Ionocytes in Fresh Water and Sea Water Using Laser Capture Microdissection Combined with Microarray Analysis.

    PubMed

    Leguen, Isabelle; Le Cam, Aurélie; Montfort, Jérôme; Peron, Sandrine; Fautrel, Alain

    2015-01-01

    Fish gills represent a complex organ composed of several cell types that perform multiple physiological functions. Among these cells, ionocytes are implicated in the maintenance of ion homeostasis. However, because the ionocyte represents only a small percent of whole gill tissue, its specific transcriptome can be overlooked among the numerous cell types included in the gill. The objective of this study is to better understand ionocyte functions by comparing the RNA expression of this cell type in freshwater and seawater acclimated rainbow trout. To realize this objective, ionocytes were captured from gill cryosections using laser capture microdissection after immunohistochemistry. Then, transcriptome analyses were performed on an Agilent trout oligonucleotide microarray. Gene expression analysis identified 108 unique annotated genes differentially expressed between freshwater and seawater ionocytes, with a fold change higher than 3. Most of these genes were up-regulated in freshwater cells. Interestingly, several genes implicated in ion transport, extracellular matrix and structural cellular proteins appeared up-regulated in freshwater ionocytes. Among them, several ion transporters, such as CIC2, SLC26A6, and NBC, were validated by qPCR and/or in situ hybridization. The latter technique allowed us to localize the transcripts of these ion transporters in only ionocytes and more particularly in the freshwater cells. Genes involved in metabolism and also several genes implicated in transcriptional regulation, cell signaling and the cell cycle were also enhanced in freshwater ionocytes. In conclusion, laser capture microdissection combined with microarray analysis allowed for the determination of the transcriptional signature of scarce cells in fish gills, such as ionocytes, and aided characterization of the transcriptome of these cells in freshwater and seawater acclimated trout.

  16. Application of laser microdissection ICP-MS for high resolution elemental mapping in mouse brain tissue: a comparative study with laser ablation ICP-MS.

    PubMed

    Sussulini, Alessandra; Becker, J Sabine

    2015-01-01

    Mapping of elements in biological tissue by laser induced mass spectrometry is a fast growing analytical methodology in life sciences. This method provides a multitude of useful information of metal, nonmetal, metalloid and isotopic distribution at major, minor and trace concentration ranges, usually with a lateral resolution of 12-160 µm. Selected applications in medical research require an improved lateral resolution of laser induced mass spectrometric technique at the low micrometre scale and below. The present work demonstrates the applicability of a recently developed analytical methodology - laser microdissection associated to inductively coupled plasma mass spectrometry (LMD ICP-MS) - to obtain elemental images of different solid biological samples at high lateral resolution. LMD ICP-MS images of mouse brain tissue samples stained with uranium and native are shown, and a direct comparison of LMD and laser ablation (LA) ICP-MS imaging methodologies, in terms of elemental quantification, is performed. PMID:25476347

  17. MicroRNA Profiling of Laser-Microdissected Hepatocellular Carcinoma Reveals an Oncogenic Phenotype of the Tumor Capsule123

    PubMed Central

    Peveling-Oberhag, Jan; Seiz, Anna; Döring, Claudia; Hartmann, Sylvia; Köberle, Verena; Liese, Juliane; Zeuzem, Stefan; Hansmann, Martin-Leo; Piiper, Albrecht

    2014-01-01

    Several microRNAs (miRNAs) are associated with the molecular pathogenesis of hepatocellular carcinoma (HCC). However, previous studies analyzing the dysregulation of miRNAs in HCC show heterogeneous results. We hypothesized that part of this heterogeneity might be attributable to variations of miRNA expression deriving from the HCC capsule or the fibrotic septa within the peritumoral tissue used as controls. Tissue from surgically resected hepatitis C–associated HCC from six well-matched patients was microdissected using laser microdissection and pressure catapulting technique. Four distinct histologic compartments were isolated: tumor parenchyma (TP), fibrous capsule of the tumor (TC), tumor-adjacent liver parenchyma (LP), and cirrhotic septa of the tumor-adjacent liver (LC). MiRNA expression profiling analysis of 1105 mature miRNAs and precursors was performed using miRNA microarray. Principal component analysis and consecutive pairwise supervised comparisons demonstrated distinct patterns of expressed miRNAs not only for TP versus LP (e.g., intratumoral down-regulation of miR-214, miR-199a, miR-146a, and miR-125a; P< .05) but also for TC versus LC (including down-regulation within TC of miR-126, miR-99a/100, miR-26a, and miR-125b; P< .05). The tumor capsule therefore demonstrates a tumor-like phenotype with down-regulation of well-known tumor-suppressive miRNAs. Variations of co-analyzed fibrotic tissue within the tumor or in controls may have profound influence on miRNA expression analyses in HCC. Several miRNAs, which are proposed to be HCC specific, may indeed be rather associated to the tumor capsule. As miRNAs evolve to be important biomarkers in liver tumors, the presented data have important translational implications on diagnostics and treatment in patients with HCC. PMID:25500075

  18. Laser capture microdissection of uredinia formed by Melampsora larici-populina revealed a transcriptional switch between biotrophy and sporulation.

    PubMed

    Hacquard, Stéphane; Delaruelle, Christine; Legué, Valérie; Tisserant, Emilie; Kohler, Annegret; Frey, Pascal; Martin, Francis; Duplessis, Sébastien

    2010-10-01

    The foliar rust caused by the basidiomycete Melampsora larici-populina is the main disease affecting poplar plantations in Europe. The biotrophic status of rust fungi is a major limitation to study gene expression of cell or tissue types during host infection. At the uredinial stage, infected poplar leaves contain distinct rust tissues such as haustoria, infection hyphae, and uredinia with sporogenous hyphae and newly formed asexual urediniospores. Laser capture microdissection (LCM) was used to isolate three areas corresponding to uredinia and subjacent zones in the host mesophyll for expression analysis with M. larici-populina whole-genome exon oligoarrays. Optimization of tissue preparation prior to LCM allowed isolation of RNA of good integrity for genome-wide expression profiling. Our results indicate that the poplar rust uredinial stage is marked by distinct genetic programs related to biotrophy in the host palisade mesophyll and to sporulation in the uredinium. A strong induction of transcripts encoding small secreted proteins, likely containing rust effectors, is observed in the mesophyll, suggesting a late maintenance of suppression of host defense in the tissue containing haustoria and infection hyphae. On the other hand, cell cycle and cell defense rescue transcripts are strongly accumulated in the sporulation area. This combined LCM-transcriptomic approach brings new insights on the molecular mechanisms underlying urediniospore formation in rust fungi.

  19. Analysis of Transcription Factor mRNAs in Identified Oxytocin and Vasopressin Magnocellular Neurons Isolated by Laser Capture Microdissection

    PubMed Central

    Rodriguez-Canales, Jaime; Lubelski, Daniel; Rashid, Omar M.; Salinas, Yasmmyn D.; Shi, YiJun; Ponzio, Todd; Fields, Raymond; Emmert-Buck, Michael R.; Gainer, Harold

    2013-01-01

    The oxytocin (Oxt) and vasopressin (Avp) magnocellular neurons (MCNs) in the hypothalamus are the only neuronal phenotypes that are present in the supraoptic nucleus (SON), and are characterized by their robust and selective expression of either the Oxt or Avp genes. In this paper, we take advantage of the differential expression of these neuropeptide genes to identify and isolate these two individual phenotypes from the rat SON by laser capture microdissection (LCM), and to analyze the differential expression of several of their transcription factor mRNAs by qRT-PCR. We identify these neuronal phenotypes by stereotaxically injecting recombinant Adeno-Associated Viral (rAAV) vectors which contain cell-type specific Oxt or Avp promoters that drive expression of EGFP selectively in either the Oxt or Avp MCNs into the SON. The fluorescent MCNs are then dissected by LCM using a novel Cap Road Map protocol described in this paper, and the purified MCNs are extracted for their RNAs. qRT-PCR of these RNAs show that some transcription factors (RORA and c-jun) are differentially expressed in the Oxt and Avp MCNs. PMID:23894472

  20. Laser capture microdissection of intestinal tissue from sea bass larvae using an optimized RNA integrity assay and validated reference genes

    PubMed Central

    Schaeck, M.; De Spiegelaere, W.; De Craene, J.; Van den Broeck, W.; De Spiegeleer, B.; Burvenich, C.; Haesebrouck, F.; Decostere, A.

    2016-01-01

    The increasing demand for a sustainable larviculture has promoted research regarding environmental parameters, diseases and nutrition, intersecting at the mucosal surface of the gastrointestinal tract of fish larvae. The combination of laser capture microdissection (LCM) and gene expression experiments allows cell specific expression profiling. This study aimed at optimizing an LCM protocol for intestinal tissue of sea bass larvae. Furthermore, a 3′/5′ integrity assay was developed for LCM samples of fish tissue, comprising low RNA concentrations. Furthermore, reliable reference genes for performing qPCR in larval sea bass gene expression studies were identified, as data normalization is critical in gene expression experiments using RT-qPCR. We demonstrate that a careful optimization of the LCM procedure allows recovery of high quality mRNA from defined cell populations in complex intestinal tissues. According to the geNorm and Normfinder algorithms, ef1a, rpl13a, rps18 and faua were the most stable genes to be implemented as reference genes for an appropriate normalization of intestinal tissue from sea bass across a range of experimental settings. The methodology developed here, offers a rapid and valuable approach to characterize cells/tissues in the intestinal tissue of fish larvae and their changes following pathogen exposure, nutritional/environmental changes, probiotic supplementation or a combination thereof. PMID:26883391

  1. Towards the finer mapping of facioscapulohumeral muscular dystrophy at 4q35: Construction of a laser microdissection library

    SciTech Connect

    Upadhyaya, M.; Osborn, M.; Maynard, J.

    1995-06-19

    Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal-dominant disorder which has been mapped to the 4q35 region. In order to saturate this distal 4q region with DNA markers, a laser-based chromosomal microdissection and microcloning procedure was used to construct a genomic library from the distal 20% of chromosome 4, derived from a single human metaphase spread. Of the 100 microclones analyzed from this library, 94 clones contained inserts sized from 80-800 bp, with an average size of 340 bp. Less than 20% of these clones hybridized to human repeat sequences. Seventy-two single-copy clones were further characterized by Southern blot hybridization against a DNA panel of somatic cell hybrids, containing various regions of chromosome 4. Forty-two clones mapped to chromosome 4, of which 8 clones mapped into the relevant 4q35 region. Twenty of these chromosome 4-specific clones were screened against {open_quotes}zoo-blots{close_quotes}; 11 clones, of which 3 mapped to 4q35, identified conserved sequences. This is the first report to describe the isolation of potential expressed sequences derived from the FSHD region. These chromosome region-specific microclones will be useful in the construction of the physical map of the region, the positional cloning of potential disease-associated genes, and the identification of additional polymorphic markers from within the distal 4q region. 47 refs., 6 figs., 1 tab.

  2. Global Proteomic Analysis of Functional Compartments in Immature Avian Follicles Using Laser Microdissection Coupled to LC-MS/MS

    PubMed Central

    Nepomuceno, Angelito I.; Muddiman, David C.; Petitte, James N.

    2015-01-01

    Laser microdissection (LMD) was utilized for the separation of the yolk, follicular wall and surrounding stromal cells of small white follicles (SWF) obtained from reproductively active domestic fowl. Herein, we provide an in-situ proteomics based approach to studying follicular development through the use of LMD and mass spectrometry. This study resulted in a total of 2,889 proteins identified from the three specific isolated compartments. White yolk from the smallest avian follicles resulted in the identification of 1,984 proteins, while isolated follicular wall and ovarian stroma yielded 2,470 and 2,456 proteins, respectively. GO annotations highlighted the functional differences between the compartments. Among the three compartments examined, the relative abundance of vitellogenins, steroidogenic enzymes, anti-Mullerian hormone, transcription factors, and proteins involved in retinoic acid receptors/retinoic acid synthesis, transcription factors and cell surface receptors such as EGFR and their associated signaling pathways reflected known cellular function of the ovary. This study has provided a global proteome for SWF, white yolk and ovarian stroma of the avian ovary that can be used as a baseline for future studies and verifies that the coupling of LMD with proteomic analysis can be used to evaluate proteins from small, physiologically functional compartments of complex tissue. PMID:26211554

  3. Gene Expression Profiles of Beta-Cell Enriched Tissue Obtained by Laser Capture Microdissection from Subjects with Type 2 Diabetes

    PubMed Central

    Marselli, Lorella; Thorne, Jeffrey; Dahiya, Sonika; Sgroi, Dennis C.; Sharma, Arun; Bonner-Weir, Susan; Marchetti, Piero; Weir, Gordon C.

    2010-01-01

    Background Changes in gene expression in pancreatic beta-cells from type 2 diabetes (T2D) should provide insights into their abnormal insulin secretion and turnover. Methodology/Principal Findings Frozen sections were obtained from cadaver pancreases of 10 control and 10 T2D human subjects. Beta-cell enriched samples were obtained by laser capture microdissection (LCM). RNA was extracted, amplified and subjected to microarray analysis. Further analysis was performed with DNA-Chip Analyzer (dChip) and Gene Set Enrichment Analysis (GSEA) software. There were changes in expression of genes linked to glucotoxicity. Evidence of oxidative stress was provided by upregulation of several metallothionein genes. There were few changes in the major genes associated with cell cycle, apoptosis or endoplasmic reticulum stress. There was differential expression of genes associated with pancreatic regeneration, most notably upregulation of members of the regenerating islet gene (REG) family and metalloproteinase 7 (MMP7). Some of the genes found in GWAS studies to be related to T2D were also found to be differentially expressed. IGF2BP2, TSPAN8, and HNF1B (TCF2) were upregulated while JAZF1 and SLC30A8 were downregulated. Conclusions/Significance This study made possible by LCM has identified many novel changes in gene expression that enhance understanding of the pathogenesis of T2D. PMID:20644627

  4. Cell type-specific transcriptome of Brassicaceae stigmatic papilla cells from a combination of laser microdissection and RNA sequencing.

    PubMed

    Osaka, Masaaki; Matsuda, Tomoki; Sakazono, Satomi; Masuko-Suzuki, Hiromi; Maeda, Shunsuke; Sewaki, Misato; Sone, Mikako; Takahashi, Hirokazu; Nakazono, Mikio; Iwano, Megumi; Takayama, Seiji; Shimizu, Kentaro K; Yano, Kentaro; Lim, Yong Pyo; Suzuki, Go; Suwabe, Keita; Watanabe, Masao

    2013-11-01

    Pollination is an early and critical step in plant reproduction, leading to successful fertilization. It consists of many sequential processes, including adhesion of pollen grains onto the surface of stigmatic papilla cells, foot formation to strengthen pollen-stigma interaction, pollen hydration and germination, and pollen tube elongation and penetration. We have focused on an examination of the expressed genes in papilla cells, to increase understanding of the molecular systems of pollination. From three representative species of Brassicaceae (Arabidopsis thaliana, A. halleri and Brassica rapa), stigmatic papilla cells were isolated precisely by laser microdissection, and cell type-specific gene expression in papilla cells was determined by RNA sequencing. As a result, 17,240, 19,260 and 21,026 unigenes were defined in papilla cells of A. thaliana, A. halleri and B. rapa, respectively, and, among these, 12,311 genes were common to all three species. Among the17,240 genes predicted in A. thaliana, one-third were papilla specific while approximately half of the genes were detected in all tissues examined. Bioinformatics analysis revealed that genes related to a wide range of reproduction and development functions are expressed in papilla cells, particularly metabolism, transcription and membrane-mediated information exchange. These results reflect the conserved features of general cellular function and also the specific reproductive role of papilla cells, highlighting a complex cellular system regulated by a diverse range of molecules in these cells. This study provides fundamental biological knowledge to dissect the molecular mechanisms of pollination in papilla cells and will shed light on our understanding of plant reproduction mechanisms.

  5. Cell type-specific transcriptome of Brassicaceae stigmatic papilla cells from a combination of laser microdissection and RNA sequencing.

    PubMed

    Osaka, Masaaki; Matsuda, Tomoki; Sakazono, Satomi; Masuko-Suzuki, Hiromi; Maeda, Shunsuke; Sewaki, Misato; Sone, Mikako; Takahashi, Hirokazu; Nakazono, Mikio; Iwano, Megumi; Takayama, Seiji; Shimizu, Kentaro K; Yano, Kentaro; Lim, Yong Pyo; Suzuki, Go; Suwabe, Keita; Watanabe, Masao

    2013-11-01

    Pollination is an early and critical step in plant reproduction, leading to successful fertilization. It consists of many sequential processes, including adhesion of pollen grains onto the surface of stigmatic papilla cells, foot formation to strengthen pollen-stigma interaction, pollen hydration and germination, and pollen tube elongation and penetration. We have focused on an examination of the expressed genes in papilla cells, to increase understanding of the molecular systems of pollination. From three representative species of Brassicaceae (Arabidopsis thaliana, A. halleri and Brassica rapa), stigmatic papilla cells were isolated precisely by laser microdissection, and cell type-specific gene expression in papilla cells was determined by RNA sequencing. As a result, 17,240, 19,260 and 21,026 unigenes were defined in papilla cells of A. thaliana, A. halleri and B. rapa, respectively, and, among these, 12,311 genes were common to all three species. Among the17,240 genes predicted in A. thaliana, one-third were papilla specific while approximately half of the genes were detected in all tissues examined. Bioinformatics analysis revealed that genes related to a wide range of reproduction and development functions are expressed in papilla cells, particularly metabolism, transcription and membrane-mediated information exchange. These results reflect the conserved features of general cellular function and also the specific reproductive role of papilla cells, highlighting a complex cellular system regulated by a diverse range of molecules in these cells. This study provides fundamental biological knowledge to dissect the molecular mechanisms of pollination in papilla cells and will shed light on our understanding of plant reproduction mechanisms. PMID:24058146

  6. Laser microdissection-assisted analysis of the functional fate of iron deficiency-induced root hairs in cucumber.

    PubMed

    Santi, Simonetta; Schmidt, Wolfgang

    2008-01-01

    Iron ranks fourth in the sequence of abundance of the elements in the Earth's crust, but its low bio-availability often limits plant growth. When present in suboptimal amounts, the acquisition of iron by plants is aided by a suite of responses, comprising molecular and developmental changes that facilitate the uptake of iron from sparingly soluble pools. The expression of genes involved in the mobilization of iron (CsHA1), the reduction of ferric chelates (CsFRO1), and in the uptake of ferrous iron (CsIRT1) was investigated in epidermal cells of Fe-sufficient and Fe-deficient cucumber (Cucumis sativum L.) roots using the Laser Microdissection and Pressure Catapulting (LMPC) method. Growing plants hydroponically in media deprived of iron induced the differentiation of almost all epidermal cells into root hairs. No root hairs were formed under iron-replete conditions. The formation of root hairs in response to Fe starvation was associated with a dramatic increase in message levels of CsFRO1, CsIRT1, and the iron-inducible H(+)-ATPase isoform CsHA1, when compared to epidermal cells of Fe-sufficient plants. On the contrary, transcripts of a housekeeping ATPase isoform, CsHA2, were not detected in root hairs, suggesting that Fe-deficiency-induced acidification is predominantly mediated by CsHA1. These data show that the formation of root hairs in response to iron deficiency is associated with cell-specific accumulation of transcripts that are involved in iron acquisition. The results also show that this includes the differential regulation of ATPase isoforms with similar function, but supposedly different characteristics, to counteract the imbalance in nutrient supply efficiently. PMID:18316319

  7. Gene Expression Analysis of Neurons and Astrocytes Isolated by Laser Capture Microdissection from Frozen Human Brain Tissues.

    PubMed

    Tagliafierro, Lidia; Bonawitz, Kirsten; Glenn, Omolara C; Chiba-Falek, Ornit

    2016-01-01

    Different cell types and multiple cellular connections characterize the human brain. Gene expression analysis using a specific population of cells is more accurate than conducting analysis of the whole tissue homogenate, particularly in the context of neurodegenerative diseases, where a specific subset of cells is affected by the different pathology. Due to the difficulty of obtaining homogenous cell populations, gene expression in specific cell-types (neurons, astrocytes, etc.) has been understudied. To leverage the use of archive resources of frozen human brains in studies of neurodegenerative diseases, we developed and calibrated a method to quantify cell-type specific-neuronal, astrocytes-expression profiles of genes implicated in neurodegenerative diseases, including Parkinson's and Alzheimer's diseases. Archive human frozen brain tissues were used to prepare slides for rapid immunostaining using cell-specific antibodies. The immunoreactive-cells were isolated by Laser Capture Microdissection (LCM). The enrichment for a particular cell-type of interest was validated in post-analysis stage by the expression of cell-specific markers. We optimized the technique to preserve the RNA integrity, so that the RNA was suitable for downstream expression analyses. Following RNA extraction, the expression levels were determined digitally using nCounter Single Cell Gene Expression assay (NanoString Technologies®). The results demonstrated that using our optimized technique we successfully isolated single neurons and astrocytes from human frozen brain tissues and obtained RNA of a good quality that was suitable for mRNA expression analysis. We present here new advancements compared to previous reported methods, which improve the method's feasibility and its applicability for a variety of downstream molecular analyses. Our new developed method can be implemented in genetic and functional genomic research of neurodegenerative diseases and has the potential to significantly

  8. Gene Expression Analysis of Neurons and Astrocytes Isolated by Laser Capture Microdissection from Frozen Human Brain Tissues

    PubMed Central

    Tagliafierro, Lidia; Bonawitz, Kirsten; Glenn, Omolara C.; Chiba-Falek, Ornit

    2016-01-01

    Different cell types and multiple cellular connections characterize the human brain. Gene expression analysis using a specific population of cells is more accurate than conducting analysis of the whole tissue homogenate, particularly in the context of neurodegenerative diseases, where a specific subset of cells is affected by the different pathology. Due to the difficulty of obtaining homogenous cell populations, gene expression in specific cell-types (neurons, astrocytes, etc.) has been understudied. To leverage the use of archive resources of frozen human brains in studies of neurodegenerative diseases, we developed and calibrated a method to quantify cell-type specific—neuronal, astrocytes—expression profiles of genes implicated in neurodegenerative diseases, including Parkinson's and Alzheimer's diseases. Archive human frozen brain tissues were used to prepare slides for rapid immunostaining using cell-specific antibodies. The immunoreactive-cells were isolated by Laser Capture Microdissection (LCM). The enrichment for a particular cell-type of interest was validated in post-analysis stage by the expression of cell-specific markers. We optimized the technique to preserve the RNA integrity, so that the RNA was suitable for downstream expression analyses. Following RNA extraction, the expression levels were determined digitally using nCounter Single Cell Gene Expression assay (NanoString Technologies®). The results demonstrated that using our optimized technique we successfully isolated single neurons and astrocytes from human frozen brain tissues and obtained RNA of a good quality that was suitable for mRNA expression analysis. We present here new advancements compared to previous reported methods, which improve the method's feasibility and its applicability for a variety of downstream molecular analyses. Our new developed method can be implemented in genetic and functional genomic research of neurodegenerative diseases and has the potential to

  9. Analysis of cannabinoids in laser-microdissected trichomes of medicinal Cannabis sativa using LCMS and cryogenic NMR.

    PubMed

    Happyana, Nizar; Agnolet, Sara; Muntendam, Remco; Van Dam, Annie; Schneider, Bernd; Kayser, Oliver

    2013-03-01

    Trichomes, especially the capitate-stalked glandular hairs, are well known as the main sites of cannabinoid and essential oil production of Cannabis sativa. In this study the distribution and density of various types of Cannabis sativa L. trichomes, have been investigated by scanning electron microscopy (SEM). Furthermore, glandular trichomes were isolated over the flowering period (8 weeks) by laser microdissection (LMD) and the cannabinoid profile analyzed by LCMS. Cannabinoids were detected in extracts of 25-143 collected cells of capitate-sessile and capitate stalked trichomes and separately in the gland (head) and the stem of the latter. Δ(9)-Tetrahydrocannabinolic acid [THCA (1)], cannabidiolic acid [CBDA (2)], and cannabigerolic acid [CBGA (3)] were identified as most-abundant compounds in all analyzed samples while their decarboxylated derivatives, Δ(9)-tetrahydrocannabinol [THC (4)], cannabidiol [CBD (5)], and cannabigerol [CBG (6)], co-detected in all samples, were present at significantly lower levels. Cannabichromene [CBC (8)] along with cannabinol (CBN (9)) were identified as minor compounds only in the samples of intact capitate-stalked trichomes and their heads harvested from 8-week old plants. Cryogenic nuclear magnetic resonance spectroscopy (NMR) was used to confirm the occurrence of major cannabinoids, THCA (1) and CBDA (2), in capitate-stalked and capitate-sessile trichomes. Cryogenic NMR enabled the additional identification of cannabichromenic acid [CBCA (7)] in the dissected trichomes, which was not possible by LCMS as standard was not available. The hereby documented detection of metabolites in the stems of capitate-stalked trichomes indicates a complex biosynthesis and localization over the trichome cells forming the glandular secretion unit.

  10. Isolating RNA from precursor and mature melanocytes from human vitiligo and normal skin using laser capture microdissection.

    PubMed

    Goldstein, Nathaniel B; Koster, Maranke I; Hoaglin, Laura G; Wright, Michael J; Robinson, Steven E; Robinson, William A; Roop, Dennis R; Norris, David A; Birlea, Stanca A

    2016-10-01

    To characterize the gene expression profile of regenerated melanocytes in the narrow band UVB (NBUVB)-treated vitiligo epidermis and their precursors in the hair follicle, we present here a strategy of RNA isolation from in situ melanocytes using human frozen skin. We developed a rapid immunostaining protocol using the NKI-beteb antibody, which labels differentiated and precursor melanocytes, followed by fluorescent laser capture microdissection. This technique enabled the direct isolation, from melanocyte and adjacent keratinocyte populations, of satisfactory quality RNA that was successfully amplified and analysed by qRT-PCR. The melanocyte-specific gene transcripts TYR, DCT, TYRP1 and PMEL were significantly upregulated in our NBUVB-treated melanocyte samples as compared with the keratinocyte samples, while keratinocyte-specific genes (KRT5 and KRT14) were expressed significantly higher in the keratinocyte samples as compared with the melanocyte samples. Furthermore, in both NBUVB-treated vitiligo skin and normal skin, when bulge melanocytes were compared with epidermal melanocytes, we found significantly lower expression of melanocyte-specific genes and significantly higher expression of three melanocytic stem cell genes (SOX9, WIF1 and SFRP1), while ALCAM and ALDH1A1 transcripts did not show significant variation. We found significantly higher expression of melanocyte-specific genes in the epidermis of NBUVB-treated vitiligo, as compared to the normal skin. When comparing bulge melanocyte samples from untreated vitiligo, NBUVB-treated vitiligo and normal skin, we did not find significant differences in the expression of melanocyte-specific genes or melanocytic stem cell genes. These techniques offer valuable opportunities to study melanocytes and their precursors in vitiligo and other pigmentation disorders.

  11. Direct PCR amplification of the 16S rRNA gene from single microbial cells isolated from an Antarctic iceberg using laser microdissection microscopy

    NASA Astrophysics Data System (ADS)

    Yanagihara, Katsuhiko; Niki, Hironori; Baba, Tomoya

    2011-09-01

    Here, we describe a technique that allows the genetic linage analysis of 16S rRNA genes in bacteria observed under a microscope. The technique includes the isolation of microbial cells using a laser microdissection microscope, lysis of the cells, and amplification of the 16S rRNA genes in the isolated cells without interference by bacterial DNA contamination from the experimental environment or reagents. Using this technique, we successfully determined 15 16S rRNA gene sequences in cells isolated from an Antarctic iceberg. These sequences showed 94%-100% identity to their closest strains, which included bacteria that occur in aqueous, marine, and soil environments.

  12. Tissue-specific metabolite profiling of Turmeric by using laser micro-dissection, ultra-high performance liquid chromatography-quadrupole time of fight-mass spectrometry and liquid chromatography-tandem mass spectrometry.

    PubMed

    Jaiswal, Yogini; Liang, Zhitao; Ho, Alan; Chen, Hubiao; Zhao, Zhongzhen

    2014-01-01

    Curcuma longa L. is recognized for its therapeutic and culinary uses both in Ayurveda and traditional Chinese medicine and is considered to be a boon to mankind. It has been extensively studied for its benefits and still continues to be an important drug with continued potential for further exploration and research. We studied the tissue-specific distribution of secondary metabolites to establish the validity of the use of rhizome samples from India and China, as substitutes for each other, based upon their metabolite profiles and curcumin contents. Laser microdissection was used for the isolation of microscopic tissues, such as cork, cortex and leaf-trace vascular bundles from rhizomes. Metabolite profiling was carried out by ultra-high performance liquid chromatography-quadrupole time of fight-mass spectrometry and curcumin content was estimated by a method validated as per the Harmonized Tripartite Guidelines. The cortex and cork revealed the presence of a higher number of secondary metabolites than in the leaf-trace vascular bundles. The curcumin contents in rhizome samples from both the countries, estimated with the help of a precise and accurate validated method, were found to be comparable. Based on the results, we conclude that turmeric rhizomes grown in India and China are qualitatively and quantitatively indistinguishable and therefore can be used as substitutes. The developed method can be widely applied for microscopic identification, authentication and analysis of the distribution of phytoconstituents in other botanical species of interest or of species with a significant commercial and therapeutic value. PMID:25707128

  13. Analysis of laser-microdissected prostate cancer tissues reveals potential tumor markers.

    PubMed

    Shaikhibrahim, Zaki; Lindstrot, Andreas; Buettner, Reinhard; Wernert, Nicolas

    2011-10-01

    Prostate cancer (PCA) is a clinically heterogeneous and often multifocal disease with a clinical outcome difficult to predict. A deeper knowledge of the molecular basis of the disease may lead to a better prediction of prognosis. Therefore, in this study we investigated the molecular basis of PCA by identifying potential tumor markers in laser-microdisected PCA tissues. Among a group of PCA patients, quantitative RT-PCR analysis was performed to compare the expression of 70 genes. These genes were selected from the results of two microarrays which investigated the gene expression profile differences between moderately or poorly differentiated prostate carcinoma glands and the corresponding normal glands. Among the genes examined, CDKN2A, GATA3, CREBBP, ITGA2, NBL1 and TGM4 were down-regulated in the prostate carcinoma glands compared to the corresponding normal glands, whereas TFF3, TMPRSS2 and ERG were up-regulated. Our findings indicate that these genes may play roles as tumor suppressor genes or oncogenes in PCA, and may serve as potential tumor markers and novel therapeutic targets.

  14. Cytogenetic Characterization of the TM4 Mouse Sertoli Cell Line. II. Chromosome Microdissection, FISH, Scanning Electron Microscopy, and Confocal Laser Scanning Microscopy.

    PubMed

    Schmid, Michael; Guttenbach, Martina; Steinlein, Claus; Wanner, Gerhard; Houben, Andreas

    2015-01-01

    The chromosomes and interphase cell nuclei of the permanent mouse Sertoli cell line TM4 were examined by chromosome microdissection, FISH, scanning electron microscopy, and confocal laser scanning microscopy. The already known marker chromosomes m1-m5 were confirmed, and 2 new large marker chromosomes m6 and m7 were characterized. The minute heterochromatic marker chromosomes m4 and m5 were microdissected and their DNA amplified by DOP-PCR. FISH of this DNA probe on TM4 metaphase chromosomes demonstrated that the m4 and m5 marker chromosomes have derived from the centromeric regions of normal telocentric mouse chromosomes. Ectopic pairing of the m4 and m5 marker chromosomes with the centromeric region of any of the other chromosomes (centromeric associations) was apparent in ∼60% of the metaphases. Scanning electron microscopy revealed DNA-protein bridges connecting the centromeric regions of normal chromosomes and the associated m4 and m5 marker chromosomes. Interphase cell nuclei of TM4 Sertoli cells did not exhibit the characteristic morphology of Sertoli cells in the testes of adult mice as shown by fluorescence microscopy and confocal laser scanning microscopy. PMID:26900862

  15. Gene expression profiling of neurochemically-defined regions of the human brain by in situ hybridization-guided laser capture microdissection

    PubMed Central

    Bernard, René; Kerman, Ilan A.; Meng, Fan; Evans, Simon J.; Amrein, Irmgard; Jones, Edward G.; Bunney, William E.; Akil, Huda; Watson, Stanley J.; Thompson, Robert C.

    2009-01-01

    Laser capture microdissection (LCM) permits isolation of specific cell types and cell groups based upon morphology, anatomical landmarks and histochemical properties. This powerful technique can be used for region-specific dissection if the target structure is clearly delineated. However, it is difficult to visualize anatomical boundaries in an unstained specimen, while histological staining can complicate the microdissection process and compromise downstream processing and analysis. We now introduce a novel method in which in situ hybridization (ISH) signal is used to guide LCM on adjacent unstained sections to collect tissue from neurochemically-defined regions of the human postmortem brain to minimize sample manipulation prior to analysis. This approach was validated in nuclei that provide monoaminergic inputs to the forebrain, and likely contribute to the pathophysiology of mood disorders. This method was used successfully to carry out gene expression profiling and quantitative real-time PCR (qPCR) confirmation from the dissected material. When compared to traditional micropunch dissections, our ISH-guided LCM method provided enhanced signal intensity for mRNAs of specific monoaminergic marker genes as measured by genome-wide gene expression microarrays. Enriched expression of specific monoaminergic genes (as determined by microarrays and qPCR) was detected within appropriate anatomical locations validating the accuracy of microdissection. Together these results support the conclusion that ISH-guided LCM permits acquisition of enriched nucleus-specific RNA that can be successfully used for downstream gene expression investigations. Future studies will utilize this approach for gene expression profiling of neurochemically-defined regions of postmortem brains collected from mood disorder patients. PMID:19070632

  16. Biomarker discovery and identification in laser microdissected head and neck squamous cell carcinoma with ProteinChip technology, two-dimensional gel electrophoresis, tandem mass spectrometry, and immunohistochemistry.

    PubMed

    Melle, Christian; Ernst, Gunther; Schimmel, Bettina; Bleul, Annett; Koscielny, Sven; Wiesner, Andreas; Bogumil, Ralf; Moller, Ursula; Osterloh, Dirk; Halbhuber, Karl-Jurgen; von Eggeling, Ferdinand

    2003-07-01

    Head and neck cancer is a frequent malignancy with a complex, and up to now not clear etiology. Therefore, despite of improvements in diagnosis and therapy, the survival rate with head and neck squamous-cell carcinomas is poor. For a better understanding of the molecular mechanisms behind the process of tumorigenesis and tumor progression, we have analyzed changes of protein expression between microdissected normal pharyngeal epithelium and tumor tissue by ProteinChip technology. For this, cryostat sections from head and neck tumors (n = 57) and adjacent mucosa (n = 44) were laser-microdissected and analyzed on ProteinChip arrays. The derived mass spectrometry profiles exhibited numerous statistical differences. One peak significantly higher expressed in the tumor (p = 0.000029) was isolated by two-dimensional gel electrophoresis and identified as annexin V by in-gel proteolytic digestion, peptide mapping, tandem mass spectrometry analysis, and immuno-deplete assay. The relevance of this single marker protein was further evaluated by immunohistochemistry. Annexin-positive tissue areas were re-analyzed on ProteinChip arrays to confirm the identity of this protein. In this study, we could show that biomarker in head and neck cancer can be found, identified, and assessed by combination of ProteinChip technology, two-dimensional gel electrophoresis, and immunohistochemistry. In our experience, however, such studies only make sense if a relatively pure microdissected tumor tissue is used. Only then minute changes in protein expression between normal pharyngeal epithelium and tumor tissue can be detected, and it will become possible to educe a tumor-associated protein pattern that might be used as a marker for tumorigenesis and progression. PMID:12824440

  17. Selective vascular injury during cutaneous laser therapy

    NASA Astrophysics Data System (ADS)

    Tunnell, James William

    Pulsed laser irradiation in conjunction with cryogen spray cooling (CSC) can induce selective vascular injury to remove cutaneous hypervascular malformations such as port wine stains (PWS), hemangiomas, and facial veins. In this group of studies, we characterized the cryogen heat removal process and determined the effects of pulsed laser irradiation in conjunction with cryogen spray cooling of human skin. First, we employed an inverse heat conduction algorithm to measure the thermal boundary condition due to CSC in in vitro skin phantoms. Second, we developed a mathematical model of laser irradiation in conjunction with CSC in human skin. We determined tissue damage and temperature profiles due to varying combinations of laser pulse duration, radiant exposure, and CSC application times. Finally, we used ex vivo and in vivo human skin to determine the effects of high radiant exposures and CSC on epidermal and vascular injury. CSC induces a dynamic cooling effect, removing heat from the skin both during and following the spurt application time. Residual cryogen, deposited on the skin surface during the cryogen spurt, remains on the skin surface several times as long as the as cryogen spurt itself. The heat removal rate during the cryogen spurt is greatest; however, the total energy removed following the cryogen spurt is also substantial (approximately half as much as during the spurt application time). CSC was effective in protecting the human skin epidermis in light to moderately pigmented skin. Mathematical modeling, ex vivo, and in vivo studies showed that the epidermal damage threshold could be increased by a factor of approximately two. Increased radiant exposures increased the risk of non-selective vascular injury observed in histology as injury to the epidermis and perivascular collagen; however, proper choice of cryogen cooling durations resulted in the elimination of epidermal injury as well as perivascular tissue injury. In addition, higher radiant

  18. Laser microdissection-based analysis of the Y sex chromosome of the Antarctic fish Chionodracohamatus (Notothenioidei, Channichthyidae).

    PubMed

    Cocca, Ennio; Petraccioli, Agnese; Morescalchi, Maria Alessandra; Odierna, Gaetano; Capriglione, Teresa

    2015-01-01

    Microdissection, DOP-PCR amplification and microcloning were used to study the large Y chromosome of Chionodracohamatus, an Antarctic fish belonging to the Notothenioidei, the dominant component of the Southern Ocean fauna. The species has evolved a multiple sex chromosome system with digametic males showing an X1YX2 karyotype and females an X1X1X2X2 karyotype. Fluorescence in situ hybridization, performed with a painting probe made from microdissected Y chromosomes, allowed a deeper insight on the chromosomal rearrangement, which underpinned the fusion event that generated the Y. Then, we used a DNA library established by microdissection and microcloning of the whole Y chromosome of Chionodracohamatus for searching sex-linked sequences. One clone provided preliminary information on the presence on the Y chromosome of the CHD1 gene homologue, which is sex-linked in birds but in no other vertebrates. Several clones from the Y-chromosome mini-library contained microsatellites and transposable elements, one of which mapped to the q arm putative fusion region of the Y chromosome. The findings confirm that interspersed repetitive sequences might have fostered chromosome rearrangements and the emergence of the Y chromosome in Chionodracohamatus. Detection of the CHD1 gene in the Y sex-determining region could be a classical example of convergent evolution in action.

  19. Laser microdissection-based analysis of the Y sex chromosome of the Antarctic fish Chionodraco hamatus (Notothenioidei, Channichthyidae)

    PubMed Central

    Cocca, Ennio; Petraccioli, Agnese; Morescalchi, Maria Alessandra; Odierna, Gaetano; Capriglione, Teresa

    2015-01-01

    Abstract Microdissection, DOP-PCR amplification and microcloning were used to study the large Y chromosome of Chionodraco hamatus, an Antarctic fish belonging to the Notothenioidei, the dominant component of the Southern Ocean fauna. The species has evolved a multiple sex chromosome system with digametic males showing an X1YX2 karyotype and females an X1X1X2X2 karyotype. Fluorescence in situ hybridization, performed with a painting probe made from microdissected Y chromosomes, allowed a deeper insight on the chromosomal rearrangement, which underpinned the fusion event that generated the Y. Then, we used a DNA library established by microdissection and microcloning of the whole Y chromosome of Chionodraco hamatus for searching sex-linked sequences. One clone provided preliminary information on the presence on the Y chromosome of the CHD1 gene homologue, which is sex-linked in birds but in no other vertebrates. Several clones from the Y-chromosome mini-library contained microsatellites and transposable elements, one of which mapped to the q arm putative fusion region of the Y chromosome. The findings confirm that interspersed repetitive sequences might have fostered chromosome rearrangements and the emergence of the Y chromosome in Chionodraco hamatus. Detection of the CHD1 gene in the Y sex-determining region could be a classical example of convergent evolution in action. PMID:25893071

  20. Redistribution of Ionotropic Glutamate Receptors Detected by Laser Microdissection of the Rat Dentate Gyrus 48 h following LTP Induction In Vivo

    PubMed Central

    Kennard, Jeremy T. T.; Guévremont, Diane; Mason-Parker, Sara E.; Abraham, Wickliffe C.; Williams, Joanna M.

    2014-01-01

    The persistence and input specificity of long-term potentiation (LTP) make it attractive as a mechanism of information storage. In its initial phase, both in vivo and in vitro studies have shown that LTP is associated with increased membrane localization of AMPA receptor subunits, but the molecular basis of LTP maintenance over the long-term is still unclear. We have previously shown that expression of AMPA and NMDA receptor subunits is elevated in whole homogenates prepared from dentate gyrus 48 h after LTP induction in vivo. In the present study, we utilized laser microdissection (LMD) techniques to determine whether AMPA and NMDA receptor upregulation occurs specifically in the stimulated regions of the dentate gyrus dendritic arbor. Receptor proteins GluN1, GluA1 and GluA2, as well as postsynaptic density protein of 95 kDa and tubulin were detected by Western blot analysis in microdissected samples. Gradients of expression were observed for GluN1 and GluA2, decreasing from the inner to the outer zones of the molecular layer, and were independent of LTP. When induced at medial perforant path synapses, LTP was associated with an apparent specific redistribution of GluA1 and GluN1 to the middle molecular layer that contains these synapses. These data indicate that glutamate receptor proteins are delivered specifically to dendritic regions possessing LTP-expressing synapses, and that these changes are preserved for at least 48 h. PMID:24667777

  1. Use of laser microdissection for the construction of Humulusjaponicus Siebold et Zuccarini, 1846 (Cannabaceae) sex chromosome-specific DNA library and cytogenetics analysis.

    PubMed

    Yakovin, Nickolay A; Divashuk, Mikhail G; Razumova, Olga V; Soloviev, Alexander A; Karlov, Gennady I

    2014-01-01

    Dioecy is relatively rare among plant species, and distinguishable sex chromosomes have been reported in few dioecious species. The multiple sex chromosome system (XX/XY1Y2) of Humulusjaponicus Siebold et Zuccarini, 1846 differs from that of other members of the family Cannabaceae, in which the XX/XY chromosome system is present. Sex chromosomes of Humulusjaponicus were isolated from meiotic chromosome spreads of males by laser microdissection with the P.A.L.M. MicroLaser system. The chromosomal DNA was directly amplified by degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR). Fast fluorescence in situ hybridization (FAST-FISH) using a labeled, chromosome-specific DOP-PCR product as a probe showed preferential hybridization to sex chromosomes. In addition, the DOP-PCR product was used to construct a short-insert, Humulusjaponicus sex chromosomes-specific DNA library. The randomly sequenced clones showed that about 12% of them have significant homology to Humuluslupulus and 88% to Cannabissativa Linnaeus, 1753 sequences from GenBank database. Forty-four percent of the sequences show homology to plant retroelements. It was concluded that laser microdissection is a useful tool for isolating the DNA of sex chromosomes of Humulusjaponicus and for the construction of chromosome-specific DNA libraries for the study of the structure and evolution of sex chromosomes. The results provide the potential for identifying unique or sex chromosome-specific sequence elements in Humulusjaponicus and could aid in the identification of sex chromosome-specific repeat and coding regions through chromosome isolation and genome complexity reduction. PMID:25610546

  2. Use of laser microdissection for the construction of Humulus japonicus Siebold et Zuccarini, 1846 (Cannabaceae) sex chromosome-specific DNA library and cytogenetics analysis

    PubMed Central

    Yakovin, Nickolay A.; Divashuk, Mikhail G.; Razumova, Olga V.; Soloviev, Alexander A.; Karlov, Gennady I.

    2014-01-01

    Abstract Dioecy is relatively rare among plant species, and distinguishable sex chromosomes have been reported in few dioecious species. The multiple sex chromosome system (XX/XY1Y2) of Humulus japonicus Siebold et Zuccarini, 1846 differs from that of other members of the family Cannabaceae, in which the XX/XY chromosome system is present. Sex chromosomes of Humulus japonicus were isolated from meiotic chromosome spreads of males by laser microdissection with the P.A.L.M. MicroLaser system. The chromosomal DNA was directly amplified by degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR). Fast fluorescence in situ hybridization (FAST-FISH) using a labeled, chromosome-specific DOP-PCR product as a probe showed preferential hybridization to sex chromosomes. In addition, the DOP-PCR product was used to construct a short-insert, Humulus japonicus sex chromosomes-specific DNA library. The randomly sequenced clones showed that about 12% of them have significant homology to Humulus lupulus and 88% to Cannabis sativa Linnaeus, 1753 sequences from GenBank database. Forty-four percent of the sequences show homology to plant retroelements. It was concluded that laser microdissection is a useful tool for isolating the DNA of sex chromosomes of Humulus japonicus and for the construction of chromosome-specific DNA libraries for the study of the structure and evolution of sex chromosomes. The results provide the potential for identifying unique or sex chromosome-specific sequence elements in Humulus japonicus and could aid in the identification of sex chromosome-specific repeat and coding regions through chromosome isolation and genome complexity reduction. PMID:25610546

  3. Laser Capture Microdissection Revisited as a Tool for Transcriptomic Analysis: Application of an Excel-Based qPCR Preparation Software (PREXCEL-Q)

    PubMed Central

    Sow, Fatoumata B.; Gallup, Jack M.; Sacco, Randy E.; Ackermann, Mark R.

    2009-01-01

    The ability to reliably analyze cellular and molecular profiles of normal or diseased tissues is frequently complicated by the inherent heterogeneous nature of tissues. Laser Capture Microdissection (LCM) is an innovative technique that allows the isolation and enrichment of pure subpopulations of cells from tissues under direct microscopic examination. Material obtained by LCM can be used for downstream assays including gene microarrays, western blotting, cDNA library generation and DNA genotyping. We describe a series of LCM protocols for cell collection, RNA extraction and qPCR gene expression analysis. Using reagents we helped develop commercially, we focus on two LCM approaches: laser cutting and laser capture. Reagent calculations have been pre-determined for 10 samples using the new PREXCEL-Q assay development and project management software. One can expect the entire procedure for laser cutting coupled to qPCR to take approximately 12.5-15 h, and laser capture coupled to qPCR to take approximately 13.5-17.5 h. PMID:20556230

  4. Laser treatment of oral vascular malformations

    NASA Astrophysics Data System (ADS)

    Romeo, U.; Gaimari, G.; Mohsen, M.; Tenore, G.; Palaia, G.

    2014-01-01

    Oral Vascular Malformations (OVM) are congenital anomalies characterized by morph-structural and/or functional changes of nature in severity and extension. OVM can affect any type of vessels arterial, venous or lymphatic and any capillary or anatomical. They are divided into two categories: low and high flow. In this study were treated 40 patients with OVM with a range size from 2 mm to 44 mm; they were subjected to clinical examination supported by Colour-Doppler Ultrasound instrumental examination and only for doubt cases the Magnetic Resonance Imaging (MRI) was prescribed. Only low flow venous and capillary malformations were treated by GaAlAs laser (Wiser®, Lambda, Brindole,Italy, 980nm) and KTP laser (SmartLite®, DEKA, Florence, Italy, 532nm) with two different techniques: the Transmucosal Thermophotocoagulation (TMT) and the Intralesional Photocoagulation (ILP). These techniques permitted a good control of haemostasis, avoiding bleeding both during surgery and in the postoperative. It is obtained an excellent and good healing respectively in 10% and 60% of cases, a moderate and poor resolution respectively in 22.5% and 7.5% of cases. A clear diagnosis allowed the management of Venous malformations (VM) by laser devices with wavelengths highly absorbed in haemoglobin in safety and efficacy and according to the principles of minimal invasive surgery. The aim of this study was to verify if the laser is effective in the treatment of OVM for the purpose of the clinical findings and the postoperative course. The Authors concluded that the laser can be considered the "gold standard" for treating OVM.

  5. Localization of ginsenosides in the rhizome and root of Panax ginseng by laser microdissection and liquid chromatography-quadrupole/time of flight-mass spectrometry.

    PubMed

    Liang, Zhitao; Chen, Yujie; Xu, Liang; Qin, Minjian; Yi, Tao; Chen, Hubiao; Zhao, Zhongzhen

    2015-02-01

    The root and rhizome of Panax ginseng C.A. Mey, known as ginseng, is a commonly used medicinal plant. Ginsenosides are the major active components responsible for the tonic effects of this herb. Here, the combination of laser microdissection and ultra-high performance liquid chromatography quadrupole/time of flight-mass spectrometry (UHPLC-QTOF-MS) was applied to investigate the localization of ginsenosides in root and rhizome of P. ginseng. Five kinds of tissue cells were separated from the rhizome, main root and branch root of ginseng. Fifty-nine ginsenosides were identified and the results showed that the cork contained more kinds of ginsenosides than did the cortex, phloem, xylem and resin canals. It is interesting that the phloem, xylem and resin canals from branch root contained a greater number of ginsenosides than did from main root. This study provides solid evidence on the accumulation of ginsenosides in cork, cortex, phloem and xylem.

  6. Laser-based microdissection of single cells from tissue sections and PCR analysis of rearranged immunoglobulin genes from isolated normal and malignant human B cells.

    PubMed

    Küppers, Ralf; Schneider, Markus; Hansmann, Martin-Leo

    2013-01-01

    Normal and malignant B cells carry rearranged immunoglobulin (Ig) variable region genes, which due to their practically limitless diversity represent ideal clonal markers for these cells. We describe here an approach to isolate single cells from frozen tissue sections by microdissection using a laser-based method. From the isolated cells rearranged IgH and Igκ genes are amplified in a semi-nested PCR approach, using a collection of V gene family-specific primers recognizing nearly all V gene segments together with primers for the J gene segments. By sequence analysis of V genes from distinct cells, the clonal relationship of the B lineage cells can unequivocally be determined and related to the histological distribution of the cells. The approach is also useful to determine V, D, and J gene usage. Moreover, the presence and pattern of somatic Ig V gene mutations give valuable insight into the stage of differentiation of the B cells.

  7. Functional genomics of a generalist parasitic plant: Laser microdissection of host-parasite interface reveals host-specific patterns of parasite gene expression

    PubMed Central

    2013-01-01

    Background Orobanchaceae is the only plant family with members representing the full range of parasitic lifestyles plus a free-living lineage sister to all parasitic lineages, Lindenbergia. A generalist member of this family, and an important parasitic plant model, Triphysaria versicolor regularly feeds upon a wide range of host plants. Here, we compare de novo assembled transcriptomes generated from laser micro-dissected tissues at the host-parasite interface to uncover details of the largely uncharacterized interaction between parasitic plants and their hosts. Results The interaction of Triphysaria with the distantly related hosts Zea mays and Medicago truncatula reveals dramatic host-specific gene expression patterns. Relative to above ground tissues, gene families are disproportionally represented at the interface including enrichment for transcription factors and genes of unknown function. Quantitative Real-Time PCR of a T. versicolor β-expansin shows strong differential (120x) upregulation in response to the monocot host Z. mays; a result that is concordant with our read count estimates. Pathogenesis-related proteins, other cell wall modifying enzymes, and orthologs of genes with unknown function (annotated as such in sequenced plant genomes) are among the parasite genes highly expressed by T. versicolor at the parasite-host interface. Conclusions Laser capture microdissection makes it possible to sample the small region of cells at the epicenter of parasite host interactions. The results of our analysis suggest that T. versicolor’s generalist strategy involves a reliance on overlapping but distinct gene sets, depending upon the host plant it is parasitizing. The massive upregulation of a T. versicolor β-expansin is suggestive of a mechanism for parasite success on grass hosts. In this preliminary study of the interface transcriptomes, we have shown that T. versicolor, and the Orobanchaceae in general, provide excellent opportunities for the

  8. Molecular analysis of cell type-specific gene expression profile during mouse spermatogenesis by laser microdissection and qRT-PCR.

    PubMed

    Esakky, Prabagaran; Hansen, Deborah A; Drury, Andrea M; Moley, Kelle H

    2013-03-01

    Laser microdissection (LMD) is a selective cell isolation technique that enables the separation of desired homogenous cell subpopulations from complex tissues such as the testes under direct microscopic visualization. The LMD accompanied by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) represents an indispensable tool in quantifying messenger RNA (mRNA) expression among defined cell populations. Gene expression is temporally and spatially regulated at 3 sequential phases of mitotic, meiotic, and postmeiotic stages of spermatogenesis. The present study demonstrates a short modified LMD protocol based upon hematoxylin and eosin (H&E) staining. Stage-specific LMD success was validated by the use of mRNA profiling of "marker genes" which are conserved across species and are known to be differentially expressed during spermatogenesis. Magea4, Hspa2, Cox6b2, Tnp1, Prm1, and Prm2 are used to differentiate among the microdissected cell populations, namely spermatogonia (group I), spermatocytes (group II), round and condensing spermatids (group III), and elongated and condensed spermatids (group IV), respectively. The LMD combined with qRT-PCR is further extended to assess the cell stage-specific distribution of selected stress response genes such as Hsp90aa1, Gpx4, Ucp2, Sod1, and Sod2. The germ cell-specific mRNA profiles are suitably complemented by Western blot of the LMD samples, immunohistochemistry, and confocal localization of the corresponding proteins. The current study suggests that LMD can successfully isolate cell subpopulations from the complex tissues of the testes; and establish cell stage-specific basal expression patterns of selected stress response genes and proteins. It is our hypothesis that the baseline expression of stress response genes will differ by cell stage to create discrete stage-specific vulnerabilities to reproductive toxicants.

  9. Reproducibility of protein identification of selected cell types in Barrett's esophagus analyzed by combining laser-capture microdissection and mass spectrometry.

    PubMed

    Stingl, Christoph; van Vilsteren, Frederike G I; Guzel, Coskun; Ten Kate, Fiebo J W; Visser, Mike; Krishnadath, Kausilia K; Bergman, Jacques J; Luider, Theo M

    2011-01-01

    Barrett's esophagus (BE) is associated with increased risk of esophageal adenocarcinoma (EAC) and characterized by replacement of normal esophageal squamous epithelium by columnar epithelium. These alterations are also reflected in changes in the protein-expression profiles of the cell types involved. To separately investigate the proteomes of selected cell-types we combined laser-capture microdissection (LCM) and liquid chromatography-mass spectrometry (LC-MS). Aims were to determine the sensitivity, specificity, and technical reproducibility of the sampling method, and the biological variability within and between biopsies and patients. Frozen biopsies were cryo-sectioned, samples of around 2000 epithelial or stroma cells microdissected, digested and measured by Orbitrap LC-MS. Proteins were then identified by MS/MS database search and quantified by label-free analysis. An average of 366 protein-groups were identified per sample, and more protein-groups were found in epithelial samples than in stromal samples (442 vs 301, p < 0.0001). Altogether, 1254 distinct protein-groups were found, 289 and 88 of them significantly more often in epithelial and stroma samples, respectively. We assessed five different types of reproducibilities (run-to-run, intrabiopsy, biopsy-to-biopsy, experiment-to-experiment, and patient-to-patient) for protein identification and protein quantification. Reproducibility of protein identification ranged from 78 to 57%, and standard deviation of protein quantification was on patient-to-patient level four times higher than for run-to-run. We conclude that sampling around 2000 cells requires groups of 32 samples to detect significant, over 10-fold differences in protein abundances and thus creates a successful compromise between throughput and quality of results. We therefore believe that this method is suitable for investigating protein-expression profiles during carcinogenesis.

  10. Laser microdissection and microarray analysis of the hippocampus of Ras-GRF1 knockout mice reveals gene expression changes affecting signal transduction pathways related to memory and learning.

    PubMed

    Fernández-Medarde, A; Porteros, A; de las Rivas, J; Núñez, A; Fuster, J J; Santos, E

    2007-04-25

    We used manual macrodissection or laser capture microdissection (LCM) to isolate tissue sections of the hippocampus area of Ras-GRF1 wild type and knockout mice brains, and analyzed their transcriptional patterns using commercial oligonucleotide microarrays. Comparison between the transcriptomes of macrodissected and microdissected samples showed that the LCM samples allowed detection of significantly higher numbers of differentially expressed genes, with higher statistical rates of significance. These results validate LCM as a reliable technique for in vivo genomic studies in the brain hippocampus, where contamination by surrounding areas (not expressing Ras-GRF1) increases background noise and impairs identification of differentially expressed genes. Comparison between wild type and knockout LCM hippocampus samples revealed that Ras-GRF1 elimination caused significant gene expression changes, mostly affecting signal transduction and related neural processes. The list of 36 most differentially expressed genes included loci concerned mainly with Ras/G protein signaling and cytoskeletal organization (i.e. 14-3-3gamma/zeta, Kcnj6, Clasp2) or related, cross-talking pathways (i.e. jag2, decorin, strap). Consistent with the phenotypes shown by Ras-GRF1 knockout mice, many of these differentially expressed genes play functional roles in processes such as sensory development and function (i.e. Sptlc1, antiquitin, jag2) and/or neurological development/neurodegeneration processes affecting memory and learning. Indeed, potential links to neurodegenerative diseases such as Alzheimer disease (AD) or Creutzfeldt-Jacobs disease (CJD), have been reported for a number of differentially expressed genes identified in this study (Ptma, Aebp2, Clasp2, Hebp1, 14-3-3gamma/zeta, Csnk1delta, etc.). These data, together with the previously described role of IRS and insulin (known Ras-GRF1 activators) in AD, warrant further investigation of a potential functional link of Ras-GRF1 to

  11. UV-laser microdissection system - A novel approach for the preparation of high-resolution stable isotope records (δ13C/δ18O) from tree rings

    NASA Astrophysics Data System (ADS)

    Schollaen, Karina; Helle, Gerhard

    2013-04-01

    Intra-annual stable isotope (δ13C and δ18O) studies of tree rings at various incremental resolutions have been attempting to extract valuable seasonal climatic and environmental information or assessing plant ecophysiological processes. For preparing high-resolution isotope samples normally wood segments or cores are mechanically divided in radial direction or cut in tangential direction. After mechanical dissection, wood samples are ground to a fine powder and either cellulose is extracted or bulk wood samples are analyzed. Here, we present a novel approach for the preparation of high-resolution stable isotope records from tree rings using an UV-laser microdissection system. Firstly, tree-ring cellulose is directly extracted from wholewood cross-sections largely leaving the wood anatomical structure intact and saving time as compared to the classical procedure. Secondly, micro-samples from cellulose cross-sections are dissected with an UV-Laser dissection microscope. Tissues of interest from cellulose cross-sections are identified and marked precisely with a screen-pen and dissected via an UV-laser beam. Dissected cellulose segments were automatically collected in capsules and are prepared for stable isotope (δ13C and δ18O) analysis. The new techniques facilitate inter- and intra-annual isotope analysis on tree-ring and open various possibilities for comparisons with wood anatomy in plant eco-physiological studies. We describe the design and the handling of this novel methodology and discuss advantages and constraints given by the example of intra-annual oxygen isotope analysis on tropical trees.

  12. Western blot analysis of a limited number of cells: a valuable adjunct to proteome analysis of paraffin wax-embedded, alcohol-fixed tissue after laser capture microdissection.

    PubMed

    Martinet, Wim; Abbeloos, Vanessa; Van Acker, Nathalie; De Meyer, Guido R Y; Herman, Arnold G; Kockx, Mark M

    2004-03-01

    In recent years, laser capture microdissection (LCM) has been used successfully to obtain distinct populations of cells for subsequent molecular analysis. Because of the limited sample availability and the absence of in vitro amplification steps for proteins, the use of LCM for proteome analysis largely depends on highly sensitive protein detection methods. In this study, a western blot protocol was developed and validated for the detection of beta-actin and the moderately expressed cell death protein caspase-3 in small numbers of cells. Initially, cultured human U937 monocytes and whole sections of paraffin wax-embedded, alcohol-fixed human tonsils were used to optimize protein electrophoresis and western blotting conditions. High-performance NuPAGE Bis-Tris gels in combination with high-quality transfer membranes, optimized antibody concentrations, and a sensitive chemiluminescent substrate provided a strong signal for beta-actin with approximately 500 U937 cells. In the same way, procaspase-3 could be identified with approximately 1000 cells. Similar results were obtained with germinal centre cells that were procured from paraffin wax-embedded, alcohol-fixed human tonsils by LCM. Treatment of U937 cells with etoposide rapidly induced cell death and allowed the detection of active caspase-3 with approximately 2500 cells (0.8 pg of protein). The findings of this study suggest that western blotting is a valuable adjunct to proteome analysis of LCM procured cells.

  13. Mitochondrial DNA deletion mutations are concomitant with ragged red regions of individual, aged muscle fibers: analysis by laser-capture microdissection

    PubMed Central

    Cao, Zhengjin; Wanagat, Jonathan; McKiernan, Susan H.; Aiken, Judd M.

    2001-01-01

    Laser-capture microdissection was coupled with PCR to define the mitochondrial genotype of aged muscle fibers exhibiting mitochondrial enzymatic abnormalities. These electron transport system (ETS) abnormalities accumulate with age, are localized segmentally along muscle fibers, are associated with fiber atrophy and may contribute to age-related fiber loss. DNA extracted from single, 10 µm thick, ETS abnormal muscle fibers, as well as sections from normal fibers, served as templates for PCR-based deletion analysis. Large mitochondrial (mt) DNA deletion mutations (4.4–9.7 kb) were detected in all 29 ETS abnormal fibers analyzed. Deleted mtDNA genomes were detected only in the regions of the fibers with ETS abnormalities; adjacent phenotypically normal portions of the same fiber contained wild-type mtDNA. In addition, identical mtDNA deletion mutations were found within different sections of the same abnormal region. These findings demonstrate that large deletion mutations are associated with ETS abnormalities in aged rat muscle and that, within a fiber, deletion mutations are clonal. The displacement of wild-type mtDNAs with mutant mtDNAs results in concomitant mitochondrial enzymatic abnormalities, fiber atrophy and fiber breakage. PMID:11691938

  14. A pilot study on the expression of microRNAs resident on chromosome 21 in laser microdissected FFPE prostate adenocarcinoma samples.

    PubMed

    Mihala, Adrian; Alexa, Andreea Anda; Samoilă, Corina; Dema, Alis; Vizitiu, Anda Cornelia; Anghel, Andrei; Tămaş, Liviu; Marian, Cătălin Valer; Sîrbu, Ioan Ovidiu

    2015-01-01

    The tremendous research effort of the last decades added a new, epigenetic layer of complexity to the already complex image of prostate cancer pathogenesis. Here we use quantitative real-time polymerase chain reaction (qRT-PCR) to investigate the expression of the microRNAs resident on chromosome 21 (miR-ch21) in laser capture microdissected (LCM) tissues from formalin-fixed paraffin-embedded (FFPE) archived, prostate adenocarcinoma samples. We show a strong, specific down-regulation of miR-ch21 in tumoral epithelia and stromae as compared to normal counterparts, results at odd with the current paradigm on the involvement of these microRNAs in prostate oncogenesis. By comparing this result with the expression of two well-known pluripotency associated microRNA, hsa-miR-372 and miR-373, we suggest that miR-ch21 down-regulation might be the result of specific silencing of miR genes mapped to chromosome 21. Further studies, of larger sample size are needed to confirm our preliminary data.

  15. Collection of Epithelial Cells from Rodent Mammary Gland Via Laser Capture Microdissection Yielding High-Quality RNA Suitable for Microarray Analysis

    PubMed Central

    2010-01-01

    Laser capture microdissection (LCM) enables collection of cell populations highly enriched for specific cell types that have the potential of yielding critical information about physiological and pathophysiological processes. One use of cells collected by LCM is for gene expression profiling. Samples intended for transcript analyses should be of the highest quality possible. RNA degradation is an ever-present concern in molecular biological assays, and LCM is no exception. This paper identifies issues related to preparation, collection, and processing in a lipid-rich tissue, rodent mammary gland, in which the epithelial to stromal cell ratio is low and the stromal component is primarily adipocytes, a situation that presents numerous technical challenges for high-quality RNA isolation. Our goal was to improve the procedure so that a greater probe set present call rate would be obtained when isolated RNA was evaluated using Affymetrix microarrays. The results showed that the quality of RNA isolated from epithelial cells of both mammary gland and mammary adenocarcinomas was high with a probe set present call rate of 65% and a high signal-to-noise ratio. PMID:21406068

  16. Olfactory Neurons Obtained through Nasal Biopsy Combined with Laser-Capture Microdissection: A Potential Approach to Study Treatment Response in Mental Disorders

    PubMed Central

    Narayan, Soumya; McLean, Charlee; Sawa, Akira; Lin, Sandra Y.; Rai, Narayan; Hipolito, MariaMananita S.; Cascella, Nicola; Nurnberger, John J.I.; Koko, Ishizuka; Nwulia, Evaristus A.

    2015-01-01

    Bipolar disorder (BD) is a severe neuropsychiatric disorder with poorly understood pathophysiology and typically treated with the mood stabilizer, lithium carbonate. Animal studies as well as human genetic studies indicate that lithium affects molecular targets that are involved in neuronal growth, survival and maturation, and notably molecules involved in Wnt signaling. Given the ethical challenge to obtaining brain biopsies for investigating dynamic molecular changes associated with lithium-response in the central nervous system (CNS), one may consider the use of neurons obtained from olfactory tissues to achieve this goal.The olfactory epithelium contains olfactory receptor neurons at different stages of development and glial-like supporting cells. This provides a unique opportunity to study dynamic changes in the CNS of patients with neuropsychiatric diseases, using olfactory tissue safely obtained from nasal biopsies. To overcome the drawback posed by substantial contamination of biopsied olfactory tissue with non-neuronal cells, a novel approach to obtain enriched neuronal cell populations was developed by combining nasal biopsies with laser-capture microdissection. In this study, a system for investigating treatment-associated dynamic molecular changes in neuronal tissue was developed and validated, using a small pilot sample of BD patients recruited for the study of the molecular mechanisms of lithium treatment response. PMID:25549156

  17. Detection of CCND1 amplification using laser capture microdissection coupled with real-time polymerase chain reaction in human esophageal squamous cell carcinoma.

    PubMed

    Huang, Chen; Yang, Lin; Li, Zhongfang; Yang, Jun; Zhao, Junjie; Dehui, Xu; Liu, Liying; Wang, Quanli; Song, Tusheng

    2007-05-01

    Several methods have been used to detect CCND1 amplification or overexpression in esophageal squamous cell carcinoma (ESCC), but problems remain, associated with heterogeneity of tumor tissue and quantification of gene copies. Laser capture microdissection coupled with real-time polymerase chain reaction (PCR) is a reliable method for the molecular analysis of gene profiles in specific tissues. All 35 specimens of ESCC studied were paraffin-embedded, cut into tissue slides, and stained by hematoxylin-eosin. The pure ESCC cell and normal squamous epithelia populations were separated by LCM and then genomic DNA was extracted from the dissected cells. CCND1 amplification was detected with real-time FQ-PCR and with PCR. Amplification was calculated by the formula X = 2(-DeltaDeltaCt) and R = (CCND1/ACTB) CANCER/(CCND1/ACTB) NORMAL. Twenty (57%) of primary ESCC cancer cell groups had a detectable CCND1 amplification (range, 2.06-fold to 25.9-fold) with real-time FQ-PCR, but only 2 of 15 primary ESCC cancer cell groups had detectable CCND1 amplification by PCR. CCND1 amplification was not correlated with age, sex, size of tumor, histological grade, and lymph node metastasis. In conclusion, LCM coupled with real-time fluorescence quantitative-PCR technique is more precise than PCR for the identifying amplified oncogenes; The role of CCND1 amplification in ESCC development and progression needs more extensive study.

  18. Adaptation of Laser Microdissection Technique for the Study of a Spontaneous Metastatic Mammary Carcinoma Mouse Model by NanoString Technologies.

    PubMed

    Castro, Nadia P; Merchant, Anand S; Saylor, Karen L; Anver, Miriam R; Salomon, David S; Golubeva, Yelena G

    2016-01-01

    Laser capture microdissection (LCM) of tissue is an established tool in medical research for collection of distinguished cell populations under direct microscopic visualization for molecular analysis. LCM samples have been successfully analyzed in a number of genomic and proteomic downstream molecular applications. However, LCM sample collection and preparation procedure has to be adapted to each downstream analysis platform. In this present manuscript we describe in detail the adaptation of LCM methodology for the collection and preparation of fresh frozen samples for NanoString analysis based on a study of a model of mouse mammary gland carcinoma and its lung metastasis. Our adaptation of LCM sample preparation and workflow to the requirements of the NanoString platform allowed acquiring samples with high RNA quality. The NanoString analysis of such samples provided sensitive detection of genes of interest and their associated molecular pathways. NanoString is a reliable gene expression analysis platform that can be effectively coupled with LCM. PMID:27077656

  19. Improved method for extraction and detection of Helicobacter pylori DNA in formalin-fixed paraffin embedded gastric biopsies using laser micro-dissection

    PubMed Central

    Loayza, María Fernanda; Villavicencio, Fernando Xavier; Santander, Stephanie Carolina; Baldeón, Manuel; Ponce, Lourdes Karina; Salvador, Iván; Vivar Díaz, Nicolás

    2014-01-01

    To assess the molecular events exerted by Helicobacter pylori interacting directly with gastric epithelial cells, an improved procedure for microbial DNA isolation from stained hematoxilin-eosin gastric biopsies was developed based on laser micro-dissection (LM) [1]. Few articles have described the use of LM to select and detect H. pylori genome from formalin-fixed paraffin embedded gastric tissue [2]. To improve the yield and quality of DNA isolated from H. pylori contacting intestinal epithelial cells, the following conditions were established after modification of the QIAamp DNA Micro kit. • Use of at least 25 cut sections of 10–20 μm of diameter and 3 μm thick with more than 10 bacteria in each cut. • Lysis with 30 μL of tissue lysis buffer and 20 μL of proteinase K (PK) with the tube in an upside-down position. • The use of thin purification columns with 35 μL of elution buffer. The mean of DNA concentration obtained from 25 LM cut sections was 1.94± 0 .16 ng/μL, and it was efficiently amplified with qPCR in a Bio Rad iCycler instrument. The LM can improve the sample selection and DNA extraction for molecular analysis of H. pylori associated with human gastric epithelium. PMID:26150965

  20. Improved method for extraction and detection of Helicobacter pylori DNA in formalin-fixed paraffin embedded gastric biopsies using laser micro-dissection.

    PubMed

    Loayza, María Fernanda; Villavicencio, Fernando Xavier; Santander, Stephanie Carolina; Baldeón, Manuel; Ponce, Lourdes Karina; Salvador, Iván; Vivar Díaz, Nicolás

    2015-01-01

    To assess the molecular events exerted by Helicobacter pylori interacting directly with gastric epithelial cells, an improved procedure for microbial DNA isolation from stained hematoxilin-eosin gastric biopsies was developed based on laser micro-dissection (LM) [1]. Few articles have described the use of LM to select and detect H. pylori genome from formalin-fixed paraffin embedded gastric tissue [2]. To improve the yield and quality of DNA isolated from H. pylori contacting intestinal epithelial cells, the following conditions were established after modification of the QIAamp DNA Micro kit. •Use of at least 25 cut sections of 10-20 μm of diameter and 3 μm thick with more than 10 bacteria in each cut.•Lysis with 30 μL of tissue lysis buffer and 20 μL of proteinase K (PK) with the tube in an upside-down position.•The use of thin purification columns with 35 μL of elution buffer. The mean of DNA concentration obtained from 25 LM cut sections was 1.94± 0 .16 ng/μL, and it was efficiently amplified with qPCR in a Bio Rad iCycler instrument. The LM can improve the sample selection and DNA extraction for molecular analysis of H. pylori associated with human gastric epithelium.

  1. Laser Capture Microdissection of Cervical Human Papillomavirus Infections: Copy Number of the Virus in Cancerous and Normal Tissue and Heterogeneous DNA Methylation

    PubMed Central

    Kalantari, Mina; Garcia-Carranca, Alejandro; Morales-Vazquez, Claudia Dalia; Zuna, Rosemary; Montiel, Delia Perez; Calleja-Macias, Itzel E.; Johansson, Bo; Andersson, Sonia; Bernard, Hans-Ulrich

    2009-01-01

    Research on the pathogenicity of human papillomaviruses (HPVs) during cervical carcinogenesis often relies on the study of homogenized tissue or cultured cells. This approach does not detect molecular heterogeneities within the infected tissue. It is desirable to understand molecular properties in specific histological contexts. We asked whether Laser Capture Microdissection (LCM) of archival cervical tumors in combination with real-time polymerase chain reaction and bisulfite sequencing permits (i) sensitive DNA diagnosis of small clusters of formalin fixed cells, (ii) quantification of HPV DNA in neoplastic and normal cells, and (iii) analysis of HPV DNA methylation, a marker of tumor progression. We analyzed 26 tumors containing HPV-16 or 18. We prepared DNA from LCM dissected thin sections of 100 to 2000 cells, and analyzed aliquots corresponding to between nine and 70 cells. We detected nine to 630 HPV-16 genome copies and one to 111 HPV-18 genome copies per tumor cell, respectively. In 17 of the 26 samples, HPV DNA existed in histologically normal cells distant from the margins of the tumors, but at much lower concentrations than in the tumor, suggesting that HPVs can infect at low levels without pathogenic changes. Methylation of HPV DNA, a biomarker of integration of the virus into cellular DNA, could be measured only in few samples due to limited sensitivity, and indicated heterogeneous methylation patterns in small clusters of cancerous and normal cells. LCM is powerful to study molecular parameters of cervical HPV infections like copy number, latency and epigenetics. PMID:19497607

  2. Adaptation of Laser Microdissection Technique for the Study of a Spontaneous Metastatic Mammary Carcinoma Mouse Model by NanoString Technologies

    PubMed Central

    Saylor, Karen L.; Anver, Miriam R.; Salomon, David S.; Golubeva, Yelena G.

    2016-01-01

    Laser capture microdissection (LCM) of tissue is an established tool in medical research for collection of distinguished cell populations under direct microscopic visualization for molecular analysis. LCM samples have been successfully analyzed in a number of genomic and proteomic downstream molecular applications. However, LCM sample collection and preparation procedure has to be adapted to each downstream analysis platform. In this present manuscript we describe in detail the adaptation of LCM methodology for the collection and preparation of fresh frozen samples for NanoString analysis based on a study of a model of mouse mammary gland carcinoma and its lung metastasis. Our adaptation of LCM sample preparation and workflow to the requirements of the NanoString platform allowed acquiring samples with high RNA quality. The NanoString analysis of such samples provided sensitive detection of genes of interest and their associated molecular pathways. NanoString is a reliable gene expression analysis platform that can be effectively coupled with LCM. PMID:27077656

  3. Laser-capture microdissection of oropharyngeal epithelium indicates restriction of Epstein-Barr virus receptor/CD21 mRNA to tonsil epithelial cells

    PubMed Central

    Jiang, Ru; Gu, Xin; Nathan, Cherie-Ann; Hutt-Fletcher, Lindsey

    2008-01-01

    Background: Epstein-Barr virus colonizes the oropharynx of a majority of individuals. It infects B lymphocytes and epithelial cells and can contribute to the development of both lymphoid and epithelial tumors. The virus uses CD21 for attachment to B cells which constitutively express the protein. Infection of epithelial cells in vitro is also more efficient if CD21 is available. However, its potential contribution to infection in vivo has been difficult to evaluate as discrepant results with antibodies have made it difficult to determine which, if any, epithelial cells in the oropharynx express CD21. Methods: To reevaluate CD21 expression by an alternative method, epithelial cells were isolated by laser-capture microdissection from formalin-fixed sections of tissues from various parts of the oropharynx and mRNA was amplified with primers specific for the exons of CD21 which code for the Epstein-Barr virus binding site. Results: CD21 mRNA was expressed in tonsil epithelium, but not in epithelium from buccal mucosa, uvula, soft palate or tongue. Conclusions: CD21 does not contribute to infection of most normal epithelial tissues in the oropharynx, but may contribute to infection of epithelial cells in the tonsil, where virus has been demonstrated in healthy carriers. PMID:18710421

  4. Expression of Intermediate Filaments and Germ Cell Markers in the Developing Bovine Ovary: An Immunohistochemical and Laser-Assisted Microdissection Study.

    PubMed

    Kenngott, Rebecca Anna-Maria; Sauer, Ulrich; Vermehren, Margarete; Sinowatz, Fred

    2014-01-01

    In the present investigation, bovine ovary prenatal development was studied using immunohistochemistry and laser-assisted microdissection (LAM). A major aim of this study was to evaluate the protein expression pattern of intermediate filaments (IF) and distinguish S100 protein (S100 alpha and S100 beta protein) isoforms during prenatal follicle differentiation, subsequently correlating them with germ cell marker expression. A development-specific expression pattern of different keratins as well as vimentin was detected in the prenatal bovine ovary; K18-specific expression was found during all developmental stages (i.e. in surface epithelium, germ cell cord somatic cells, and follicle cells), and keratins 5, 7, 8, 14, and 19 and vimentin had a stage-specific expression pattern in the different cell populations of the prenatal ovaries. Additionally, our results represent new data on the expression pattern of germ cell markers during bovine ovary prenatal development. S100 alpha and beta protein was localized to oocyte cytoplasm of different follicle stages, and S100 alpha staining could be observed in granulosa cells. Furthermore, through isolation of characteristic ovary cell populations using LAM, specific confirmation of some genes of interest (KRT8, KRT18, S100 alpha, S100 beta, and OCT4, DDX4) could be obtained by RT-PCR in single cell groups of the developing bovine ovary.

  5. Laser microdissection coupled to transcriptional profiling of Arabidopsis roots inoculated by Plasmodiophora brassicae indicates a role for brassinosteroids in clubroot formation.

    PubMed

    Schuller, Astrid; Kehr, Julia; Ludwig-Müller, Jutta

    2014-02-01

    The clubroot disease caused by the obligate biotrophic protist Plasmodiophora brassicae on host plants of the Brassicaceae family is characterized by enhanced cell division and cell expansion. Since a typical root section of an infected plant always includes different stages of the pathogen as well as uninfected cells, we were interested in investigating specific developmental stages of the pathogen and their effect on host transcriptional changes. We extended previous microarray studies on whole roots by using laser microdissection and pressure catapulting (LMPC) to isolate individual cells harboring defined developmental stages of the pathogen. In addition, we compared the central cylinder of infected plants with that of control plants. We were especially interested in elucidating the stage-specific hormonal network. The up-regulation of genes involved in auxin and cytokinin metabolism and signaling was confirmed. In addition, we found evidence that brassinosteroid (BR) synthesis and signal perception genes were in many cases up-regulated in enlarged cells and the central cylinder. This was confirmed by quantitative PCR. Treatment of wild-type plants with the BR biosynthesis inhibitor propiconazole reduced gall formation, and the analysis of the BR receptor mutant bri1-6 revealed less severe gall formation than in the respective wild type. Our results identify novel hormone pathways involved in clubroot development. Using LMPC to generate pools of homogeneous cell type populations combined with transcriptome analysis has been very useful to elucidate the regulation of gall growth by this obligate biotropic pathogen in a cell- and stage-specific manner.

  6. Copper-bromide laser for treating vascular lesions

    NASA Astrophysics Data System (ADS)

    Kim, Ryun Kyung; Lee, TaeBum; Lee, SunWoo; Kim, JungSoo; Eun, HyeJun; Huh, Young

    2014-03-01

    A stable and effective CuBr laser is demonstrated for the treatment of vascular lesions. The two lasing wavelengths of 511 nm and 578 nm can be adjusted to any mixing ratio by using a 6-hole filter. The output power is stabilized to maintain the same peak power by using a proportional integral differential (PID) modulator. For various treatments and treatment effects, the laser outputs, such as the output power, spot size, and beam mode can be controlled. Preclinical test using Korean rabbits showed that the fabricated CuBr laser had a positive effect when treating vascular lesions. The blood vessels of the rabbit's ear were well ablated after irradiation at a laser output power of 8 W in a 1 mm spot in the top-hat mode.

  7. Analysis of Genome-Wide Monoallelic Expression Patterns in Three Major Cell Types of Mouse Visual Cortex Using Laser Capture Microdissection

    PubMed Central

    Gau, Susan Shur-Fen

    2016-01-01

    Genomic imprinting is an epigenetic mechanism causing monoallelic expression in a parent-of-origin-specific manner. Disruption of imprinted genes causes various neurological and psychiatric disorders. However, the role of imprinted genes in the brain is largely unknown. Different cell types within distinct brain regions can influence the genomic imprinting status, but imprinted genes in single cell types within distinct brain regions have not been characterized on a genome-wide scale. To address this critical question, we used a multi-stage approach, which combined genetically engineered mice with fluorescence-based laser capture microdissection (LCM) to capture excitatory neurons, inhibitory neurons and astrocytes as single cells in layer 2/3 of mouse visual cortex. RNA sequencing determined parental expression patterns on a genome-wide scale in the captured cells within specific brain regions. The expression level of cell-type-specific genes for excitatory neurons (13 genes), inhibitory neurons (16 genes) and astrocytes (20 genes) confirmed the LCM-captured cells maintained their cellular identities. The parent-of-origin-specific expression pattern of imprinted genes, including maternally expressed Meg3 and paternally expressed Peg3, provided evidence that the status of known imprinted genes was also maintained. Although our platform remains to be improved, our findings demonstrate the parental expression pattern can be analysed not only at the level of a single cell type but also at the level of specific cortical layers. Our approach has the potential to reveal novel regulatory modules associated with plasticity through genomic imprinting mechanisms in different cell types, not only in the visual cortex but also in other brain regions. PMID:27662371

  8. Gene expression profiling of the hyperplastic growth zones of the late trout embryo myotome using laser capture microdissection and microarray analysis

    PubMed Central

    2013-01-01

    Background A unique feature of fish is that new muscle fibres continue to be produced throughout much of the life cycle; a process termed muscle hyperplasia. In trout, this process begins in the late embryo stage and occurs in both a discrete, continuous layer at the surface of the primary myotome (stratified hyperplasia) and between existing muscle fibres throughout the myotome (mosaic hyperplasia). In post-larval stages, muscle hyperplasia is only of the mosaic type and persists until 40% of the maximum body length is reached. To characterise the genetic basis of myotube neoformation in trout, we combined laser capture microdissection and microarray analysis to compare the transcriptome of hyperplastic regions of the late embryo myotome with that of adult myotomal muscle, which displays only limited hyperplasia. Results Gene expression was analysed using Agilent trout oligo microarrays. Our analysis identified more than 6800 transcripts that were significantly up-regulated in the superficial hyperplastic zones of the late embryonic myotome compared to adult myotomal muscle. In addition to Pax3, Pax7 and the fundamental myogenic basic helix-loop-helix regulators, we identified a large set of up-regulated transcriptional factors, including Myc paralogs, members of Hes family and many homeobox-containing transcriptional regulators. Other cell-autonomous regulators overexpressed in hyperplastic zones included a large set of cell surface proteins belonging to the Ig superfamily. Among the secreted molecules found to be overexpressed in hyperplastic areas, we noted growth factors as well as signalling molecules. A novel finding in our study is that many genes that regulate planar cell polarity (PCP) were overexpressed in superficial hyperplastic zones, suggesting that the PCP pathway is involved in the oriented elongation of the neofibres. Conclusion The results obtained in this study provide a valuable resource for further analysis of novel genes potentially involved

  9. Laser Microdissection Unravels Cell-Type-Specific Transcription in Arbuscular Mycorrhizal Roots, Including CAAT-Box Transcription Factor Gene Expression Correlating with Fungal Contact and Spread1[W

    PubMed Central

    Hogekamp, Claudia; Arndt, Damaris; Pereira, Patrícia A.; Becker, Jörg D.; Hohnjec, Natalija; Küster, Helge

    2011-01-01

    Arbuscular mycorrhizae (AM) are the most widespread symbioses on Earth, promoting nutrient supply of most terrestrial plant species. To unravel gene expression in defined stages of Medicago truncatula root colonization by AM fungi, we here combined genome-wide transcriptome profiling based on whole mycorrhizal roots with real-time reverse transcription-PCR experiments that relied on characteristic cell types obtained via laser microdissection. Our genome-wide approach delivered a core set of 512 genes significantly activated by the two mycorrhizal fungi Glomus intraradices and Glomus mossae. Focusing on 62 of these genes being related to membrane transport, signaling, and transcriptional regulation, we distinguished whether they are activated in arbuscule-containing or the neighboring cortical cells harboring fungal hyphae. In addition, cortical cells from nonmycorrhizal roots served as a reference for gene expression under noncolonized conditions. Our analysis identified 25 novel arbuscule-specific genes and 37 genes expressed both in the arbuscule-containing and the adjacent cortical cells colonized by fungal hyphae. Among the AM-induced genes specifying transcriptional regulators were two members encoding CAAT-box binding transcription factors (CBFs), designated MtCbf1 and MtCbf2. Promoter analyses demonstrated that both genes were already activated by the first physical contact between the symbionts. Subsequently, and corresponding to our cell-type expression patterns, they were progressively up-regulated in those cortical areas colonized by fungal hyphae, including the arbuscule-containing cells. The encoded CBFs thus represent excellent candidates for regulators that mediate a sequential reprogramming of root tissues during the establishment of an AM symbiosis. PMID:22034628

  10. Comprehensive network analysis of anther-expressed genes in rice by the combination of 33 laser microdissection and 143 spatiotemporal microarrays.

    PubMed

    Aya, Koichiro; Suzuki, Go; Suwabe, Keita; Hobo, Tokunori; Takahashi, Hirokazu; Shiono, Katsuhiro; Yano, Kentaro; Tsutsumi, Nobuhiro; Nakazono, Mikio; Nagamura, Yoshiaki; Matsuoka, Makoto; Watanabe, Masao

    2011-01-01

    Co-expression networks systematically constructed from large-scale transcriptome data reflect the interactions and functions of genes with similar expression patterns and are a powerful tool for the comprehensive understanding of biological events and mining of novel genes. In Arabidopsis (a model dicot plant), high-resolution co-expression networks have been constructed from very large microarray datasets and these are publicly available as online information resources. However, the available transcriptome data of rice (a model monocot plant) have been limited so far, making it difficult for rice researchers to achieve reliable co-expression analysis. In this study, we performed co-expression network analysis by using combined 44 K agilent microarray datasets of rice, which consisted of 33 laser microdissection (LM)-microarray datasets of anthers, and 143 spatiotemporal transcriptome datasets deposited in RicexPro. The entire data of the rice co-expression network, which was generated from the 176 microarray datasets by the Pearson correlation coefficient (PCC) method with the mutual rank (MR)-based cut-off, contained 24,258 genes and 60,441 genes pairs. Using these datasets, we constructed high-resolution co-expression subnetworks of two specific biological events in the anther, "meiosis" and "pollen wall synthesis". The meiosis network contained many known or putative meiotic genes, including genes related to meiosis initiation and recombination. In the pollen wall synthesis network, several candidate genes involved in the sporopollenin biosynthesis pathway were efficiently identified. Hence, these two subnetworks are important demonstrations of the efficiency of co-expression network analysis in rice. Our co-expression analysis included the separated transcriptomes of pollen and tapetum cells in the anther, which are able to provide precise information on transcriptional regulation during male gametophyte development in rice. The co-expression network data

  11. Vascular tissue engineering by computer-aided laser micromachining.

    PubMed

    Doraiswamy, Anand; Narayan, Roger J

    2010-04-28

    Many conventional technologies for fabricating tissue engineering scaffolds are not suitable for fabricating scaffolds with patient-specific attributes. For example, many conventional technologies for fabricating tissue engineering scaffolds do not provide control over overall scaffold geometry or over cell position within the scaffold. In this study, the use of computer-aided laser micromachining to create scaffolds for vascular tissue networks was investigated. Computer-aided laser micromachining was used to construct patterned surfaces in agarose or in silicon, which were used for differential adherence and growth of cells into vascular tissue networks. Concentric three-ring structures were fabricated on agarose hydrogel substrates, in which the inner ring contained human aortic endothelial cells, the middle ring contained HA587 human elastin and the outer ring contained human aortic vascular smooth muscle cells. Basement membrane matrix containing vascular endothelial growth factor and heparin was to promote proliferation of human aortic endothelial cells within the vascular tissue networks. Computer-aided laser micromachining provides a unique approach to fabricate small-diameter blood vessels for bypass surgery as well as other artificial tissues with complex geometries.

  12. Vascular tissue engineering by computer-aided laser micromachining.

    PubMed

    Doraiswamy, Anand; Narayan, Roger J

    2010-04-28

    Many conventional technologies for fabricating tissue engineering scaffolds are not suitable for fabricating scaffolds with patient-specific attributes. For example, many conventional technologies for fabricating tissue engineering scaffolds do not provide control over overall scaffold geometry or over cell position within the scaffold. In this study, the use of computer-aided laser micromachining to create scaffolds for vascular tissue networks was investigated. Computer-aided laser micromachining was used to construct patterned surfaces in agarose or in silicon, which were used for differential adherence and growth of cells into vascular tissue networks. Concentric three-ring structures were fabricated on agarose hydrogel substrates, in which the inner ring contained human aortic endothelial cells, the middle ring contained HA587 human elastin and the outer ring contained human aortic vascular smooth muscle cells. Basement membrane matrix containing vascular endothelial growth factor and heparin was to promote proliferation of human aortic endothelial cells within the vascular tissue networks. Computer-aided laser micromachining provides a unique approach to fabricate small-diameter blood vessels for bypass surgery as well as other artificial tissues with complex geometries. PMID:20308108

  13. Vascular Welding Using The Argon Laser

    NASA Astrophysics Data System (ADS)

    White, Rodney A.; Donayre, Carlos; Kopchok, George; White, Geoffrey; Abergel, R. Patrick; Lyons, Richard; Klein, Stanley; Dwyer, Richard; Uitto, Jouni

    1987-03-01

    This study compared the histology, biochemistry, and tensile strength of laser welded and sutured canine venotomies, arteriotomies and arteriovenous fistulas. Bilateral femoral, carotid or jugular vessels were studied with one repair (control) closed with interrupted 6-0 polypropylene sutures, and the contralatral repair (experimental) welded with the argon laser. Specimens were examined at weekly intervals from 1 to 4 weeks for each type of repair and evaluated histologically by hematoxylineosin, elastin and trichrome stains, biochemically by the formation of [3H] hyaroxyproline as an index of collagen synthesis, ana mechanically by tensile strength determinations. At removal, all experimental closures were patent without hematomas, aneurysms or luminal dilatation. Histologic and biochemical examination and tensile strength determinations suggest that laser welaing may be an alternative to sutures for repair of large diameter venotomies, arteriotomies and arteriovenous fistulas, as they heal comparable to suture repairs up to 4 weeks postoperatively.

  14. A roadmap for zinc trafficking in the developing barley grain based on laser capture microdissection and gene expression profiling

    PubMed Central

    Tauris, Birgitte; Borg, Søren; Gregersen, Per L.; Holm, Preben B.

    2009-01-01

    Nutrients destined for the developing cereal grain encounter several restricting barriers on their path towards their final storage sites in the grain. In order to identify transporters and chelating agents that may be involved in transport and deposition of zinc in the barley grain, expression profiles have been generated of four different tissue types: the transfer cells, the aleurone layer, the endosperm, and the embryo. Cells from these tissues were isolated with the ‘laser capture microdissection’ technology and the extracted RNA was subjected to three rounds of T7-based amplification. The amplified RNA was subsequently hybridized to Affymetrix 22K Barley GeneChips. Due to the short average length of the amplified transcripts and the positioning of numerous probe sets at locations more than 400 base pairs (bp) from the poly(A)-tail, a normalization approach was used where the probe positions were taken into account. On the basis of the expression levels of a number of metal homeostasis genes, a working model is proposed for the translocation of zinc from the phloem to the storage sites in the developing grain. PMID:19297552

  15. Comedones Induced by Vascular Laser Therapy

    PubMed Central

    Demirci, Gulsen Tukenmez; Mansur, Ayse Tulin; Gulec, Ayse Tulin

    2016-01-01

    A 21-year-old female presented with acne-like blackheads on brownish areas located on the cheek. She had been treated with neodymium-doped yttrium aluminium garnet (Nd-YAG) laser (1071 nm), 160 j/cm2, three months ago for erythema and telangiectasia of her face. Afterwards, she developed atrophic, slightly depressed, hyperpigmented, 3-4 mm scars with superimposed tiny comedones within the treated areas. Topical treatment with tretinoin 0.05% cream on alternate days, and Sun Protection Factor (SPF) 50 sunscreen daily were commenced. After 2 months, comedones and hyperpigmentation mostly resolved but mild superficial atrophy persisted. According to our knowledge, this is the first case of atrophic scars studded with open comedones, developing shortly after laser therapy used for facial telangiectasia. PMID:27081249

  16. Comedones Induced by Vascular Laser Therapy.

    PubMed

    Demirci, Gulsen Tukenmez; Mansur, Ayse Tulin; Gulec, Ayse Tulin

    2016-01-01

    A 21-year-old female presented with acne-like blackheads on brownish areas located on the cheek. She had been treated with neodymium-doped yttrium aluminium garnet (Nd-YAG) laser (1071 nm), 160 j/cm(2), three months ago for erythema and telangiectasia of her face. Afterwards, she developed atrophic, slightly depressed, hyperpigmented, 3-4 mm scars with superimposed tiny comedones within the treated areas. Topical treatment with tretinoin 0.05% cream on alternate days, and Sun Protection Factor (SPF) 50 sunscreen daily were commenced. After 2 months, comedones and hyperpigmentation mostly resolved but mild superficial atrophy persisted. According to our knowledge, this is the first case of atrophic scars studded with open comedones, developing shortly after laser therapy used for facial telangiectasia. PMID:27081249

  17. Laser capture microdissection as a tool to evaluate human papillomavirus genotyping and methylation as biomarkers of persistence and progression of anal lesions

    PubMed Central

    Cornall, Alyssa M; Roberts, Jennifer M; Molano, Monica; Machalek, Dorothy A; Phillips, Samuel; Hillman, Richard J; Grulich, Andrew E; Jin, Fengyi; Poynten, I Mary; Templeton, David J; Garland, Suzanne M; Tabrizi, Sepehr N

    2015-01-01

    Introduction Anal squamous cell carcinoma is preceded by persistent infection with high-risk human papillomavirus (HPV) and the cancer precursor, high-grade squamous intraepithelial lesion (HSIL). Detection of specific HPV genotypes and HPV-related biomarkers may be an option for primary anal screening. However, more data on the natural history of HPV-related anal lesions are required. The outcomes from this study will enhance our understanding of the clinical and biological behaviour of HPV-related anal lesions and inform the development of future HPV genotype and/or biomarker screening tests. Methods and analysis HIV-negative and HIV-positive men who have sex with men, aged 35 years and over, recruited from community-based settings in Sydney, Australia, attend 6 clinic visits over 3 years. At the first 5 visits, participants undergo a digital anorectal examination, an anal swab for HPV genotyping and anal cytology, and high-resolution anoscopy with directed biopsy of any visible abnormalities that are suggestive of any abnormality suspicious of SIL. Tissue sections from participants diagnosed with histologically confirmed HSIL at the baseline clinic visit will undergo laser capture microdissection, HPV detection and genotyping, and quantitation of CpG methylation in baseline and follow-up biopsies. Histological and cytological findings in combination with HPV genotyping data will be used to identify persistent HSIL. HSIL will be stratified as non-persistent and persistent based on their status at 12 months. The performance of HPV genotype and methylation status in predicting disease persistence at 12 months will be assessed, along with associations with HIV status and other covariates such as age. Ethics and dissemination The St Vincent's Hospital Ethics Committee granted ethics approval for the study. Written informed consent is obtained from all individuals before any study-specific procedures are performed. Findings from this study will be disseminated

  18. Comparative transcriptional survey between laser-microdissected cells from laminar abscission zone and petiolar cortical tissue during ethylene-promoted abscission in citrus leaves

    PubMed Central

    Agustí, Javier; Merelo, Paz; Cercós, Manuel; Tadeo, Francisco R; Talón, Manuel

    2009-01-01

    Background Abscission is the cell separation process by which plants are able to shed organs. It has a great impact on the yield of most crop plants. At the same time, the process itself also constitutes an excellent model to study cell separation processes, since it occurs in concrete areas known as abscission zones (AZs) which are composed of a specific cell type. However, molecular approaches are generally hampered by the limited area and cell number constituting the AZ. Therefore, detailed studies at the resolution of cell type are of great relevance in order to accurately describe the process and to identify potential candidate genes for biotechnological applications. Results Efficient protocols for the isolation of specific citrus cell types, namely laminar abscission zone (LAZ) and petiolar cortical (Pet) cells based on laser capture microdissection (LCM) and for RNA microextraction and amplification have been developed. A comparative transcriptome analysis between LAZ and Pet from citrus leaf explants subjected to an in-vitro 24 h ethylene treatment was performed utilising microarray hybridization and analysis. Our analyses of gene functional classes differentially represented in ethylene-treated LAZ revealed an activation program dominated by the expression of genes associated with protein synthesis, protein fate, cell type differentiation, development and transcription. The extensive repertoire of genes associated with cell wall biosynthesis and metabolism strongly suggests that LAZ layers activate both catabolic and anabolic wall modification pathways during the abscission program. In addition, over-representation of particular members of different transcription factor families suggests important roles for these genes in the differentiation of the effective cell separation layer within the many layers contained in the citrus LAZ. Preferential expression of stress-related and defensive genes in Pet reveals that this tissue is likely to be reprogrammed to

  19. In Situ Staining and Laser Capture Microdissection of Lymph Node Residing SIV Gag-Specific CD8+ T cells--A Tool to Interrogate a Functional Immune Response Ex Vivo.

    PubMed

    Tjernlund, Annelie; Burgener, Adam; Lindvall, Jessica M; Peng, Tao; Zhu, Jia; Öhrmalm, Lars; Picker, Louis J; Broliden, Kristina; McElrath, M Juliana; Corey, Lawrence

    2016-01-01

    While a plethora of data describes the essential role of systemic CD8+ T cells in the control of SIV replication little is known about the local in situ CD8+ T cell immune responses against SIV at the intact tissue level, due to technical limitations. In situ staining, using GagCM9 Qdot 655 multimers, were here combined with laser capture microdissection to detect and collect SIV Gag CM9 specific CD8+ T cells in lymph node tissue from SIV infected rhesus macaques. CD8+ T cells from SIV infected and uninfected rhesus macaques were also collected and compared to the SIV GagCM9 specific CD8+ T cells. Illumina bead array and transcriptional analyses were used to assess the transcriptional profiles and the three different CD8+ T cell populations displayed unique transcriptional patterns. This pilot study demonstrates that rapid and specific immunostaining combined with laser capture microdissection in concert with transcriptional profiling may be used to elucidate phenotypic differences between CD8+ T cells in SIV infection. Such technologies may be useful to determine differences in functional activities of HIV/SIV specific T cells.

  20. In Situ Staining and Laser Capture Microdissection of Lymph Node Residing SIV Gag-Specific CD8+ T cells—A Tool to Interrogate a Functional Immune Response Ex Vivo

    PubMed Central

    Tjernlund, Annelie; Burgener, Adam; Lindvall, Jessica M.; Peng, Tao; Zhu, Jia; Öhrmalm, Lars; Picker, Louis J.; Broliden, Kristina; McElrath, M. Juliana; Corey, Lawrence

    2016-01-01

    While a plethora of data describes the essential role of systemic CD8+ T cells in the control of SIV replication little is known about the local in situ CD8+ T cell immune responses against SIV at the intact tissue level, due to technical limitations. In situ staining, using GagCM9 Qdot 655 multimers, were here combined with laser capture microdissection to detect and collect SIV Gag CM9 specific CD8+ T cells in lymph node tissue from SIV infected rhesus macaques. CD8+ T cells from SIV infected and uninfected rhesus macaques were also collected and compared to the SIV GagCM9 specific CD8+ T cells. Illumina bead array and transcriptional analyses were used to assess the transcriptional profiles and the three different CD8+ T cell populations displayed unique transcriptional patterns. This pilot study demonstrates that rapid and specific immunostaining combined with laser capture microdissection in concert with transcriptional profiling may be used to elucidate phenotypic differences between CD8+ T cells in SIV infection. Such technologies may be useful to determine differences in functional activities of HIV/SIV specific T cells. PMID:26986062

  1. In Situ Staining and Laser Capture Microdissection of Lymph Node Residing SIV Gag-Specific CD8+ T cells--A Tool to Interrogate a Functional Immune Response Ex Vivo.

    PubMed

    Tjernlund, Annelie; Burgener, Adam; Lindvall, Jessica M; Peng, Tao; Zhu, Jia; Öhrmalm, Lars; Picker, Louis J; Broliden, Kristina; McElrath, M Juliana; Corey, Lawrence

    2016-01-01

    While a plethora of data describes the essential role of systemic CD8+ T cells in the control of SIV replication little is known about the local in situ CD8+ T cell immune responses against SIV at the intact tissue level, due to technical limitations. In situ staining, using GagCM9 Qdot 655 multimers, were here combined with laser capture microdissection to detect and collect SIV Gag CM9 specific CD8+ T cells in lymph node tissue from SIV infected rhesus macaques. CD8+ T cells from SIV infected and uninfected rhesus macaques were also collected and compared to the SIV GagCM9 specific CD8+ T cells. Illumina bead array and transcriptional analyses were used to assess the transcriptional profiles and the three different CD8+ T cell populations displayed unique transcriptional patterns. This pilot study demonstrates that rapid and specific immunostaining combined with laser capture microdissection in concert with transcriptional profiling may be used to elucidate phenotypic differences between CD8+ T cells in SIV infection. Such technologies may be useful to determine differences in functional activities of HIV/SIV specific T cells. PMID:26986062

  2. Laser application in the field of vascular anastomosis: Experimental and clinical study

    NASA Astrophysics Data System (ADS)

    Okada, Masayoshi

    2005-07-01

    Nowadays, much attention has been paid to keep long-term patency after vascular anastomosis especially for small-caliber vessels in vascular surgery. From these standpoints, low energy CO2 laser was used to make an anastomosis especially for small -caliber vessels. And then an availability of vascular anastomosis by laser could be apparently recognized.

  3. Online, Absolute Quantitation of Propranolol from Spatially Distinct 20- and 40-μm Dissections of Brain, Liver, and Kidney Thin Tissue Sections by Laser Microdissection-Liquid Vortex Capture-Mass Spectrometry.

    PubMed

    Cahill, John F; Kertesz, Vilmos; Weiskittel, Taylor M; Vavrek, Marissa; Freddo, Carol; Van Berkel, Gary J

    2016-06-01

    Spatial resolved quantitation of chemical species in thin tissue sections by mass spectrometric methods has been constrained by the need for matrix-matched standards or other arduous calibration protocols and procedures to mitigate matrix effects (e.g., spatially varying ionization suppression). Reported here is the use of laser "cut and drop" sampling with a laser microdissection-liquid vortex capture electrospray ionization tandem mass spectrometry (LMD-LVC/ESI-MS/MS) system for online and absolute quantitation of propranolol in mouse brain, kidney, and liver thin tissue sections of mice administered with the drug at a 7.5 mg/kg dose, intravenously. In this procedure either 20 μm × 20 μm or 40 μm × 40 μm tissue microdissections were cut and dropped into the flowing solvent of the capture probe. During transport to the ESI source drug related material was completely extracted from the tissue into the solvent, which contained a known concentration of propranolol-d7 as an internal standard. This allowed absolute quantitation to be achieved with an external calibration curve generated from standards containing the same fixed concentration of propranolol-d7 and varied concentrations of propranolol. Average propranolol concentrations determined with the laser "cut and drop" sampling method closely agreed with concentration values obtained from 2.3 mm diameter tissue punches from serial sections that were extracted and quantified by HPLC/ESI-MS/MS measurements. In addition, the relative abundance of hydroxypropranolol glucuronide metabolites were recorded and found to be consistent with previous findings. PMID:27214103

  4. Online, absolute quantitation of propranolol from spatially distinct 20-μm and 40-μm dissections of brain, liver, and kidney thin tissue sections by laser microdissection – liquid vortex capture – mass spectrometry

    DOE PAGES

    Kertesz, Vilmos; Vavrek, Marissa; Freddo, Carol; Van Berkel, Gary J.; Cahill, John F.; Weiskittel, Taylor M.

    2016-05-23

    Here, spatial resolved quantitation of chemical species in thin tissue sections by mass spectrometric methods has been constrained by the need for matrix-matched standards or other arduous calibration protocols and procedures to mitigate matrix effects (e.g., spatially varying ionization suppression). Reported here is the use of laser cut and drop sampling with a laser microdissection-liquid vortex capture electrospray ionization tandem mass spectrometry (LMD-LVC/ESI-MS/MS) system for online and absolute quantitation of propranolol in mouse brain, kidney, and liver thin tissue sections of mice administered with the drug at a 7.5 mg/kg dose, intravenously. In this procedure either 20 μm x 20more » μm or 40 μm x 40 μm tissue microdissections were cut and dropped into the flowing solvent of the capture probe. During transport to the ESI source drug related material was completely extracted from the tissue into the solvent, which contained a known concentration of propranolol-d7 as an internal standard. This allowed absolute quantitation to be achieved with an external calibration curve generated from standards containing the same fixed concentration of propranolold-d7 and varied concentrations of propranolol. Average propranolol concentrations determined with the laser cut and drop sampling method closely agreed with concentration values obtained from 2.3 mm diameter tissue punches from serial sections that were extracted and quantified by HPLC/ESI-MS/MS measurements. In addition, the relative abundance of hydroxypropranolol glucuronide metabolites were recorded and found to be consistent with previous findings.« less

  5. Flashlamp-excited dye laser therapy for treatment of cutaneous vascular lesions

    NASA Astrophysics Data System (ADS)

    Goldberg, David J.

    1990-06-01

    Flashlamp excited dye laser therapy represents an exciting new advance in the treatment of a variety of cutaneous vascular lesions. Portwine stains, angiomas and telangiectases can be treated in all age groups with this laser system. This paper will review the physics of flashlamp dye laser photothermolysis. The differences between argon laser photocoagulation and flashlamp excited dye laser therapy will be reviewed.

  6. Laser therapy and sclerotherapy in the treatment of oral and maxillofacial hemangioma and vascular malformations

    NASA Astrophysics Data System (ADS)

    Crişan, Bogdan; BǎciuÅ£, Mihaela; BǎciuÅ£, Grigore; Crişan, Liana; Bran, Simion; Rotar, Horatiu; Moldovan, Iuliu; Vǎcǎraş, Sergiu; Mitre, Ileana; Barbur, Ioan; Magdaş, Andreea; Dinu, Cristian

    2016-03-01

    Hemangioma and vascular malformations in the field of oral and maxillofacial surgery is a pathology more often found in recent years in patients. The aim of this study was to evaluate the efficacy of the laser photocoagulation performed with a diode laser (Ga-Al-As) 980 nm wavelength in the treatment of vascular lesions which are located on the oral and maxillofacial areas, using color Doppler ultrasonography for evaluation of the results. We also made a comparison between laser therapy and sclerotherapy in order to establish treatment protocols and recommendations associated with this pathology. We conducted a controlled study on a group of 92 patients (38 male and 54 female patients, with an average age of 36 years) having low flow hemangioma and vascular malformations. Patients in this trial received one of the methods of treatment for vascular lesions such as hemangioma and vascular malformations: laser therapy or sclerotherapy. After laser therapy we have achieved a reduction in size of hemangioma and vascular malformations treated with such a procedure, and the aesthetic results were favorable. No reperfusion or recanalization of laser treated vascular lesions was observed after an average follow-up of 6 to 12 months. In case of sclerotherapy a reduction in the size of vascular lesions was also obtained. The 980 nm diode laser has been proved to be an effective tool in the treatment of hemangioma and vascular malformations in oral and maxillofacial area. Laser therapy in the treatment of vascular lesions was more effective than the sclerotherapy procedure.

  7. A Cross-Platform Comparison of Genome-Wide Expression Changes of Laser Microdissected Lung Tissue of C-Raf Transgenic Mice Using 3′IVT and Exon Array

    PubMed Central

    Londhe, Kishor Bapu; Borlak, Juergen

    2012-01-01

    Microarrays are widely used to study genome-wide gene expression changes in different conditions most notably disease, growth, or to investigate the effects of drugs on entire genomes. While the number and gene probe sequences to investigate individual gene expression changes differs amongst manufactures, the design for all of the probes is biased towards the 3′ region. With the advent of exon arrays, transcripts of any known or predicted exon can be investigated to facilitate the study of genome-wide alternative splicing events. Thus, the use of exon arrays provides unprecedented opportunities in gene expression studies. However, it remains a major challenge to directly compare gene expression data derived from oligonucleotide to exon arrays. In the present study, genome-wide expression profiling of Laser Micro-dissected Pressure Catapulted (LMPC) samples of c-Raf mouse lung adenocarcinoma, dysplasia, unaltered transgenic and non-transgenic tissues was performed using the Affymetrix GeneChip Mouse Genome 430 2.0 Array and whole genome Mouse Exon 1.0 ST Array. Based on individual group comparisons 52 to 83% of regulated genes were similar in direction, but fold changes of regulated genes disagreed when data amongst the two platforms were compared. Furthermore, for 27 regulated genes opposite direction of gene expression was observed when the two platforms were compared pointing to the need to assess alternative splicing events at the 3′ end. Taken collectively, exon arrays can be performed even with laser microdissected samples but fold change gene expression changes differ considerably between 3′IVT array and exon arrays with alternative splicing events contributing to apparent differences in gene expression changes. PMID:22815814

  8. Treatment of facial vascular lesions with an argon laser

    NASA Astrophysics Data System (ADS)

    Szymanczyk, Jacek; Golebiowska, Aleksandra; Michalska, I.

    1996-03-01

    Two-hundred-ninety-six patients with various vascular lesions of the face have been treated with argon laser LAK-1 in the Department of Dermatology Warsaw Medical Academy since April 1992. The diagnosis of the treated lesions was port-wine stains, multiple telangiectasiae and small, most often induced by trauma hemangioma cavernosum of the lip. Best results were achieved in the patients with small hemangiomas cavernosum of the lip and multiple telangiectasiae on the face. Cure rate in this group was 100%. In 112 port-wine stain cases fading of 50 - 75% comparing with the adjacent skin was achieved. With stress, the argon laser therapy is a method of choice for the treatment of hemangioma cavernosum, port-wine stains and multiple teleagiectasiae of the face.

  9. [Therapeutic indications for percutaneous laser in patients with vascular malformations and tumors].

    PubMed

    Labau, D; Cadic, P; Ouroussoff, G; Ligeron, C; Laroche, J-P; Guillot, B; Dereure, O; Quéré, I; Galanaud, J-P

    2014-12-01

    Lasers are increasingly used to treat vascular abnormalities. Indeed, this technique is non-invasive and allows a specific treatment. The aim of this review is to present some biophysical principles of the lasers, to describe the different sorts of lasers available for treatment in vascular medicine indications. Three principal lasers exist in vascular medicine: the pulsed-dye laser, for the treatment of superficial pink lesions, the NdYAG-KTP laser for purple and bigger lesions, and the NdYAG long pulse laser for even deeper and bigger vascular lesions. In vascular malformations, port wine stains can also be treated by pulsed-dye laser, KTP or NdYAG when they are old and thick. Telangiectasias are good indications for the three sorts of lasers, depending on their depth, color and size. Microcystic lymphatic malformations can be improved by laser treatment. Arterio-venous malformations constitute a contraindication of laser treatment. In vascular tumors, involuted infantile hemangiomas constitute an excellent indication of pulsed-dye laser treatment. Controlled studies are necessary to evaluate and to compare the efficacy of each laser, in order to determine their optimal indications and optimal parameters for each machine.

  10. Vascular lasers and IPLS: guidelines for care from the European Society for Laser Dermatology (ESLD).

    PubMed

    Adamic, Metka; Troilius, Agneta; Adatto, Maurice; Drosner, Michael; Dahmane, Raja

    2007-06-01

    Dermatology and dermatologic surgery have rapidly evolved during the last two decades thanks to the numerous technological and scientific acquisitions focused on improved precision in the diagnosis and treatment of skin alterations. Given the proliferation of new devices for the treatment of vascular lesions, we have considerably changed our treatment approach. Lasers and non-coherent intense pulse light sources (IPLS) are based on the principle of selective photothermolysis and can be used for the treatment of many vascular skin lesions. A variety of lasers has recently been developed for the treatment of congenital and acquired vascular lesions which incorporate these concepts into their design. The list is a long one and includes pulsed dye (FPDL, APDL) lasers (577 nm, 585 nm and 595 nm), KTP lasers (532 nm), long pulsed alexandrite lasers (755 nm), pulsed diode lasers (in the range of 800 to 900 nm), long pulsed 1064 Nd:YAG lasers and intense pulsed light sources (IPLS, also called flash-lights or pulsed light sources). Several vascular lasers (such as argon, tunable dye, copper vapour, krypton lasers) which were used in the past are no longer useful as they pose a higher risk of complications such as dyschromia (hypopigmentation or hyperpigmentation) and scarring. By properly selecting the wavelength which is maximally absorbed by the target--also called the chromophore (haemoglobin in the red blood cells within the vessels)--and a corresponding pulse duration which is shorter than the thermal relaxation time of that target, the target can be preferentially injured without transferring significant amounts of energy to surrounding tissues (epidermis and surrounding dermal tissue). Larger structures require more time for sufficient heat absorption. Therefore, a longer laser-pulse duration has to be used. In addition, more deeply situated vessels require the use of longer laser wavelengths (in the infrared range) which can penetrate deeper into the skin. Although

  11. Microdissection approaches in tuberculosis research

    PubMed Central

    Hudock, Teresa A.; Lackner, Andrew A.; Kaushal, Deepak

    2015-01-01

    Mycobacterium tuberculosis, a globally significant pathogen, results in active or latent tuberculosis. The granuloma is the characteristic lesion that offers insight into host-pathogen interactions in these distinct states. Microdissection provides a way to isolate and consequently investigate specific tissue sections. We review various techniques available and in use. PMID:25164280

  12. Comparison of different lasers in terms of thrombogenicity of the laser-treated vascular wall.

    PubMed

    Ragimov, S E; Belyaev, A A; Vertepa, I A; Dolgov, V V; Furzikov, N P; Akchurin, R S; Repin, V S; Trubetskoy, A V

    1988-01-01

    The thrombogenic properties of the laser-treated vascular wall are reported as quantitatively assessed on the basis of 33 experiments with peripheral canine vessel segments. Three types of surfaces, namely, 1) intact, 2) mechanically de-endothelized, and 3) postlaser, have been simultaneously exposed to platelet-enriched plasma in a sequentially organized artificial circulation system. Then the adherent platelets have been counted on the treated surfaces, and the relative thrombogenicity index has been calculated according to the equation T = (A1 - Aint)/(Ad - Aint), where Aint, Ad, A1 = the adherent platelet counts on intact, mechanically de-endothelized, and laser-treated surfaces, respectively. The following lasers have been evaluated: 1) Nd-YAG, 1,060 nm, continuous wave, 4W; 2) argon-ion, 480 and 514 nm and argon-ion, 350 nm, continuous-wave, 1 W and 400 mW [corrected], respectively; 3) excimer XeCl, 308 nm, pulsed, 30 mJ per pulse, repetition rate 10 Hz [corrected]; and also 4) the laser-heated metal probe (2 mm diameter, Trimedyne, Nd-YAG) 1,060 nm, 8 W. The thrombogenicity index values obtained were 83 +/- 7, 72 +/- 8, 57 +/- 9, 63 +/- 7, and 82 +/- 9%, respectively. The differences between these values were statistically insignificant. The data are suggestive of the essential requirement of, at least, anticoagulant therapy after laser angioplasty irrespective of the laser type.

  13. Early transcriptomic events in microdissected Arabidopsis nematode-induced giant cells.

    PubMed

    Barcala, Marta; García, Alejandra; Cabrera, Javier; Casson, Stuart; Lindsey, Keith; Favery, Bruno; García-Casado, Gloria; Solano, Roberto; Fenoll, Carmen; Escobar, Carolina

    2010-02-01

    Root-knot nematodes differentiate highly specialized feeding cells in roots (giant cells, GCs), through poorly characterized mechanisms that include extensive transcriptional changes. While global transcriptome analyses have used galls, which are complex root structures that include GCs and surrounding tissues, no global gene expression changes specific to GCs have been described. We report on the differential transcriptome of GCs versus root vascular cells, induced in Arabidopsis by Meloidogyne javanica at a very early stage of their development, 3 days after infection (d.p.i.). Laser microdissection was used to capture GCs and root vascular cells for microarray analysis, which was validated through qPCR and by a promoter-GUS fusion study. Results show that by 3 d.p.i., GCs exhibit major gene repression. Although some genes showed similar regulation in both galls and GCs, the majority had different expression patterns, confirming the molecular distinctiveness of the GCs within the gall. Most of the differentially regulated genes in GCs have no previously assigned function. Comparisons with other transcriptome analyses revealed similarities between GCs and cell suspensions differentiating into xylem cells. This suggests a molecular link between GCs and developing vascular cells, which represent putative GC stem cells. Gene expression in GCs at 3 d.p.i. was also found to be similar to crown galls induced by Agrobacterium tumefaciens, a specialized root biotroph. PMID:20003167

  14. Nd:YAG laser photocoagulation of benign oral vascular lesions: a case series.

    PubMed

    Medeiros, Rui; Silva, Igor Henrique; Carvalho, Alessandra Tavares; Leão, Jair Carneiro; Gueiros, Luiz Alcino

    2015-11-01

    Vascular anomalies of the head and neck are common lesions usually associated with functional and/or aesthetic limitations. The aim of the present paper was to report a case series of oral vascular malformations treated with Nd:YAG laser photocoagulation, highlighting the clinical evolution and post-surgical complications. Fifteen patients diagnosed with oral vascular malformations were treated with Nd:YAG laser followed by three sessions of biostimulation. None of the patients presented post-surgical pain, but 6 of 15 patients (40%) experienced minimal post-surgical complications. All cases presented complete resolution of the lesions after laser treatment. More importantly, 12 out of 15 (80%) resolved after a single session. Low morbidity, minimal patient discomfort, and satisfactory aesthetic results point Nd:YAG laser photocoagulation as a promising option for the management of benign oral vascular lesions.

  15. Laser therapy for selected cutaneous vascular lesions in the pediatric population: a review.

    PubMed

    Tan, O T; Gilchrest, B A

    1988-10-01

    Three cutaneous vascular lesions of childhood (the spider angioma [nevus araneus], the strawberry hemangioma, and the port wine stain) are reviewed, with particular emphasis on the present and future role of laser therapy in their management. PMID:3050868

  16. The stage-specific testicular germ cell apoptotic response to low-dose X-irradiation and 2,5-hexanedione combined exposure. I: Validation of the laser capture microdissection method for qRT-PCR array application.

    PubMed

    Catlin, Natasha R; Huse, Susan M; Boekelheide, Kim

    2014-12-01

    Over the past decade, laser capture microdissection (LCM) has grown as a tool for gene expression profiling of small numbers of cells from tumor samples and of specific cell populations in complex tissues. LCM can be used to study toxicant effects on selected cell populations within the testis at different stages of spermatogenesis. There are several LCM-related hurdles to overcome, including issues inherent to the method itself, as well as biases that result from amplifying the LCM-isolated RNA. Many technical issues associated with the LCM method are addressed here, including increasing RNA yield and obtaining more accurate quantification of RNA yields. We optimized the LCM method optimized to generate RNA quantities sufficient for quantitative reverse transcription polymerase chain reaction (qRT-PCR) array analysis without amplification and were able to validate the method through direct comparison of results from unamplified and amplified RNA from individual samples. The addition of an amplification step for gene expression studies using LCM RNA resulted in a bias, especially for low abundance transcripts. Although the amplification bias was consistent across samples, researchers should use caution when comparing results generated from amplified and unamplified LCM RNA. Here, we have validated the use of LCM-derived RNA with the qRT-PCR array, improving our ability to investigate cell-type and stage-specific responses to toxicant exposures.

  17. Whole transverse section and specific-tissue analysis of secondary metabolites in seven different grades of root of Paeonia lactiflora using laser microdissection and liquid chromatography-quadrupole/time of flight-mass spectrometry.

    PubMed

    Wang, Qiuling; Liang, Zhitao; Peng, Yong; Hou, Jun Ling; Wei, Sheng Li; Zhao, Zhong Zhen; Wang, Wen Quan

    2015-01-25

    The root of Paeonia lactiflora Pall. is widely used in the pharmaceutical, food and cosmetic industries. For these purposes, roots are graded according to diameter, with larger roots considered to be of better quality. To assess the inherent quality of different grades and of different tissues in roots of P. lactiflora, here laser microdissection coupled with UPLC-Q/TOF-MS was applied. The results show the quantity of pharmaceutically important components decreased with increase in root diameter from 0.3cm to 0.7cm. Above 0.7cm of diameter, quantity and diversity of these components increased proportionally with increase in root diameter. The tissue-specific study indicated that the high content of paeoniflorin and albiflorin are mainly distributed in the cork and cortex. According to the results of this study, the roots of P. lactiflora greater than 1.7cm in diameter are of better quality medicinal use than smaller, and the commercial grades chose was best cutoff points. The fine roots and the outer bank of roots, which besides the commercial grades, contain such significant amounts of chemical components too. This study provides a new and practical method for evaluating the different grades of P. lactiflora. PMID:25462115

  18. A gene expression analysis of syncytia laser microdissected from the roots of the Glycine max (soybean) genotype PI 548402 (Peking) undergoing a resistant reaction after infection by Heterodera glycines (soybean cyst nematode).

    PubMed

    Klink, Vincent P; Hosseini, Parsa; Matsye, Prachi; Alkharouf, Nadim W; Matthews, Benjamin F

    2009-12-01

    The syncytium is a nurse cell formed within the roots of Glycine max by the plant parasitic nematode Heterodera glycines. Its development and maintenance are essential for nematode survival. The syncytium appears to undergo two developmental phases during its maturation into a functional nurse cell. The first phase is a parasitism phase where the nematode establishes the molecular circuitry that during the second phase ensures a compatible interaction with the plant cell. The cytological features of syncytia undergoing susceptible or resistant reactions appear the same during the parasitism phase. Depending on the outcome of any defense response, the second phase is a period of syncytium maintenance (susceptible reaction) or failure (resistant reaction). In the analyses presented here, the localized gene expression occurring at the syncytium during the resistant reaction was studied. This was accomplished by isolating syncytial cells from Glycine max genotype Peking (PI 548402) by laser capture microdissection. Microarray analyses using the Affymetrix soybean GeneChip directly compared Peking syncytia undergoing a resistant reaction to those undergoing a susceptible reaction during the parasitism phase of the resistant reaction. Those analyses revealed lipoxygenase-9 and lipoxygenase-4 as the most highly induced genes in the resistant reaction. The analysis also identified induced levels of components of the phenylpropanoid pathway. These genes included phenylalanine ammonia lyase, chalcone isomerase, isoflavone reductase, cinnamoyl-CoA reductase and caffeic acid O-methyltransferase. The presence of induced levels of these genes implies the importance of jasmonic acid and phenylpropanoid signaling pathways locally at the site of the syncytium during the resistance phase of the resistant reaction. The analysis also identified highly induced levels of four S-adenosylmethionine synthetase genes, the EARLY-RESPONSIVE TO DEHYDRATION 2 gene and the 14-3-3 gene known as

  19. Comparison of the treatment of vascular lesions with the copper-vapor laser and flashlamp-pumped dye laser

    NASA Astrophysics Data System (ADS)

    Flock, Stephen T.; Waner, Milton; McGrew, Ben; Colvin, G. B.; Montague, Donna

    1992-08-01

    Vascular lesions such as port-wine stains and telangiectases are sometimes treated with carbon-dioxide lasers, argon lasers or argon-pumped dye lasers; however these lasers are non- specific in their thermal effect on tissues and as a result often cause significant scarring. Recently, evidence has accumulated that the flashlamp-pumped dye (585 nm) and copper- vapor (578 nm) lasers, which produce pulsed light that is efficiently absorbed by hemoglobin, are more selective in coagulating abnormal vascular tissue and as a result give a superior clinical result. It is not yet clear what the most important physical and biological mechanisms are during the light-tissue interaction mediated by these two lasers. The post-treatment sequence of events is different for tissue irradiated by each laser; most significantly, the flashlamp-pumped dye laser causes significant transient purpura, whereas the copper vapor laser causes blanching and eschar formation. The clinical outcome, that is regression of the lesion, is equally successful with either laser although some evidence has accumulated showing that the flashlamp-pumped dye laser is best suited to the treatment of small vessel disease while the copper-vapor laser is better for the treatment of large vessel disease. In this paper, we will discuss our observations of the treatment of vascular lesions on humans with the copper-vapor and flashlamp-pumped dye lasers using empirically derived efficacious treatment parameters. Mathematical models of light and heat propagation and in vivo experiments involving mice ears and rat skin flaps will be used to elucidate what we feel are the important underlying mechanisms of this vascular lesion laser therapy.

  20. Pulsed dye laser application in ablation of vascular ectasias of the larynx: a preliminary animal study

    NASA Astrophysics Data System (ADS)

    Woo, Peak; Wang, Zhi; Perrault, Donald F., Jr.; McMillan, Kathleen; Pankratov, Michail M.

    1995-05-01

    Vascular ectasias (dilatation) and vascular lesions of the larynx are difficult to treat with exciting modalities. Varix (enlarged vessel) of the vocal folds, vocal fold hemorrhage, vascular polyp, hemangioma, intubation or contact granuloma are common problems which disturb voice. Current applications of CO2 laser and cautery often damage the delicate vocal fold cover. The 585 nm dermatologic pulsed dye laser may be an ideal substitute. Two adult canines were examined under anesthesia via microlaryngoscopy technique. Pulsed dye laser (SPTL-1a, Candela Laser Corp., Wayland, MA) energy was delivered via the micromanipulator with the 3.1-mm spot size in single pulses of 6, 8, and 10 Joules/cm2 and applied to the vessels of the vocal folds, epiglottis, and arytenoid cartilage. Endoscopic examination was carried out immediately after the treatment and at 4 weeks postoperatively. The animals were sacrificed at 3 weeks, larynges excised, and whole organ laryngeal section were prepared for histology. Pulsed dye laser thrombosed vessels of the vocal fold using 6 or 8 Joules/cm2. Vascular break and leakage occurred at 10 Joules/cm2. Follow up examination showed excellent vessel obliteration or thrombosis without scarring or injury to the overlying tissues. Histologic examination shows vascular thrombosis without inflammation and fibrosis in the vocal fold cover. Pulsed dye laser may have promise in treatment of vascular lesions of the larynx and upper airway.

  1. Ten years of laser treatment of congenital vascular disorders: techniques and results

    NASA Astrophysics Data System (ADS)

    Philipp, Carsten M.; Berlien, Hans-Peter; Poetke, Margitta; Waldschmidt, Juergen

    1994-12-01

    During the period of January 1984 - July 1993, we have treated 611 children with more than 2000 lesions of congenital vascular disorders (CVD) such as hemangiomas and vascular malformations. This number does not include the patients with port wine stains, which also have been treated by means of laser. Most of the CVD patients (n equals 467) presented hemangiomas of the face, the anogenital region, and the extremities, some were located in the trachea or mouth or in the urogenital tract. All of these hemangiomas were growing prior to intervention or showed complications such as bleeding, ulceration, superinfection, or obstruction. Nearly a quarter (n equals 144) of the patients presented vascular malformations, either of singular vessel type involvement or of mixed vascular genesis (venous, arterio- venous, veno-lymphatic or lymphatic) with various complications like tracheal obstruction or recurrent thrombophlebitis. According to our step program, which is based on a clinical classification, the hemangiomas were treated as early as possible, while the vascular malformations were only treated with laser when no other therapeutic technique (embolization, resection) was suitable. All patients were referred for laser treatment from other centers. The lasers used were Nd:YAG and Argon lasers with transcutaneous application with or without continuous ice-cube surface cooling or interstitial laser application. The treatments were performed either on in- or outpatient basis according to age, localization and with good to excellent results in most cases and a complication rate of less than 2%.

  2. Laser assisted vascular anastomosis (LAVA): a promising nonsuture technique for surgery

    NASA Astrophysics Data System (ADS)

    Chen, Chen; Peng, Fei; Xu, Dahai; Cheng, Qinghua

    2009-08-01

    The first successful experiment of laser vascular welding was reported in 1979. Laser assisted vascular anastomosis (LAVA) is looked as a particularly promising non-suture method in future. We performed a Medline literature search on laser vessel welding combined with cross-referencing. According to the former experimental animal studies, CO2-, argon-, diode-, KTP-, Holmium:YAG-, and Nd:YAG-lasers have been used for LAVA. Almost all lasers have been used in combination with stay suture and/or solders in order to improve the strength on anastomosis site. Advantages of LAVA are minimal vessel damage, faster operation and the potential for minimally invasive application. However, the clinical application of LAVA is still seldom employed because of aneurysm formation. In conclusion of the literature study, the diode laser is the most popular, but long-term evaluation is required.

  3. Production of high quality brain-derived neurotrophic factor (BDNF) and tropomyosin receptor kinase B (TrkB) RNA from isolated populations of rat spinal cord motor neurons obtained by Laser Capture Microdissection (LCM).

    PubMed

    Mehta, Prachi; Premkumar, Brian; Morris, Renée

    2016-08-01

    The mammalian central nervous system (CNS) is composed of multiple cellular elements, making it challenging to segregate one particular cell type to study their gene expression profile. For instance, as motor neurons represent only 5-10% of the total cell population of the spinal cord, meaningful transcriptional analysis on these neurons is almost impossible to achieve from homogenized spinal cord tissue. A major challenge faced by scientists is to obtain good quality RNA from small amounts of starting material. In this paper, we used Laser Capture Microdissection (LCM) techniques to identify and isolate spinal cord motor neurons. The present analysis revealed that perfusion with paraformaldehyde (PFA) does not alter RNA quality. RNA integrity numbers (RINs) of tissue samples from rubrospinal tract (RST)-transected, intact spinal cord or from whole spinal cord homogenate were all above 8, which indicates intact, high-quality RNA. Levels of mRNA for brain-derived neurotrophic factor (BDNF) or for its tropomyosin receptor kinase B (TrkB) were not affected by rubrospinal tract (RST) transection, a surgical procedure that deprive motor neurons from one of their main supraspinal input. The isolation of pure populations of neurons with LCM techniques allows for robust transcriptional characterization that cannot be achieved with spinal cord homogenates. Such preparations of pure population of motor neurons will provide valuable tools to advance our understanding of the molecular mechanisms underlying spinal cord injury and neuromuscular diseases. In the near future, LCM techniques might be instrumental to the success of gene therapy for these debilitating conditions.

  4. Fiber laser micromachining of thin NiTi tubes for shape memory vascular stents

    NASA Astrophysics Data System (ADS)

    Liu, Lei; Li, Dong Bo; Tong, Yi Fei; Zhu, Yu Fu

    2016-07-01

    Nickel titanium (NiTi) alloy has widely been used in the vascular stent manufacturing due to its excellent properties. Neodymium-doped yttrium aluminum garnet (Nd:YAG) laser is commonly used for the preparation of metal vascular stents. Recently, fiber lasers have been used for stent profiling for better cutting quality. To investigate the cutting-kerf characters of NiTi vascular stents fabricated by fiber laser cutting, laser cutting experiments with thin NiTi tubes were conducted in this study, while NiTi sheets were used in other fiber laser cutting studies. Different with striation topography, new topographies such as layer topography and topography mixed with layers and striations were observed, and the underlying reason for new topographies was also discussed. Comparative research on different topographies was conducted through analyzing the surface roughness, kerf width, heat-affected zone (HAZ) and dross formation. Laser cutting process parameters have a comprehensive influence on the cutting quality; in this study, the process parameters' influences on the cutting quality were studied from the view of power density along the cutting direction. The present research provides a guideline for improving the cutting quality of NiTi vascular stents.

  5. High-flow vascular malformation treatment using ultrasound-guided laser combined with polidocanol sclerotherapy.

    PubMed

    Zhang, Yan; Zhou, Ping; Li, Lan; Li, Jia-le

    2015-07-01

    The current treatment for vascular malformations includes surgery, sclerotherapy, and embolization. However, each method has its limitations, such as recurrence, complications, scarring, and radiation exposure. Therefore, identifying an effective, minimally invasive treatment that reduces lesion recurrence is particularly important. We describe in detail a patient who received treatment with ultrasound-guided laser interruption of feeding vessels combined with polidocanol sclerotherapy after the recurrence of forearm high-flow vascular malformation.

  6. Selective disruption of vascular endothelium of zebrafish embryos by ultrafast laser microsurgical treatment

    PubMed Central

    Woo, Suk-Yi; Moon, Heh-Young; Kim, Tag Gyum; Lee, Heung Soon; Sidhu, Mehra S.; Kim, Changho; Jeon, Jae-Phil; Jeoung, Sae Chae

    2015-01-01

    In this work, we demonstrate that ultrafast laser irradiation could selectively disrupt vascular endothelium of zebrafish embryos in vivo. Ultrafast lasers minimize the collateral damage in the vicinity of the laser focus and eventually reduce coagulation in the tissues. We have also found that the threshold fluence for lesion formation of the vascular endothelium strongly depends on the developmental stage of the embryos. The threshold laser fluence required to induce apparent lesions in the vascular structure for Somite 14, 20 and 25 stages is about 5 J/cm2 ~7 J/cm2, which is much lower than that for the later development stages of Prim 16 and Prim 20 of 30 J/cm2 ~50 J/cm2. The proposed method for treating the vascular cord of zebrafish embryos in the early stage of development has potential as a selective and effective method to induce a fatal lesion in the vascular endothelium without damaging the developed blood vessels. PMID:26713187

  7. Optical diagnostics of vascular reactions triggered by weak allergens using laser speckle-contrast imaging technique

    SciTech Connect

    Kuznetsov, Yu L; Kalchenko, V V; Astaf'eva, N G; Meglinski, I V

    2014-08-31

    The capability of using the laser speckle contrast imaging technique with a long exposure time for visualisation of primary acute skin vascular reactions caused by a topical application of a weak contact allergen is considered. The method is shown to provide efficient and accurate detection of irritant-induced primary acute vascular reactions of skin. The presented technique possesses a high potential in everyday diagnostic practice, preclinical studies, as well as in the prognosis of skin reactions to the interaction with potentially allergenic materials. (laser biophotonics)

  8. Excimer laser phototherapy for the dissolution of vascular obstruction

    DOEpatents

    Gruen, D.M.; Young, C.E.; Pellin, M.J.

    1984-01-09

    Removal of abnormal human tissue with reduced thermal damage is achieved by selecting a laser having a wavelength in the order of 290 to 400 nm, orienting a laser-transmitting glass member toward the abnormal tissue and directing the laser through the glass member at power densities, pulse rates, and times sufficient to cause multiphoton absorption and bond breaking by Coulomb repulsion rather than thermal destruction. 2 figures.

  9. Effect of dye laser pulse duration on selective cutaneous vascular injury

    SciTech Connect

    Garden, J.M.; Tan, O.T.; Kerschmann, R.; Boll, J.; Furumoto, H.; Anderson, R.R.; Parrish, J.A.

    1986-11-01

    The pulsed dye laser at 577 nm, a wavelength well absorbed by oxyhemoglobin, causes highly selective thermal injury to cutaneous blood vessels. Confinement of thermal damage to microvessels is, in theory, related to the laser exposure time (pulsewidth) on selective vascular injury. This study investigates the effect of 577 nm dye laser pulsewidth on selective vascular injury. Nine Caucasian, normal volunteers received 577 nm dye laser exposures at pulsewidths of 1.5-350 microseconds to their skin. Clinical purpura threshold exposure doses were determined in each volunteer, and biopsies of threshold and suprathreshold doses were examined in each volunteer. The laser exposure dose required to produce purpura increased as pulsewidth increased in all 9 subjects (p less than 0.001). This finding corresponds to laser pulsewidths equal to or exceeding the thermal relaxation times for dermal blood vessels. Histologically, vessel damage was selectively, but qualitatively, different for short vs long pulsewidths. Pulsewidths shorter than 20 microseconds caused vessel wall fragmentation and hemorrhage, whereas longer pulsewidths caused no significant hemorrhage. The purpura noted clinically appears to be due to a coagulum of intralumenal denatured erythrocytes. At 24 h, there was marked vessel wall necrosis at all pulsewidths. The short pulsewidths may cause erythrocyte vaporization, rapid thermal expansion, and mechanical vessel rupture with hemorrhage. Long pulsewidths appear to cause thermal denaturation with less mechanical vessel damage. The selective, nonhemorrhagic, vascular necrosis caused by the long-pulsewidth dye laser may lead to a more desirable clinical outcome in the therapy of blood vessel disease processes.

  10. The Chromosome Microdissection and Microcloning Technique.

    PubMed

    Zhang, Ying-Xin; Deng, Chuan-Liang; Hu, Zan-Min

    2016-01-01

    Chromosome microdissection followed by microcloning is an efficient tool combining cytogenetics and molecular genetics that can be used for the construction of the high density molecular marker linkage map and fine physical map, the generation of probes for chromosome painting, and the localization and cloning of important genes. Here, we describe a modified technique to microdissect a single chromosome, paint individual chromosomes, and construct single-chromosome DNA libraries. PMID:27511173

  11. CpG Methylation Analysis of HPV16 in Laser Capture Microdissected Archival Tissue and Whole Tissue Sections from High Grade Anal Squamous Intraepithelial Lesions: A Potential Disease Biomarker.

    PubMed

    Molano, Monica; Tabrizi, Sepehr N; Garland, Suzanne M; Roberts, Jennifer M; Machalek, Dorothy A; Phillips, Samuel; Chandler, David; Hillman, Richard J; Grulich, Andrew E; Jin, Fengyi; Poynten, I Mary; Templeton, David J; Cornall, Alyssa M

    2016-01-01

    Incidence and mortality rates of anal cancer are increasing globally. More than 90% of anal squamous cell carcinomas (ASCC) are associated with human papillomavirus (HPV). Studies on HPV-related anogenital lesions have shown that patterns of methylation of viral and cellular DNA targets could potentially be developed as disease biomarkers. Lesion-specific DNA isolated from formalin-fixed paraffin-embedded (FFPE) tissues from existing or prospective patient cohorts may constitute a valuable resource for methylation analysis. However, low concentrations of DNA make these samples technically challenging to analyse using existing methods. We therefore set out to develop a sensitive and reproducible nested PCR-pyrosequencing based method to accurately quantify methylation at 10 CpG sites within the E2BS1, E2BS2,3,4 and Sp1 binding sites in the viral upstream regulatory region of HPV16 genome. Methylation analyses using primary and nested PCR-pyrosequencing on 52 FFPE tissue [26 paired whole tissue sections (WTS) and laser capture microdissected (LCM) tissues] from patients with anal squamous intraepithelial lesions was performed. Using nested PCR, methylation results were obtained for the E2BS1, E2BS2,3,4 and Sp1 binding sites in 86.4% of the WTS and 81.8% of the LCM samples. Methylation patterns were strongly correlated within median values of matched pairs of WTS and LCM sections, but overall methylation was higher in LCM samples at different CpG sites. High grade lesions showed low methylation levels in the E2BS1 and E2BS2 regions, with increased methylation detected in the E2BS,3,4/Sp1 regions, showing the highest methylation at CpG site 37. The method developed is highly sensitive in samples with low amounts of DNA and demonstrated to be suitable for archival samples. Our data shows a possible role of specific methylation in the HPV16 URR for detection of HSIL.

  12. CpG Methylation Analysis of HPV16 in Laser Capture Microdissected Archival Tissue and Whole Tissue Sections from High Grade Anal Squamous Intraepithelial Lesions: A Potential Disease Biomarker.

    PubMed

    Molano, Monica; Tabrizi, Sepehr N; Garland, Suzanne M; Roberts, Jennifer M; Machalek, Dorothy A; Phillips, Samuel; Chandler, David; Hillman, Richard J; Grulich, Andrew E; Jin, Fengyi; Poynten, I Mary; Templeton, David J; Cornall, Alyssa M

    2016-01-01

    Incidence and mortality rates of anal cancer are increasing globally. More than 90% of anal squamous cell carcinomas (ASCC) are associated with human papillomavirus (HPV). Studies on HPV-related anogenital lesions have shown that patterns of methylation of viral and cellular DNA targets could potentially be developed as disease biomarkers. Lesion-specific DNA isolated from formalin-fixed paraffin-embedded (FFPE) tissues from existing or prospective patient cohorts may constitute a valuable resource for methylation analysis. However, low concentrations of DNA make these samples technically challenging to analyse using existing methods. We therefore set out to develop a sensitive and reproducible nested PCR-pyrosequencing based method to accurately quantify methylation at 10 CpG sites within the E2BS1, E2BS2,3,4 and Sp1 binding sites in the viral upstream regulatory region of HPV16 genome. Methylation analyses using primary and nested PCR-pyrosequencing on 52 FFPE tissue [26 paired whole tissue sections (WTS) and laser capture microdissected (LCM) tissues] from patients with anal squamous intraepithelial lesions was performed. Using nested PCR, methylation results were obtained for the E2BS1, E2BS2,3,4 and Sp1 binding sites in 86.4% of the WTS and 81.8% of the LCM samples. Methylation patterns were strongly correlated within median values of matched pairs of WTS and LCM sections, but overall methylation was higher in LCM samples at different CpG sites. High grade lesions showed low methylation levels in the E2BS1 and E2BS2 regions, with increased methylation detected in the E2BS,3,4/Sp1 regions, showing the highest methylation at CpG site 37. The method developed is highly sensitive in samples with low amounts of DNA and demonstrated to be suitable for archival samples. Our data shows a possible role of specific methylation in the HPV16 URR for detection of HSIL. PMID:27529629

  13. CpG Methylation Analysis of HPV16 in Laser Capture Microdissected Archival Tissue and Whole Tissue Sections from High Grade Anal Squamous Intraepithelial Lesions: A Potential Disease Biomarker

    PubMed Central

    Molano, Monica; Tabrizi, Sepehr N.; Garland, Suzanne M.; Roberts, Jennifer M.; Machalek, Dorothy A.; Phillips, Samuel; Chandler, David; Hillman, Richard J.; Grulich, Andrew E.; Jin, Fengyi; Poynten, I. Mary; Templeton, David J.; Cornall, Alyssa M.

    2016-01-01

    Incidence and mortality rates of anal cancer are increasing globally. More than 90% of anal squamous cell carcinomas (ASCC) are associated with human papillomavirus (HPV). Studies on HPV-related anogenital lesions have shown that patterns of methylation of viral and cellular DNA targets could potentially be developed as disease biomarkers. Lesion-specific DNA isolated from formalin-fixed paraffin-embedded (FFPE) tissues from existing or prospective patient cohorts may constitute a valuable resource for methylation analysis. However, low concentrations of DNA make these samples technically challenging to analyse using existing methods. We therefore set out to develop a sensitive and reproducible nested PCR-pyrosequencing based method to accurately quantify methylation at 10 CpG sites within the E2BS1, E2BS2,3,4 and Sp1 binding sites in the viral upstream regulatory region of HPV16 genome. Methylation analyses using primary and nested PCR-pyrosequencing on 52 FFPE tissue [26 paired whole tissue sections (WTS) and laser capture microdissected (LCM) tissues] from patients with anal squamous intraepithelial lesions was performed. Using nested PCR, methylation results were obtained for the E2BS1, E2BS2,3,4 and Sp1 binding sites in 86.4% of the WTS and 81.8% of the LCM samples. Methylation patterns were strongly correlated within median values of matched pairs of WTS and LCM sections, but overall methylation was higher in LCM samples at different CpG sites. High grade lesions showed low methylation levels in the E2BS1 and E2BS2 regions, with increased methylation detected in the E2BS,3,4/Sp1 regions, showing the highest methylation at CpG site 37. The method developed is highly sensitive in samples with low amounts of DNA and demonstrated to be suitable for archival samples. Our data shows a possible role of specific methylation in the HPV16 URR for detection of HSIL. PMID:27529629

  14. Heterogeneous rpoS and rhlR mRNA levels and 16S rRNA/rDNA (rRNA gene) ratios within Pseudomonas aeruginosa biofilms, sampled by laser capture microdissection.

    PubMed

    Pérez-Osorio, Ailyn C; Williamson, Kerry S; Franklin, Michael J

    2010-06-01

    The local environmental conditions in biofilms are dependent on the impinging aqueous solution, chemical diffusion, and the metabolic activities of cells within the biofilms. Chemical gradients established in biofilms lead to physiological heterogeneities in bacterial gene expression. Previously, we used laser capture microdissection (LCM) and quantitative reverse transcription (RT)-PCR to target defined biofilm subpopulations for gene expression studies. Here, we combined this approach with quantitative PCR of bacterial DNA to normalize the amount of gene expression per cell. By comparing the ratio of 16S rRNA to 16S rDNA (rRNA gene), we demonstrated that cells at the top of thick Pseudomonas aeruginosa biofilms have 16S rRNA/genome ratios similar to those of cells in a transition from the exponential phase to the stationary phase. Cells in the middle and bottom layers of these biofilms have ratios that are not significantly different from those of stationary-phase planktonic cultures. Since much of each biofilm appeared to be in a stationary-phase-like state, we analyzed the local amounts of the stationary-phase sigma factor rpoS gene and the quorum-sensing regulator rhlR gene per cell. Surprisingly, the amount of rpoS mRNA was largest at the top of the biofilms at the air-biofilm interface. Less than one rpoS mRNA transcript per cell was observed in the middle or base of the biofilms. The rhlR mRNA content was also greatest at the top of the biofilms, and there was little detectable rhlR expression at the middle or bottom of the biofilms. While the cell density was slightly greater at the bottom of the biofilms, expression of the quorum-sensing regulator occurred primarily at the top of the biofilms, where the cell metabolic activity was greatest, as indicated by local expression of the housekeeping gene acpP and by expression from a constitutive P(trc) promoter. The results indicate that in thick P. aeruginosa biofilms, cells in the 30 microm adjacent to the

  15. Statistical characteristics of surface integrity by fiber laser cutting of Nitinol vascular stents

    NASA Astrophysics Data System (ADS)

    Fu, C. H.; Liu, J. F.; Guo, Andrew

    2015-10-01

    Nitinol alloys have been widely used in manufacturing of vascular stents due to the outstanding properties such as superelasticity, shape memory, and superior biocompatibility. Laser cutting is the dominant process for manufacturing Nitinol stents. Conventional laser cutting usually produces unsatisfactory surface integrity which has a significant detrimental impact on stent performance. Emerging as a competitive process, fiber laser with high beam quality is expected to produce much less thermal damage such as striation, dross, heat affected zone (HAZ), and recast layer. To understand the process capability of fiber laser cutting of Nitinol alloy, a design-of-experiment based laser cutting experiment was performed. The kerf geometry, roughness, topography, microstructure, and hardness were studied to better understand the nature of the HAZ and recast layer in fiber laser cutting. Moreover, effect size analysis was conducted to investigate the relationship between surface integrity and process parameters.

  16. Effect of epidermal pigmentation on selective vascular effects of pulsed laser

    SciTech Connect

    Tan, O.T.; Kerschmann, R.; Parrish, J.A.

    1984-01-01

    The effect of epidermal pigmentation on the threshold exposure dose for inducing purpura with a tunable dye laser at 577 nm, 1.5 microseconds pulse duration, was studied in 21 human volunteers with varied genetically determined amounts of melanin. More laser energy was required to produce purpura as constitutive skin pigmentation increased. Histology showed that, in lighter skin, the laser threshold dose produced the most specific vascular injury with no disruption of surrounding structures. In more pigmented skin, damage occurred in the epidermal basal layer and very few changes were seen in blood vessels below.

  17. Argon and YAG laser photocoagulation and excision of hemangiomas and vascular malformations of the nose.

    PubMed Central

    Apfelberg, D B

    1995-01-01

    A total of 22 patients--19 children, 3 adults--with a variety of hemangiomas and vascular malformations of the nose were treated over a 5-year period. Various laser modalities were used. Some lesions could be photocoagulated by the argon or the yttrium-aluminum-garnet (YAG) laser. Larger lesions were resected with the YAG laser and sapphire tips. Preliminary arteriography with superselective embolization was necessary in 1 patient. Total removal was possible in 13 patients, and no complications or side effects were noted. Images Figure 1. Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. Figure 6. PMID:7571558

  18. DNA from microdissected tissues may be extracted and stored on microscopic slides.

    PubMed

    Korabecna, M; Steiner, P; Jirkovska, M

    2016-01-01

    With regard to complex structure of tissues, laser capture microdissection represents an important step in analytical workflow streaming to proper molecular characterization of different cell types in examined samples. Therefore the simple method for simultaneous processing of higher numbers of microdissected tissues leading not only to rapid and efficient DNA isolation but allowing also the repeated sampling and easy storage may be useful in the practice of histopathological laboratories. We elaborated such a methodology applicable downstream after the microdissection from formalin-fixed paraffin embedded tissues.The tissues for examination are microdissected directly into the circular areas having the diameter 2 mm and marked on the microscopic slide. In this way, one slide is able to accommodate multiple samples. The DNA extraction is performed in low volume of buffer with Proteinase K in a droplet covered by mineral oil just on the slide. Mineral oil in the quality for molecular biology not only avoids evaporation during DNA extraction, but it helps to position the microdisssected tissue, to control the level of cell lysis microscopically and to protect the DNA sample during subsequent manipulations. We provided the evidence that DNA isolated by our methodology remains in the positions on microscopic slide for months without any changes in the lengths of available fragments and that it may be removed from each position repetitively for different kinds of analysis. The new methodological approach presented by us can be practically applied in broad spectrum of laboratories performing routinely genetic analysis on microdissected tissues. PMID:27268914

  19. DNA from microdissected tissues may be extracted and stored on microscopic slides.

    PubMed

    Korabecna, M; Steiner, P; Jirkovska, M

    2016-01-01

    With regard to complex structure of tissues, laser capture microdissection represents an important step in analytical workflow streaming to proper molecular characterization of different cell types in examined samples. Therefore the simple method for simultaneous processing of higher numbers of microdissected tissues leading not only to rapid and efficient DNA isolation but allowing also the repeated sampling and easy storage may be useful in the practice of histopathological laboratories. We elaborated such a methodology applicable downstream after the microdissection from formalin-fixed paraffin embedded tissues.The tissues for examination are microdissected directly into the circular areas having the diameter 2 mm and marked on the microscopic slide. In this way, one slide is able to accommodate multiple samples. The DNA extraction is performed in low volume of buffer with Proteinase K in a droplet covered by mineral oil just on the slide. Mineral oil in the quality for molecular biology not only avoids evaporation during DNA extraction, but it helps to position the microdisssected tissue, to control the level of cell lysis microscopically and to protect the DNA sample during subsequent manipulations. We provided the evidence that DNA isolated by our methodology remains in the positions on microscopic slide for months without any changes in the lengths of available fragments and that it may be removed from each position repetitively for different kinds of analysis. The new methodological approach presented by us can be practically applied in broad spectrum of laboratories performing routinely genetic analysis on microdissected tissues.

  20. Relevance of laser Doppler and laser speckle techniques for assessing vascular function: state of the art and future trends.

    PubMed

    Humeau-Heurtier, A; Guerreschi, E; Abraham, P; Mahé, G

    2013-03-01

    In clinical and research applications, the assessment of vascular function has become of major importance to evaluate and follow the evolution of cardiovascular pathologies, diabetes, hypertension, or foot ulcers. Therefore, the development of engineering methodologies able to monitor noninvasively blood vessel activities-such as endothelial function-is a significant and emerging challenge. Laser-based techniques have been used to respond-as much as possible-to these requirements. Among them, laser Doppler flowmetry (LDF) and laser Doppler imaging (LDI) were proposed a few decades ago. They provide interesting vascular information but possess drawbacks that prevent an easy use in some clinical situations. Recently, the laser speckle contrast imaging (LSCI) technique, a noninvasive camera-based tool, was commercialized and overcomes some of the LDF and LDI weaknesses. Our paper describes how-using engineering methodologies-LDF, LDI, and LSCI can meet the challenging clinician needs in assessing vascular function, with a special focus on the state of the art and future trends.

  1. Optimizing treatment parameters for the vascular malformations using 1064-nm Nd:YAG laser

    NASA Astrophysics Data System (ADS)

    Gong, Wei; Lin, He; Xie, Shusen

    2010-02-01

    Near infrared Nd:YAG pulsed laser treatment had been proved to be an efficient method to treat large-sized vascular malformations like leg telangiectasia for deep penetrating depth into skin and uniform light distribution in vessel. However, optimal clinical outcome was achieved by various laser irradiation parameters and the key factor governing the treatment efficacy was still unclear. A mathematical model in combination with Monte Carlo algorithm and finite difference method was developed to estimate the light distribution, temperature profile and thermal damage in epidermis, dermis and vessel during and after 1064 nm pulsed Nd:YAG laser irradiation. Simulation results showed that epidermal protection could be achieved during 1064 nm Nd:YAG pulsed laser irradiation in conjunction with cryogen spray cooling. However, optimal vessel closure and blood coagulation depend on a compromise between laser spot size and pulse duration.

  2. Fabrication and In Vitro Deployment of a Laser-Activated Shape Memory Polymer Vascular Stent

    SciTech Connect

    Baer, G M; Small IV, W; Wilson, T S; Benett, W J; Matthews, D L; Hartman, J; Maitland, D J

    2007-04-25

    Vascular stents are small tubular scaffolds used in the treatment of arterial stenosis (narrowing of the vessel). Most vascular stents are metallic and are deployed either by balloon expansion or by self-expansion. A shape memory polymer (SMP) stent may enhance flexibility, compliance, and drug elution compared to its current metallic counterparts. The purpose of this study was to describe the fabrication of a laser-activated SMP stent and demonstrate photothermal expansion of the stent in an in vitro artery model. A novel SMP stent was fabricated from thermoplastic polyurethane. A solid SMP tube formed by dip coating a stainless steel pin was laser-etched to create the mesh pattern of the finished stent. The stent was crimped over a fiber-optic cylindrical light diffuser coupled to an infrared diode laser. Photothermal actuation of the stent was performed in a water-filled mock artery. At a physiological flow rate, the stent did not fully expand at the maximum laser power (8.6 W) due to convective cooling. However, under zero flow, simulating the technique of endovascular flow occlusion, complete laser actuation was achieved in the mock artery at a laser power of {approx}8 W. We have shown the design and fabrication of an SMP stent and a means of light delivery for photothermal actuation. Though further studies are required to optimize the device and assess thermal tissue damage, photothermal actuation of the SMP stent was demonstrated.

  3. Fabrication and in vitro deployment of a laser-activated shape memory polymer vascular stent

    PubMed Central

    Baer, Géraldine M; Small, Ward; Wilson, Thomas S; Benett, William J; Matthews, Dennis L; Hartman, Jonathan; Maitland, Duncan J

    2007-01-01

    Background Vascular stents are small tubular scaffolds used in the treatment of arterial stenosis (narrowing of the vessel). Most vascular stents are metallic and are deployed either by balloon expansion or by self-expansion. A shape memory polymer (SMP) stent may enhance flexibility, compliance, and drug elution compared to its current metallic counterparts. The purpose of this study was to describe the fabrication of a laser-activated SMP stent and demonstrate photothermal expansion of the stent in an in vitro artery model. Methods A novel SMP stent was fabricated from thermoplastic polyurethane. A solid SMP tube formed by dip coating a stainless steel pin was laser-etched to create the mesh pattern of the finished stent. The stent was crimped over a fiber-optic cylindrical light diffuser coupled to an infrared diode laser. Photothermal actuation of the stent was performed in a water-filled mock artery. Results At a physiological flow rate, the stent did not fully expand at the maximum laser power (8.6 W) due to convective cooling. However, under zero flow, simulating the technique of endovascular flow occlusion, complete laser actuation was achieved in the mock artery at a laser power of ~8 W. Conclusion We have shown the design and fabrication of an SMP stent and a means of light delivery for photothermal actuation. Though further studies are required to optimize the device and assess thermal tissue damage, photothermal actuation of the SMP stent was demonstrated. PMID:18042294

  4. Comparison of different surface-cooling methods for transcutaneous laser treatment of vascular lesions

    NASA Astrophysics Data System (ADS)

    Philipp, Carsten M.; Sokoll, C.; Nowak, W.; Berlien, Hans-Peter

    1997-12-01

    The purpose of this study was the evaluation of different combined cooling and compression techniques for the treatment of vascular disorders of the skin and subdermal layers. In combination with flashlamp pumped dye lasers, argon lasers and Nd:YAG-lasers the effectiveness of glass plates, a cooling chamber with a flexible membrane and continuous ice cube cooling were evaluated in vitro by temperature measurements with thermocouples and thermographic camera readings and in vivo by laser doppler flowmetry, color coded duplex sonography and comparison of photographic documents for effectiveness and occurrence of side effects. Experimental and clinical evaluations show excellent results for skin protection, effective treatment depth enhancement and minimalization of side effects as well as for pain perception.

  5. Photosclerosis of cutaneous vascular malformations with a pulsed 810-nm diode laser

    NASA Astrophysics Data System (ADS)

    Bass, Lawrence S.

    1995-05-01

    Various continuous wave lasers have been effectively used for sclerosis of cutaneous vascular lesions. The risk of scarring has been as high as 15% in some applications, however. Pulse dye lasers have virtually eliminated scarring as a complication, substituting high cost and cumbersome operation. Pre-clinical studies have demonstrated the ability of the 810 nm gallium-aluminum-arsenide diode laser in photosclerosis. In this study, a small group of patients (n equals 6) were treated with a pulsed 810 nm diode laser (Surgimedics, The Woodlands, TX). Pulse widths used were 5 - 15 msec with corresponding energy densities during the square wave pulse of 14 - 42 J/cm2. Pulse interval was 32 msec. Lesions treated include telangiectasias (n equals 5), spider veins, (n equals 4), capillary dermal malformation (n equals 1) and a cutaneous venous malformation (n equals 1). Lower extremity spider veins were the most resistent to treatment, rarely disappearing entirely. Telangiectasias were most responsive, usually disappearing after one treatment. No scarring was noted and textural changes were seen in only one patient. While these data are preliminary and parameters have not been optimized, pulsed diode laser exposure can be an effective treatment for cutaneous vascular lesions.

  6. Optimal dye concentration and power density for laser-assisted vascular anatomosis (LAVA)

    NASA Astrophysics Data System (ADS)

    Ren, Zhen; Furnary, Anthony; Xie, Hua; Lagerquist, Kathryn A.; Burke, Allen; Prahl, Scott A.; Gregory, Kenton W.

    2003-06-01

    Laser tissue welding with albumin solder/indocyanine green (ICG) dye is an effective technique in surgical reconstruction. This study was carried out in vitro to find optimal ICG concentration and power density (PD) in laser assisted vascular anastomosis (LAVA). Fresh porcine carotid arteries incised into vascular strips (n = 120) were welded by diode laser in end-to-end with 50% albumin solder of 0.01, 0.1, and 1.0 mM ICG and at power density of 27.7, 56.7, and 76.9 W/cm2. Direct temperature was measured by inserting thermocouples outside and inside vessel. Tensile strength was tested immediately and histological study was performed. Temperature (both outside and inside vessel) significantly gradually decreasd (p < 0.01) with the increasing of ICG concentration at PD 56.7 W/cm2. Tensile strength significantly gradually decreased (p < 0.01) with increasing of ICG concentration at PD 56.7 W/cm2. Histological study showed minimal thermal injury limited to adventitia of vessels and no appreciable difference in all groups. We find that ICG concentration within solder is most important factor affecting both tissue temperature and tensile strength during laser vessel welding. The optimal balance between stronger strength and minimal thermal injury of vessel may be achieved primarily by using PD 56.7 W/cm2 at 0.01 mM ICG within solder during LAVA.

  7. Treatment of superficial vascular lesions with the KTP 532-nm laser: experience with 647 patients.

    PubMed

    Becher, G L; Cameron, H; Moseley, H

    2014-01-01

    Superficial vascular lesions are a common dermatological diagnosis but are often difficult to treat. Numerous lasers (especially the dye laser) and intense pulsed light sources have been used, but there have been very few reports on the effectiveness of the potassium-titanyl phosphate (KTP) laser. We have extensive experience of this modality at our institution, and the purpose of this survey is to report on the safety and efficacy of the KTP laser. Using an in-house database, we retrospectively collected data from patients who had undergone treatment with the KTP laser for superficial vascular lesions. Patients of Fitzpatrick skin type I-IV were included. Exclusion criteria were Fitzpatrick skin type V, patients with obvious suntan and those on potentially phototoxic medications or minocycline therapy. Diagnoses included discrete or matted telangiectasia, strawberry naevus, spider angioma, rosaceal erythema, rosaceal telangiectasia, telangiectatic naevus, angioma, combined rosaceal erythema/telangiectasia, port-wine stain, venous lake haemangioma and hereditary haemorrhagic telangiectasia. Patients underwent an initial test treatment and further treatment at 6-week intervals as required. Clinical photographs were taken pre- and post-treatment, and outcome was graded by patient and physician. Adverse effects were recorded including scarring, hypo- or hyperpigmentation, marked swelling, blistering, scabbing and bruising. Six hundred forty-seven patients with 13 diagnoses on 9 different body sites were recorded. Four hundred eighty-six were female, and the median age was 39.5 years. Of the lesions treated, 33.7 % (n = 218) were discrete telangiectases and 31.8 % (n = 206) were spider angiomas. A 92.7 % of lesions were on the face. Four hundred thirteen (77.6 %) patients who had outcomes recorded at 6 weeks were graded as "clearance" or "marked improvement". Only 38 (5.8 %) patients experienced adverse effects, all of which were minor; the main adverse

  8. Tissue-specific metabolite profiling of Cyperus rotundus L. rhizomes and (+)-nootkatone quantitation by laser microdissection, ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry, and gas chromatography-mass spectrometry techniques.

    PubMed

    Jaiswal, Yogini; Liang, Zhitao; Guo, Ping; Ho, Hing-Man; Chen, Hubiao; Zhao, Zhongzhen

    2014-07-23

    Cyperus rotundus L. is a plant species commonly found in both India and China. The caused destruction of this plant is of critical concern for agricultural produce. Nevertheless, it can serve as a potential source of the commercially important sesquiterpenoid (+)-nootkatone. The present work describes comparative metabolite profiling and (+)-nootkatone content determination in rhizome samples collected from these two countries. Laser dissected tissues, namely, the cortex, hypodermal fiber bundles, endodermis, amphivasal vascular bundles, and whole rhizomes were analyzed by ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF MS). Gas chromatography-mass spectrometry (GC-MS) analysis was used for profiling of essential oil constituents and quantitation of (+)-nootkatone. The content of (+)-nootkatone was found to be higher in samples from India (30.47 μg/10 g) compared to samples from China (21.72 μg/10 g). The method was validated as per International Conference on Harmonisation (ICH) guidelines (Q2 R1). The results from this study can be applied for quality control and efficient utilization of this terpenoid-rich plant for several applications in food-based industries. PMID:24938835

  9. Krypton laser photocoagulation induces retinal vascular remodeling rather than choroidal neovascularization.

    PubMed

    Behar-Cohen, F; Benezra, D; Soubrane, G; Jonet, L; Jeanny, J C

    2006-08-01

    The purpose of this study is to analyze the retina and choroid response following krypton laser photocoagulation. Ninety-two C57BL6/Sev129 and 32 C57BL/6J, 5-6-week-old mice received one single krypton (630 nm) laser lesion: 50 microm, 0.05 s, 400 mW. On the following day, every day thereafter for 1 week and every 2-3 days for the following 3 weeks, serial sections throughout the lesion were systematically collected and studied. Immunohistology using specific markers or antibodies for glial fibrillary acidic protein (GFAP) (astrocytes, glia and Muller's cells), von Willebrand (vW) (vascular endothelial cells), TUNEL (cells undergoing caspase dependent apoptosis), PCNA (proliferating cell nuclear antigen) p36, CD4 and F4/80 (infiltrating inflammatory and T cells), DAPI (cell nuclei) and routine histology were carried out. Laser confocal microscopy was also performed on flat mounts. Temporal and spatial observations of the created photocoagulation lesions demonstrate that, after a few hours, activated glial cells within the retinal path of the laser beam express GFAP. After 48 h, GFAP-positive staining was also detected within the choroid lesion center. "Movement" of this GFAP-positive expression towards the lasered choroid was preceded by a well-demarcated and localized apoptosis of the retina outer nuclear layer cells within the laser beam path. Later, death of retinal outer nuclear cells and layer thinning at this site was followed by evagination of the inner nuclear retinal layer. Funneling of the entire inner nuclear and the thinned outer nuclear layers into the choroid lesion center was accompanied by "dragging" of the retinal capillaries. Thus, from days 10 to 14 after krypton laser photocoagulation onward, well-formed blood capillaries (of retinal origin) were observed within the lesion. Only a few of the vW-positive capillary endothelial cells stained also for PCNA p36. In the choroid, dilatation of the vascular bed occurred at the vicinity of the

  10. Micro-vascular shape-memory polymer actuators with complex geometries obtained by laser stereolithography

    NASA Astrophysics Data System (ADS)

    Díaz Lantada, Andrés; de Blas Romero, Adrián; Chacón Tanarro, Enrique

    2016-06-01

    In our work we present the complete development process of geometrically complex micro-vascular shape-memory polymer actuators. The complex geometries and three-dimensional networks are designed by means of computer aided design resources. Manufacture is accomplished, in a single step, by means of laser stereolithography, directly from the computer-aided design files with the three dimensional geometries of the different actuators under development. To our knowledge, laser stereolithography is applied here for the first time to the development of shape memory polymer devices with complex geometries and inner micro-vasculatures for their activation using a thermal fluid. Final testing of the developed actuators helps to validate the approach and to put forward some present challenges.

  11. Confocal laser endomicroscopy and immunoendoscopy for real-time assessment of vascularization in gastrointestinal malignancies.

    PubMed

    Gheonea, Dan Ionuţ; Cârţână, Tatiana; Ciurea, Tudorel; Popescu, Carmen; Bădărău, Anca; Săftoiu, Adrian

    2011-01-01

    Gastrointestinal cancers represent a major cause of morbidity and mortality, with incomplete response to chemotherapy in the advanced stages and poor prognosis. Angiogenesis plays a crucial part in tumor growth and metastasis, with most gastrointestinal cancers depending strictly on the development of a new and devoted capillary network. Confocal laser endomicroscopy is a new technology which allows in vivo microscopic analysis of the gastrointestinal mucosa and its microvascularization during ongoing endoscopy by using topically or systemically administered contrast agents. Targeting markers of angiogenesis in association with confocal laser endomicroscopic examination (immunoendoscopy), as a future challenge, will add functional analysis to the morphological aspect of the neoplastic process. This review describes previous experience in endomicroscopic examination of the upper and lower digestive tract with emphasis on vascularization, resulting in a broad spectrum of potential clinical applications, and also preclinical research that could be translated to human studies.

  12. The influence of immunohistochemistry on mRNA recovery from microdissected frozen and formalin-fixed, paraffin-embedded sections.

    PubMed

    Gjerdrum, Lise Mette; Abrahamsen, Helene N; Villegas, Berta; Sorensen, Boe S; Schmidt, Henrik; Hamilton-Dutoit, Stephen J

    2004-12-01

    Laser-assisted microdissection (LAM) is now widely used to obtain specific cell populations from heterogeneous tissues. A major disadvantage of LAM is poor tissue morphology during microscopy, in part because coverslips are not used. Immunohistochemical labeling can improve identification of target cells but may affect the subsequent analysis of the microdissected tissue. We studied the effect of immunohistochemistry (IHC) on mRNA recovery from labeled cells after microdissection from both frozen and formalin-fixed, paraffin-embedded (FFPE) sections, using Melan-A and Ki-67 staining in lymph nodes with metastatic melanoma as a model. We developed rapid protocols for immunostaining in an attempt to limit loss of mRNA during procedures. A sensitive real-time quantitative reverse transcription-PCR was used to measure mRNA. We found a marked decrease in the mRNA yield from 500 microdissected cells from frozen and paraffin sections after immunostaining for both markers. Recovery of mRNA decreased by up to 89%, comparing the immunostained with the routinely stained sections. Interestingly, the ratio between mRNA for the two markers was similar in all stains, indicating that immunostained sections may be used for mRNA analysis. We also investigated the effect of storing membrane-mounted sections for microdissection under different conditions. Slides mounted with paraffin sections could be stored at room temperature for up to 90 days with no significant decrease in mRNA recovery.

  13. Quantitative gene expression analysis in microdissected archival formalin-fixed and paraffin-embedded tumor tissue.

    PubMed

    Specht, K; Richter, T; Müller, U; Walch, A; Werner, M; Höfler, H

    2001-02-01

    Formalin-fixed, paraffin-embedded tissue is the most widely available material for retrospective clinical studies. In combination with the potential of genomics, these tissues represent an invaluable resource for the elucidation of disease mechanisms and validation of differentially expressed genes as novel therapeutic targets or prognostic indicators. We describe here an approach that, in combination with laser-assisted microdissection allows quantitative gene expression analysis in formalin-fixed, paraffin-embedded archival tissue. Using an optimized RNA microscale extraction procedure in conjunction with real-time quantitative reverse transcriptase-polymerase chain reaction based on fluorogenic TaqMan methodology, we analyzed the expression of a panel of cancer-relevant genes, EGF-R, HER-2/neu, FGF-R4, p21/WAF1/Cip1, MDM2, and HPRT and PGK as controls. We demonstrate that expression level determinations from formalin-fixed, paraffin-embedded tissues are accurate and reproducible. Measurements were comparable to those obtained with matching fresh-frozen tissue and neither fixation grade nor time significantly affected the results. Laser microdissection studies with 5-microm thick sections and defined numbers of tumor cells demonstrated that reproducible quantitation of specific mRNAs can be achieved with only 50 cells. We applied our approach to HER-2/neu quantitative gene expression analysis in 54 microdissected tumor and nonneoplastic archival samples from patients with Barrett's esophageal adenocarcinoma and showed that the results matched those obtained in parallel by fluorescence in situ hybridization and immunohistochemistry. Thus, the combination of laser-assisted microdissection and real-time TaqMan reverse transcriptase-polymerase chain reaction opens new avenues for the investigation and clinical validation of gene expression changes in archival tissue specimens.

  14. Real-time measurements of endogenous CO production from vascular cells using an ultrasensitive laser sensor

    NASA Technical Reports Server (NTRS)

    Morimoto, Y.; Durante, W.; Lancaster, D. G.; Klattenhoff, J.; Tittel, F. K.

    2001-01-01

    Carbon monoxide (CO) has been implicated as a biological messenger molecule analogous to nitric oxide. A compact gas sensor based on a midinfrared laser absorption spectroscopy was developed for direct and real-time measurement of trace levels (in approximate pmol) of CO release by vascular cells. The midinfrared light is generated by difference frequency mixing of two nearinfrared lasers in a nonlinear optical crystal. A strong infrared absorption line of CO (4.61 microm) is chosen for convenient CO detection without interference from other gas species. The generation of CO from cultured vascular smooth muscle cells was detected every 20 s without any chemical modification to the CO. The sensitivity of the sensor reached 6.9 pmol CO. CO synthesis was measured from untreated control cells (0.25 nmol per 10(7) cells/h), sodium nitroprusside-treated cells (0.29 nmol per 10(7) cells/h), and hemin-treated cells (0.49 nmol per 10(7) cells/h). The sensor also detected decreases in CO production after the addition of the heme oxygenase (HO) inhibitor tin protoporphyrin-IX (from 0.49 to 0.02 nmol per 10(7) cells/h) and increases after the administration of the HO substrate hemin (from 0.27 to 0.64 nmol per 10(7) cells/h). These results demonstrate that midinfrared laser absorption spectroscopy is a useful technique for the noninvasive and real-time detection of trace levels of CO from biological tissues.

  15. Dermoscopy, confocal laser microscopy, and hi-tech evaluation of vascular skin lesions: diagnostic and therapeutic perspectives.

    PubMed

    Grazzini, Marta; Stanganelli, Ignazio; Rossari, Susanna; Gori, Alessia; Oranges, Teresa; Longo, Anna Sara; Lotti, Torello; Bencini, Pier Luca; De Giorgi, Vincenzo

    2012-01-01

    Vascular skin lesions comprise a wide and heterogeneous group of malformations and tumors that can be correctly diagnosed based on natural history and physical examination. However, considering the high incidence of such lesions, a great number of them can be misdiagnosed. In addition, it is not so rare that an aggressive amelanotic melanoma can be misdiagnosed as a vascular lesion. In this regard, dermoscopy and confocal laser microscopy examination can play a central role in increasing the specificity of the diagnosis of such lesions. In fact, the superiority of these tools over clinical examination has encouraged dermatologists to adopt these devices for routine clinical practice, with a progressive spread of their use. In this review, we will go through the dermoscopic and the confocal laser microscopy of diagnosis of most frequent vascular lesions (i.e., hemangiomas angiokeratoma, pyogenic granuloma, angiosarcoma) taking into particular consideration the differential diagnosis with amelanotic melanoma. PMID:22950556

  16. Micro-PIV quantification of capillary blood flow redistribution caused by laser-assisted vascular occlusion

    NASA Astrophysics Data System (ADS)

    Kurochkin, Maxim A.; Stiukhina, Elena S.; Fedosov, Ivan V.; Postnov, Dmitry E.; Tuchin, Valery V.

    2016-04-01

    We propose μPIV-based technique for quantitative assessment of blood flow redistribution in microcirculatory networks. Our approach is based on per-segment averaging of measured quantities so we can avoid most of problems that are typical for point-wise measurements. The key point of our technique is the digital processing algorithms of recorded data that include: capillary network axial line construction; interrogation regions centering; blood flow velocity local estimate using PIV approach; blood flow velocity calculation by means of averaging over entire vessel segment; the calculation of blood volume flow rate map. We illustrate the application of developed technique with in vivo measurements and blood flow velocity map reconstruction for chorioallantoic membrane (CAM) of chicken embryo, in which the local vascular occlusion was produced using continuous wave laser light irradiation..

  17. A meta-analysis of aneurysm formation in laser assisted vascular anastomosis (LAVA)

    NASA Astrophysics Data System (ADS)

    Chen, Chen; Peng, Fei; Xu, Dahai; Cheng, Qinghua

    2009-08-01

    Laser assisted vascular anastomosis (LAVA) is looked as a particularly promising non-suture method in future. However, aneurysm formation is one of the main reasons delay the clinical application of LAVA. Some scientists investigated the incidence of aneurysms in animal model. To systematically analyze the literature on reported incidence of aneurysm formation in LAVA therapy, we performed a meta-analysis comparing LAVA with conventional suture anastomosis (CSA) in animal model. Data were systematically retrieved and selected from PUBMED. In total, 23 studies were retrieved. 18 studies were excluded, and 5 studies involving 647 animals were included. Analysis suggested no statistically significant difference between LAVA and CSA (OR 1.24, 95%CI 0.66-2.32, P=0.51). Result of meta analysis shows that the technology of LAVA is very close to clinical application.

  18. Comparison of NIRS, laser Doppler flowmetry, photoplethysmography, and pulse oximetry during vascular occlusion challenges.

    PubMed

    Abay, T Y; Kyriacou, P A

    2016-04-01

    Monitoring changes in blood volume, blood flow, and oxygenation in tissues is of vital importance in fields such as reconstructive surgery and trauma medicine. Near infrared spectroscopy (NIRS), laser Doppler (LDF) flowmetry, photoplethysmography (PPG), and pulse oximetry (PO) contribute to such fields due to their safe and noninvasive nature. However, the techniques have been rarely investigated simultaneously or altogether. The aim of this study was to investigate all the techniques simultaneously on healthy subjects during vascular occlusion challenges. Sensors were attached on the forearm (NIRS and LDF) and fingers (PPG and PO) of 19 healthy volunteers. Different degrees of vascular occlusion were induced by inflating a pressure cuff on the upper arm. The responses of tissue oxygenation index (NIRS), tissue haemoglobin index (NIRS), flux (LDF), perfusion index (PPG), and arterial oxygen saturation (PO) have been recorded and analyzed. Moreover, the optical densities were calculated from slow varying dc PPG, in order to distinguish changes in venous blood volumes. The indexes showed significant changes (p  <  0.05) in almost all occlusions, either venous or over-systolic occlusions. However, differentiation between venous and arterial occlusion by LDF may be challenging and the perfusion index (PI) may not be adequate to indicate venous occlusions. Optical densities may be an additional tool to detect venous occlusions by PPG.

  19. In vivo argon laser vascular welding using thermal feedback: open and closed loop patency and collagen crosslinking

    SciTech Connect

    Small, W., LLNL

    1997-02-28

    An in vivo study of vascular welding with a fiber-delivered argon laser was conducted using a canine model. Longitudinal arteriotomies and venotomies were treated on femoral vein and artery. Laser energy was delivered to the vessel wall via a 400 {micro}m optical fiber. The surface temperature at the center of the laser spot was monitored in real time using a hollow glass optical fiber-based two-color infrared thermometer. The surface temperature was limited by either a room-temperature saline drip or direct feedback control of the laser using a mechanical shutter to alternately pass and block the laser. Acute patency was evaluated either visually (leak/no leak) or by in vivo burst pressure measurements. Biochemical assays were performed to investigate the possible laser-induced formation or destruction of enzymatically mediated covalent crosslinks between collagen molecules. Viable welds were created both with and without the use of feedback control. Tissues maintained at 50 C using feedback control had an elevated crosslink count compared to controls, while those irradiated without feedback control experienced a decrease. Differences between the volumetric heating associated with open and closed loop protocols may account for the different effects on collagen crosslinks. Covalent mechanisms may play a role in argon laser vascular fusion.

  20. Different imaging methods in the comparative assessment of vascular lesions: color-coded duplex sonography, laser Doppler perfusion imaging, and infrared thermography

    NASA Astrophysics Data System (ADS)

    Urban, Peter; Philipp, Carsten M.; Weinberg, Lutz; Berlien, Hans-Peter

    1997-12-01

    Aim of the study was the comparative investigation of cutaneous and subcutaneous vascular lesions. By means of color coded duplex sonography (CCDS), laser doppler perfusion imaging (LDPI) and infrared thermography (IT) we examined hemangiomas, vascular malformations and portwine stains to get some evidence about depth, perfusion and vascularity. LDI is a helpful method to get an impression of the capillary part of vascular lesions and the course of superficial vessels. CCDS has disadvantages in the superficial perfusion's detection but connections to deeper vascularizations can be examined precisely, in some cases it is the only method for visualizing vascular malformations. IT gives additive hints on low blood flow areas or indicates arterial-venous-shunts. Only the combination of all imaging methods allows a complete assessment, not only for planning but also for controlling the laser treatment of vascular lesions.

  1. Proteomic analysis of microdissected facial nuclei of the rat following facial nerve injury.

    PubMed

    Melle, Christian; Ernst, Günther; Grosheva, Maria; Angelov, Doychin N; Irintchev, Andrey; Guntinas-Lichius, Orlando; von Eggeling, Ferdinand

    2009-12-15

    Recent studies using molecular and genetic techniques just have started to elucidate the complex process that drives successful peripheral nerve regeneration. Introducing proteomics to this field, we unilaterally performed a facial nerve axotomy in 13 adult Wistar rats. Seven days later, a total of 40 20-microm coronary cryostat sections of the operated and contralateral unoperated nucleus facialis were microdissected. On the one hand, microdissected areas were pooled for each side, lysed and applied to ProteinChip Arrays. On the other hand, one microdissected area from the right and left facial nucleus each was directly placed on the affinity chromatographic array. Facial motoneurons were lysed in situ and released their proteins to spatially defined points. 215 laser addressable distinct positions across the surface of the spot enabled a high spatial resolution of measured protein profiles for the analysed tissue area. Protein profiles of the single positions were plotted over the used tissue section to visualize their distribution. The comparative analysis of the protein lysates from operated and normal nuclei facialis revealed, for both approaches used, differentially expressed proteins. Although by direct application of one cryostat section only a few hundred motoneurons were analysed, results comparable to these using lysates were obtained. Additionally, the applied technique revealed differences in the intensity distribution of several proteins of unknown function in the lesioned in comparison to the contralateral normal facial nucleus. This proteomic analysis with ultra high sensitivity paired with potential for a spatial resolution is a promising methodology for peripheral nerve regeneration studies. PMID:19748522

  2. Necessity of Microdissecting Different Tumor Components in Pulmonary Tumor Pyrosequencing

    PubMed Central

    Zheng, Zhong; Shen, Shanxiang; Smith, Prudence; Khalil, Farah K.

    2016-01-01

    Microdissection is a useful method in tissue sampling prior to molecular testing. Tumor heterogeneity imposes new challenges for tissue sampling. Different microdissecting methods have been employed in face of such challenge. We improved our microdissection method by separately microdissecting the morphologically different tumor components. This improvement helped the pyrosequencing data analysis of two specimens. One specimen consisted of both adenocarcinoma and neuroendocrine components. When both tumor components were sequenced together for KRAS (Kirsten rat sarcoma viral oncogene homolog) gene mutations, the resulting pyrogram indicated that it was not a wild type, suggesting that it contained KRAS mutation. However, the pyrogram did not match any KRAS mutations and a conclusion could not be reached. After microdissecting and testing the adenocarcinoma and neuroendocrine components separately, it was found that the adenocarcinoma was positive for KRAS G12C mutation and the neuroendocrine component was positive for KRAS G12D mutation. The second specimen consisted of two morphologically different tumor nodules. When microdissected and sequenced separately, one nodule was positive for BRAF (v-raf murine sarcoma viral oncogene homolog B1) V600E and the other nodule was wild type at the BRAF codon 600. These examples demonstrate that it is necessary to microdissect morphologically different tumor components for pyrosequencing.

  3. Necessity of Microdissecting Different Tumor Components in Pulmonary Tumor Pyrosequencing

    PubMed Central

    Zheng, Zhong; Shen, Shanxiang; Smith, Prudence; Khalil, Farah K.

    2016-01-01

    Microdissection is a useful method in tissue sampling prior to molecular testing. Tumor heterogeneity imposes new challenges for tissue sampling. Different microdissecting methods have been employed in face of such challenge. We improved our microdissection method by separately microdissecting the morphologically different tumor components. This improvement helped the pyrosequencing data analysis of two specimens. One specimen consisted of both adenocarcinoma and neuroendocrine components. When both tumor components were sequenced together for KRAS (Kirsten rat sarcoma viral oncogene homolog) gene mutations, the resulting pyrogram indicated that it was not a wild type, suggesting that it contained KRAS mutation. However, the pyrogram did not match any KRAS mutations and a conclusion could not be reached. After microdissecting and testing the adenocarcinoma and neuroendocrine components separately, it was found that the adenocarcinoma was positive for KRAS G12C mutation and the neuroendocrine component was positive for KRAS G12D mutation. The second specimen consisted of two morphologically different tumor nodules. When microdissected and sequenced separately, one nodule was positive for BRAF (v-raf murine sarcoma viral oncogene homolog B1) V600E and the other nodule was wild type at the BRAF codon 600. These examples demonstrate that it is necessary to microdissect morphologically different tumor components for pyrosequencing. PMID:27597976

  4. Necessity of Microdissecting Different Tumor Components in Pulmonary Tumor Pyrosequencing.

    PubMed

    Qin, Dahui; Zheng, Zhong; Shen, Shanxiang; Smith, Prudence; Khalil, Farah K

    2016-01-01

    Microdissection is a useful method in tissue sampling prior to molecular testing. Tumor heterogeneity imposes new challenges for tissue sampling. Different microdissecting methods have been employed in face of such challenge. We improved our microdissection method by separately microdissecting the morphologically different tumor components. This improvement helped the pyrosequencing data analysis of two specimens. One specimen consisted of both adenocarcinoma and neuroendocrine components. When both tumor components were sequenced together for KRAS (Kirsten rat sarcoma viral oncogene homolog) gene mutations, the resulting pyrogram indicated that it was not a wild type, suggesting that it contained KRAS mutation. However, the pyrogram did not match any KRAS mutations and a conclusion could not be reached. After microdissecting and testing the adenocarcinoma and neuroendocrine components separately, it was found that the adenocarcinoma was positive for KRAS G12C mutation and the neuroendocrine component was positive for KRAS G12D mutation. The second specimen consisted of two morphologically different tumor nodules. When microdissected and sequenced separately, one nodule was positive for BRAF (v-raf murine sarcoma viral oncogene homolog B1) V600E and the other nodule was wild type at the BRAF codon 600. These examples demonstrate that it is necessary to microdissect morphologically different tumor components for pyrosequencing. PMID:27597976

  5. 808-nm diode lasers with and without exogenous chromophore in the treatment of benign facial pigmented and vascular lesions

    NASA Astrophysics Data System (ADS)

    Marangoni, Ovidio; Magaton Rizzi, G.; Trevisan, G.

    2001-10-01

    Aim: To evaluate the safety and efficacy of an 808 nm diode laser for the treatment of benign facial pigmented and vascular lesions, with and without the use of an exogenous chromophore. Method: Thirty-eight patients were treated with an 808 nm diode laser (Eufoton, Italy), in some cases using a chromophore (1% methylene blue, SALF, Bergamo). Pigmented lesions: 21 patients, (15 pigmented keratoses, 6 melanoses). All the lesions were evaluated by dermatoscopy (Videocap 200, DS Medica, Italy) before the treatment. Fluence levels were 10 - 30 J/cmq; pulse lengths were 10 - 50 ms; spot size was 2 mm. Five hypopigmented keratoses were artificially pigmented using exogenous chromophore. Two melanoses required an additional laser session. Vascular lesions: 17 patients, (12 small angiomas, 5 teleangectasias). Fluences were 50 - 100 J/cmq; pulse lengths were 10 - 50 ms; spot size was 2 mm. Eight angiomas were pigmented with exogenous chromophore prior to the treatment. In all cases the areas surrounding the lesions were cooled. The patients were followed at 1, 4 and 8 weeks after the procedure. Results: The keratoses healed completely within two weeks. Four melanoses healed after four weeks. In the two melanoses that were re-treated after eight weeks there remained slight hypopigmentation of the area. All the vascular lesions healed after 15 days without any residual scarring. Considerations: The use of the 808 nm diode laser in the treatment of benign facial pigmented and vascular lesions appears to be justified on the grounds of efficacy and safety of the device, and good degree of acceptance by the patients. By increasing absorption of the 808 nm beam and reducing its penetration, the pigmentation of superficial lesions with exogenous chromophore allowed us to decrease fluences and reduce irradiation times.

  6. Laser Doppler flowmetry: in the assessment of peripheral vascular disorders? A preliminary evaluation.

    PubMed

    Cochrane, T; Sheriff, S B; Boulton, A J; Ward, J D; Atkins, R M

    1986-02-01

    Skin is a representative microvascular bed providing easy access for the study of capillary haemodynamics. In this preliminary evaluation we have used laser Doppler flowmetry to measure the response of skin capillary blood flow to local thermal stimulation. The purpose of our study was to see if such measurements can be used to derive useful clinical information. Thirty subjects, 15 male and 15 female, were selected to establish "normal' patterns of behaviour. All normal subjects studied showed similar patterns of response. There was a slight downward trend in red-blood-cell flux with age. Periods of vasomotor activity were present at rates ranging from 4-17 cycles/min. The mean temperature rise in the heated skin area was 4 degrees C. Blood flow over this range increased by a factor of 10 or more. Clinical cases studied were strictly limited to three categories of disease with known or suspected microvascular changes: algodystrophy, lower-limb ischaemia and diabetic neuropathy. All the clinical cases studied showed deviations from the normal pattern. Patients with algodystrophy had impaired peripheral vascular function in the affected area. Patients with lower-limb ischaemic problems all had poor peripheral perfusion. In the limited number of diabetic neuropathic feet studied two patterns emerged: patients with no evidence of ulceration displayed striking vasomotor activity; those with more severe problems had reduced red-blood-cell flux and little evidence of vasomotor control.

  7. Nanotechnology and vascular neurosurgery: an in vivo experimental study on microvessels repair using laser photoactivation of a nanostructured hyaluronan solder.

    PubMed

    Esposito, G; Rossi, F; Matteini, P; Ratto, F; Sabatino, G; Puca, A; Albanese, A; Rossi, G; Marchese, E; Maira, G; Pini, R

    2012-01-01

    Sealing tissues by laser in neurosurgical procedures may overcome problems related to the use of conventional suturing methods which can be associated with various degrees of vascular wall damage. Despite the significant experimental and clinical achievements of the past, a standardized clinical application of laser-welding technology has not yet been implemented. The main problem is related to the use of common organic chromophores. A substantial breakthrough in the laser welding of biological tissues may come from the advent of nanotechnologies. In this paper we describe an experimental study, to confirm the feasibility of an innovative laser-assisted vascular repair (LAVR) technique based on diode laser irradiation and subsequent photoactivation of a hyaluronan solder embedded with near infrared (NIR) absorbing gold nanorods (GNRs), and to analyze the induced closuring effect in a follow-up study performed in animal model. Twenty New Zealand rabbits underwent closure of a 3-mm longitudinal incision performed on the common carotid artery (CCA) by means of 810 nm diode laser irradiation, in conjunction with the topical application of an optimized GNR composite. Effective closure of the arterial wound was accomplished by using very low laser intensity (30 W/cm2). The average CCA occlusion time was as low as 50 sec. Animals underwent different follow-up periods (2, 8, 30 days). After follow-up, they were re-anesthetized, the patency of the treated vessels was tested (Doppler analysis) and then the irradiated vessels were excised and subjected to histological evaluations. Morphological examinations of the samples documented the integrity of the vascular wall. No host reaction to nanoparticles occurred. Collagen and elastic fibers returned to their normal architecture 30 days after treatment. A Scanning Electron Microscopy (SEM) examination and immuno-histochemical analysis demonstrated a full re-endothelization of the vessel walls. We thus confirmed that a laser

  8. Non-invasive technique for assessment of vascular wall stiffness using laser Doppler vibrometry

    NASA Astrophysics Data System (ADS)

    Campo, Adriaan; Segers, Patrick; Heuten, Hilde; Goovaerts, Inge; Ennekens, Guy; Vrints, Christiaan; Baets, Roel; Dirckx, Joris

    2014-06-01

    It has been shown that in cardiovascular risk management, stiffness of large arteries has a very good predictive value for cardiovascular disease and mortality. This parameter is best known when estimated from the pulse wave velocity (PWV) measured between the common carotid artery (CCA) in the neck and femoral artery in the groin, but may also be determined locally from short-distance measurements on a short vessel segment. In this work, we propose a novel, non-invasive, non-contact laser Doppler vibrometry (LDV) technique for evaluating PWV locally in an elastic vessel. First, the method was evaluated in a phantom setup using LDV and a reference method. Values correlated significantly between methods (R ≤ 0.973 (p ≤ 0.01)); and a Bland-Altman analysis indicated that the mean bias was reasonably small (mean bias ≤ -2.33 ms). Additionally, PWV was measured locally on the skin surface of the CCA in 14 young healthy volunteers. As a preliminary validation, PWV measured on two locations along the same artery was compared. Local PWV was found to be between 3 and 20 m s-1, which is in line with the literature (PWV = 5-13 m s-1). PWV assessed on two different locations on the same artery correlated significantly (R = 0.684 (p < 0.01)). In summary, we conclude that this new non-contact method is a promising technique to measure local vascular stiffness in a fully non-invasive way, providing new opportunities for clinical diagnosing.

  9. Microdissection and microcloning of mid-chromosome 4: Genetic mapping of 41 microdissection clones

    SciTech Connect

    Bahary, N.; McGraw, D.E.; Shilling, R.; Friedman, J.M. )

    1993-04-01

    Available genetic information places the mouse db (diabetes) gene approximately 5 cM distal to Ifa on mid/distal mouse chromosome 4. These data have indicated that there is a relevant paucity of genetic markers that map to this region of chromosome 4. To increase the density of the genetic map on mid-chromosome 4, the authors have applied the techniques of microdissection and microcloning of the mid-portion of mouse chromosome 4. A total of 47 RFLPs from the microdissection library were used to type the progeny of three C57BL/6J Mus spretus backcrosses. The resulting composite genetic map positions seven known genes, 41 microclones, and three other anonymous markers to a region of approximately 21 cM on mid-chromosome 4 extending from b to Lck. The density of markers in this region of chromosome 4 should be sufficient to initiate the physical mapping of this subchromosomal segment, facilitating efforts to clone the db gene, as well as other uncloned mutant loci in this region of chromosome 4. 30 refs., 3 figs., 1 tab.

  10. Improved laser-assisted vascular tissue fusion using solder-doped polymer membranes on a canine model

    NASA Astrophysics Data System (ADS)

    McNally-Heintzelman, Karen M.; Sorg, Brian S.; Hammer, Daniel X.; Heintzelman, Douglas L.; Hodges, Diane E.; Welch, Ashley J.

    2000-05-01

    Newly developed light-activated surgical adhesives have been investigated as a substitute to traditional protein solders for vascular tissue fusion without the need for sutures. Canine femoral arteries (n equals 14), femoral veins (n equals 14) and carotid arteries (n equals 10) were exposed, and a 0.3 to 0.6 cm longitudinal incision was made in the vessel walls. The surgical adhesive, composed of a poly(L-lactic-co-glycolic acid) scaffold doped with the traditional protein solder mix of bovine serum albumin and indocyanine green dye, was used to close the incisions in conjunction with an 805 nm diode laser. Blood flow was restored to the vessels immediately after the procedure and the incision sites were checked for patency. The new adhesives were flexible enough to be wrapped around the vessels while their solid nature avoided the problems associated with 'runaway' of the less viscous liquid protein solders widely used by researchers. Assessment parameters included measurement of the ex vivo intraluminal bursting pressure one to two hours after surgery, as well as histology. The acute intraluminal bursting pressures were significantly higher in the laser-solder group (greater than 300 mmHg) compared to the suture control group (less than 150 mmHg) where four evenly spaced sutures were used to repair the vessel (n equals 4). Histological analysis showed negligible evidence of collateral thermal damage to the underlying tissue in the laser-solder repair group. These initial results indicated that laser-assisted vascular repair using the new adhesives is safe, easy to perform, and contrary to conventional suturing, provides an immediate leak-free closure. In addition, the flexible and moldable nature of the new adhesives should allow them to be tailored to a wide range of tissue geometries, thus greatly improving the clinical applicability of laser-assisted tissue repair.

  11. Laser capture.

    PubMed

    Potter, S Steven; Brunskill, Eric W

    2012-01-01

    This chapter describes detailed methods used for laser capture microdissection (LCM) of discrete subpopulations of cells. Topics covered include preparing tissue blocks, cryostat sectioning, processing slides, performing the LCM, and purification of RNA from LCM samples. Notes describe the fine points of each operation, which can often mean the difference between success and failure. PMID:22639264

  12. The use of Intravenous Laser Blood Irradiation (ILBI) at 630–640 nm to prevent vascular diseases and to increase life expectancy

    PubMed Central

    2015-01-01

    Background and Aims: The mortality rate from vascular diseases is one of the highest. The use of Intravenous Laser Blood Irradiation (ILBI) within the last 30 years has demonstrated high efficacy in the treatment of vascular, cardiac and other systemic diseases. Rationale: Laser energy at 630-640 nanometers is arguably the most effective for irradiation of blood and the vascular wall. Photons at this wavelength are absorbed by oxygen, improve microcirculation, can change the viscosity of the blood and affect vascular endothelium. Conclusions: In summary, more than 25 years of experience of using laser energy at 630-640 nm has shown that this waveband directly influences the parameters of all cells in the blood, blood plasma, the coagulation process and all the structural components of the vascular wall. Additionally, ILBI directly or indirectly affects the cells of the immune system, hormones, and exchange processes in an organism, thereby not only improving the function of the vascular system, but also the other systems of an organism. It can finally lead to lower the incidence and number of vascular diseases, and indirectly to the reduction of the number of diseases in other organs and even systemically, thus helping to prolong the lifespan. PMID:25941421

  13. Detection of biogenic CO production above vascular cell cultures using a near-room-temperature QC-DFB laser

    NASA Technical Reports Server (NTRS)

    Kosterev, A. A.; Tittel, F. K.; Durante, W.; Allen, M.; Kohler, R.; Gmachl, C.; Capasso, F.; Sivco, D. L.; Cho, A. Y.

    2002-01-01

    We report the first application of pulsed, near-room-temperature quantum cascade laser technology to the continuous detection of biogenic CO production rates above viable cultures of vascular smooth muscle cells. A computer-controlled sequence of measurements over a 9-h period was obtained, resulting in a minimum detectable CO production of 20 ppb in a 1-m optical path above a standard cell-culture flask. Data-processing procedures for real-time monitoring of both biogenic and ambient atmospheric CO concentrations are described.

  14. Real-time ultrasonography as a monitoring technique for interstitial Nd:YAG laser treatment of voluminous hemangiomas and vascular malformations

    NASA Astrophysics Data System (ADS)

    Werner, Jochen A.; Gottschlich, Stefan; Lippert, Burkard M.; Folz, Benedikt J.

    1998-01-01

    Voluminous vascular anomalies of the head and neck region are still treated with conventional surgery although Neodymium:Yttrium-Aluminum-Garnet (Nd:YAG) laser therapy is an effective treatment method. One hundred thirty give patients with voluminous hemangiomas and vascular malformations were treated with interstitial Nd:YAG laser therapy, partly complemented by a non-contact mode Nd:YAG laser light application. The vascular tumors had a diameter of more than 3 cm in two or all three dimensions. Treatment was carried out under ultrasound and manual control. Nearly 60% of the patients showed a complete clinical regression of the vascular tumor, a third of the patients had a partial regression and were satisfied with the treatment outcome. Four patients were treated unsuccessfully with the laser and three of them subsequently underwent conventional surgery. Only 10 patients showed cosmetic and functional deficits. These results on the interstitial Nd:YAG laser therapy of voluminous hemangiomas and vascular malformations in a large patient group demonstrated the high effectiveness of this novel and innovative therapy modality.

  15. Acute effects of vascular modifying agents in solid tumors assessed by noninvasive laser Doppler flowmetry and near infrared spectroscopy.

    PubMed

    Kragh, Michael; Quistorff, Bjørn; Horsman, Michael R; Kristjansen, Paul E G

    2002-01-01

    The potential of noninvasive laser Doppler flowmetry (LDF) and near infrared spectroscopy (NIRS) to detect acute effects of different vascular-modifying agents on perfusion and blood volume in tumors was evaluated. C3H mouse mammary carcinomas (approximately 200 mm(3)) in the rear foot of CDF1 mice were treated with flavone acetic acid (FAA, 150 mg/kg), 5,6-dimethylxanthenone-4-acetic acid (DMXAA, 20 mg/kg), combretastatin A-4 disodium phosphate (CA4DP, 250 mg/kg), hydralazine (HDZ, 5 mg/kg), or nicotinamide (NTA, 500 mg/kg). Tumor perfusion before and after treatment was evaluated by noninvasive LDF, using a 41 degrees C heated custom-built LDF probe with four integrated laser/receiver units, and tumor blood volume was estimated by NIRS, using light guide coupled reflectance measurements at 800+/-10 nm. FAA, DMXAA, CA4DP, and HDZ significantly decreased tumor perfusion by 50%, 47%, 73%, and 78%, respectively. In addition, FAA, DMXAA, and HDZ significantly reduced the blood volume within the tumor, indicating that these compounds to some degree shunted blood from the tumor to adjacent tissue, HDZ being most potent. CA4DP caused no change in the tumor blood volume, indicating that the mechanism of action of CA4DP was vascular shut down with the blood pool trapped in the tumor. NTA caused no change in either tumor perfusion or tumor blood volume. We conclude that noninvasive LDF and NIRS can determine acute effects of vascular modifying agents on tumor perfusion and blood volume.

  16. [Selection of various types of lasers in the treatment of surface and deep vascular anomalies].

    PubMed

    Waldschmidt, J; Berlien, H P; Hauck, G W; el-Dessouky, M

    1988-02-01

    Laser therapy can be used to great advantage in the treatment of haemangiomas in childhood in specially selected cases. We were able to collect experiences in 101 children and to utilise both the coagulation effect for percutaneous induction of regression and the cutting effect in the resection of haemangiomas at the body surface and the thoracic and abdominal cavities. Among the commercially available laser equipment we would prefer the argon laser for intracutaneous lesions (naevi teleangiectatici, spider naevi, plane haemangiomas), whereas for the cavernous or planotuberous or tuberonodous haemangiomas it is better to use the neodym-YAG laser. Patients must be very carefully selected and selection must be restricted to haemangiomas with complications. Treatment via neodym-YAG laser must be complemented by protecting the skin against cold, by compressing the angiomas and by an appropriate after treatment.

  17. RGB imaging system for monitoring of skin vascular malformation's laser therapy

    NASA Astrophysics Data System (ADS)

    Jakovels, Dainis; Kuzmina, Ilona; Berzina, Anna; Spigulis, Janis

    2012-06-01

    A prototype RGB imaging system for mapping of skin chromophores consists of a commercial RGB CMOS sensor, RGB LEDs ring-light illuminator and orthogonally orientated polarizers for reducing specular reflectance. The system was used for monitoring of vascular malformations (hemagiomas and telangiectasias) therapy.

  18. Assessment of the Microbiota in Microdissected Tissues of Crohn's Disease Patients

    PubMed Central

    De Hertogh, Gert; Lemmens, Bart; Verhasselt, Peter; de Hoogt, Ronald; Sagaert, Xavier; Joossens, Marie; Van Assche, Gert; Rutgeerts, Paul; Vermeire, Severine; Aerssens, Jeroen

    2012-01-01

    The microbiota of the gastrointestinal tract is frequently mentioned as one of the key players in the etiopathogenesis of Crohn's disease (CD). Four hypotheses have been suggested: the single, still unknown bacterial pathogen, an abnormal overall composition of the bowel microbiota (“dysbiosis”), an abnormal immunological reaction to an essentially normally composed microbiota, and increased bacterial translocation. We propose that laser capture microdissection of selected microscopic structures, followed by broad-range 16S rRNA gene sequencing, is an excellent method to assess spatiotemporal alterations in the composition of the bowel microbiota in CD. Using this approach, we demonstrated significant changes of the composition, abundance, and location of the gut microbiome in this disease. Some of these abnormal findings persisted even after macroscopic mucosal healing. Further investigations along these lines may lead to a better understanding of the possible involvement of the bowel bacteria in the development of clinical Crohn's disease. PMID:22191064

  19. Analytical modeling of laser pulse heating of embedded biological targets: An application to cutaneous vascular lesions

    NASA Astrophysics Data System (ADS)

    Mirkov, Mirko; Sherr, Evan A.; Sierra, Rafael A.; Lloyd, Jenifer R.; Tanghetti, Emil

    2006-06-01

    Detailed understanding of the thermal processes in biological targets undergoing laser irradiation continues to be a challenging problem. For example, the contemporary pulsed dye laser (PDL) delivers a complex pulse format which presents specific challenges for theoretical understanding and further development. Numerical methods allow for adequate description of the thermal processes, but are lacking for clarifying the effects of the laser parameters. The purpose of this work is to derive a simplified analytical model that can guide the development of future laser designs. A mathematical model of heating and cooling processes in tissue is developed. Exact analytical solutions of the model are found when applied to specific temporal and spatial profiles of heat sources. Solutions are reduced to simple algebraic expressions. An algorithm is presented for approximating realistic cases of laser heating of skin structures by heat sources of the type found to have exact solutions. The simple algebraic expressions are used to provide insight into realistic laser irradiation cases. The model is compared with experiments on purpura threshold radiant exposure for PDL. These include data from four independent groups over a period of 20 years. Two of the data sets are taken from previously published articles. Two more data sets were collected from two groups of patients that were treated with two PDLs (585 and 595 nm) on normal buttocks skin. Laser pulse durations were varied between 0.5 and 40 ms radiant exposures were varied between 3 and 20 J/cm2. Treatment sites were evaluated 0.5, 1, and 24 hours later to determine purpuric threshold. The analytical model is in excellent agreement with a wide range of experimental data for purpura threshold radiant exposure. The data collected by independent research groups over the last 20 years with PDLs with wavelengths ranged from 577 to 595 nm were described accurately by this model. The simple analytical model provides an accurate

  20. [Changes in the blood rheological properties in the transcutaneous irradiation of the ulnar vascular fascicle with a helium-neon laser].

    PubMed

    Paleev, N R; Karandashov, V I; Voronina, M A; Fin'ko, I A

    1993-10-01

    An investigation of blood rheologic properties in 12 patients with acute pneumonia has been made by using low-energy He-Ne laser (LG-79-2) irradiation of vascular fascicle. 37 exposures have been performed, 40 min each. Immediate effects of He-Ne laser were studied comparing blood samples taken before and after the radiation. Results obtained have demonstrated transcutaneous blood irradiation causing prompt and pronounced effects on blood rheologic characteristics: reduced blood viscosity, improved both viscous-elastic properties and osmotic resistance of erythrocytes, activated platelet aggregation. These effects excluding the latter are rather positive for the human body. As for platelet aggregation, its activation following transcutaneous laser irradiation of blood might be expected to grow into a factor of risk provoking pathologic thrombogenesis in venous congestion, hypercoagulation and vascular wall injury.

  1. [Intravenous laser irradiation of the blood in occlusive vascular diseases of the extremities].

    PubMed

    Shval'b, P G; Zakharchenko, A Ia; Sigaev, A A; Kataev, M I

    1990-01-01

    The authors analyze the results of clinical application of intravenous He-Ne laser irradiation of the blood in patients with obliterating diseases of the limb vessels. Starting from 1984, this method was employed in the treatment of 133 patients, of these 102 ones with atherosclerosis obliterans of the lower limb vessels, 17 with endarteritis obliterans, and 14 with Raynaud's syndrome. Intravenous laser therapy proved to the most effective in atherosclerotic involvement of the vessels, when positive result was achieved in 77.5 percent of patients. The length of remission was up to 6 months. the method of treatment is described. PMID:2367895

  2. Optimum pulse duration and radiant exposure for vascular laser therapy of dark port-wine skin: a theoretical study.

    PubMed

    Tunnell, James W; Wang, Lihong V; Anvari, Bahman

    2003-03-01

    Laser therapy for cutaneous hypervascular malformations such as port-wine stain birthmarks is currently not feasible for dark-skinned individuals. We study the effects of pulse duration, radiant exposure, and cryogen spray cooling (CSC) on the thermal response of skin, using a Monte Carlo based optical-thermal model. Thermal injury to the epidermis decreases with increasing pulse duration during irradiation at a constant radiant exposure; however, maintaining vascular injury requires that the radiant exposure also increase. At short pulse durations, only a minimal increase in radiant exposure is necessary for a therapeutic effect to be achieved because thermal diffusion from the vessels is minimal. However, at longer pulse durations the radiant exposure must be greatly increased. There exists an optimum pulse duration at which minimal damage to the epidermis and significant injury within the targeted vasculature occur. For example, the model predicts optimum pulse durations of approximately 1.5, 6, and 20 ms for vessel diameters of 40, 80, and 120 microm, respectively. Optimization of laser pulse duration and radiant exposure in combination with CSC may offer a means to treat cutaneous lesions in dark-skinned individuals. PMID:12638894

  3. Optimum Pulse Duration and Radiant Exposure for Vascular Laser Therapy of Dark port-wine Skin: A Theoretical Study

    NASA Astrophysics Data System (ADS)

    Tunnell, James W.; Wang, Lihong V.; Anvari, Bahman

    2003-03-01

    Laser therapy for cutaneous hypervascular malformations such as port-wine stain birthmarks is currently not feasible for dark-skinned individuals. We study the effects of pulse duration, radiant exposure, and cryogen spray cooling (CSC) on the thermal response of skin, using a Monte Carlo based optical-thermal model. Thermal injury to the epidermis decreases with increasing pulse duration during irradiation at a constant radiant exposure; however, maintaining vascular injury requires that the radiant exposure also increase. At short pulse durations, only a minimal increase in radiant exposure is necessary for a therapeutic effect to be achieved because thermal diffusion from the vessels is minimal. However, at longer pulse durations the radiant exposure must be greatly increased. There exists an optimum pulse duration at which minimal damage to the epidermis and significant injury within the targeted vasculature occur. For example, the model predicts optimum pulse durations of approximately 1.5, 6, and 20 ms for vessel diameters of 40, 80, and 120 μm, respectively. Optimization of laser pulse duration and radiant exposure in combination with CSC may offer a means to treat cutaneous lesions in dark-skinned individuals.

  4. Effects of endovenous laser ablation on vascular tissue: molecular genetics approach

    PubMed Central

    Alur, İhsan; Dodurga, Yavuz; Güneş, Tevfik; Eroglu, Canan; Durna, Fırat; Türk, Nilay Şen; Adıgüzel, Esat; Emrecan, Bilgin

    2015-01-01

    Background: Endovenous laser ablation (EVLA) is a treatment option for lower extremity varicose veins. In the present study, we investigate to the genetic changes and possibility of living tissue in the saphenous vein wall after the EVLA procedure. Methods: Eleven saphenous vein grafts were randomized in two groups: (1) 4 cm SVG segments of performed EVLA procedure in study group, (2) 4 cm segments of SVG none performed EVLA procedure in control group. SVG were taken from the remnants of distal saphenous vein grafts prepared for the bypass procedure but not used. SVG was approximately 8 cm in length and was divided into two parts 4 cm in length. One half was exposed to laser energy, while the other half of the same vein graft was untreated as a control. EVLA was performed on complete saphenous veins in the study group. Abnormal genetic changes of the SVG were observed with a Tri-Reagent method and quantified with a Nanodrop™ spectrophotometer. Results: Histopathological changes indicated that the intima including the endothelium was completely necrotized in the study group. It was observed that intimal thermal-energy-induced injury did not reach the media. Histopathological examination showed that homogenous eosinophilic discoloration and coagulation necrosis characterized the laser related thermal damage as well. Conclusions: In this preliminary study, we found that living tissue remained in the SVG wall after application of laser ablation, and we also detected abnormal genetic changes in the study group compared with the control group. PMID:26379903

  5. Determination of EGFR mutations in single cells microdissected from enriched lung tumor cells in peripheral blood.

    PubMed

    Ran, Ran; Li, Longyun; Wang, Mengzhao; Wang, Shulan; Zheng, Zhi; Lin, Peter Ping

    2013-09-01

    A minimally invasive and repeatable approach for real-time epidermal growth factor receptor (EGFR) mutation surveillance would be highly beneficial for individualized therapy of late stage lung cancer patients whose surgical specimens are often not available. We aim to develop a viable method to detect EGFR mutations in single circulating tumor cells (CTCs). Using a model CTC system of spiked tumor cells in whole blood, we evaluated EGFR mutation determination in single tumor cells enriched from blood. We used magnetic beads labeled with antibody against leukocyte surface antigens to deplete leukocytes and enrich native CTCs independent of epithelial marker expression level. We then used laser cell microdissection (LCM) to isolate individual CTCs, followed by whole-genome amplification of the DNA for exon 19 microdeletion, L858R and T790M mutation detection by PCR sequencing. EGFR mutations were successfully measured in individual spiked tumor cells enriched from 7.5 ml whole blood. Whole-genome amplification provided sufficient DNA for mutation determination at multiple sites. Ninety-five percent of the single CTCs microdissected by LCM (19/20) yielded PCR amplicons for at least one of the three mutation sites. The amplification success rates were 55 % (11/20) for exon 19 deletion, 45 % (9/20) for T790M, and 85 % (17/20) for L858R. Sequencing of the amplicons showed allele dropout in the amplification reactions, but mutations were correctly identified in 80 % of the amplicons. EGFR mutation determination from single captured tumor cells from blood is feasible with the approach described here. However, to overcome allele dropout and to obtain reliable information about the tumor's EGFR status, multiple individual tumor cells should be assayed. PMID:23828210

  6. Laser Doppler line scanner for monitoring skin perfusion changes of port wine stains during vascular-targeted photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Chen, Defu; Ren, Jie; Wang, Ying; Gu, Ying

    2014-11-01

    Vascular-targeted photodynamic therapy (V-PDT) is known to be an effective therapeutic modality for the treatment of port wine stains (PWS). Monitoring the PWS microvascular response to the V-PDT is crucial for improving the effectiveness of PWS treatment. The objective of this study was to use laser Doppler technique to directly assess the skin perfusion in PWS before and during V-PDT. In this study, 30 patients with PWS were treated with V-PDT. A commercially laser Doppler line scanner (LDLS) was used to record the skin perfusion of PWS immediately before; and at 1, 3, 5, 7, 10, 15 and 20 minutes during V-PDT treatment. Our results showed that there was substantial inter- and intra-patient perfusion heterogeneity in PWS lesion. Before V-PDT, the comparison of skin perfusion in PWS and contralateral healthy control normal skin indicated that PWS skin perfusion could be larger than, or occasionally equivalent to, that of control normal skin. During V-PDT, the skin perfusion in PWS significantly increased after the initiation of V-PDT treatment, then reached a peak within 10 minutes, followed by a slowly decrease to a relatively lower level. Furthermore, the time for reaching peak and the subsequent magnitude of decrease in skin perfusion varied with different patients, as well as different PWS lesion locations. In conclusion, the LDLS system is capable of assessing skin perfusion changes in PWS during V-PDT, and has potential for elucidating the mechanisms of PWS microvascular response to V-PDT.

  7. [Focal Laser Ablation and Photodynamic Vascular Therapy with soluble TOOKAD® in the treatment of low risk prostate cancer].

    PubMed

    Domínguez-Escrig, J L; Casanova Ramón-Borja, J

    2016-07-01

    The increase of the diagnosis of low risk prostate cancer translates into a new clinical entity, for which active surveillance may not be always enough and conventional therapies are clearly overtreatment. Faced with the necessity of giving a therapeutic answer to these patients, and facilitated by the technological advances in the imaging field and new energy sources, the interest is centered in the clinical development of focal therapies as an alternative with minimal morbidity and oncologically safe. As a part of the review carried out in this monographic issue, this article focus on the features relative to the preclinical and clinical development of laser ablative therapy and the innovative photodynamic vascular therapy with soluble TOOKAD®. With this aim we performed an exhaustive bibliographic search, updated to February 2016, in the greater databases, including original articles and reviews in reference to the object of this review, without restrictions for year of publication. This article reviews the preclinical and clinical development of these innovative ablative techniques in the field of focal therapy for low risk prostate cancer. PMID:27416636

  8. Assessment of the Effects of Low-Level Laser Therapy on the Thyroid Vascularization of Patients with Autoimmune Hypothyroidism by Color Doppler Ultrasound

    PubMed Central

    Höfling, Danilo Bianchini; Chavantes, Maria Cristina; Juliano, Adriana G.; Cerri, Giovanni G.; Knobel, Meyer; Yoshimura, Elisabeth M.; Chammas, Maria Cristina

    2012-01-01

    Background. Chronic autoimmune thyroiditis (CAT) frequently alters thyroid vascularization, likely as a result of the autoimmune process. Objective. To evaluate the effects of low-level laser therapy (LLLT) on the thyroid vascularization of patients with hypothyroidism induced by CAT using color Doppler ultrasound parameters. Methods. In this randomized clinical trial, 43 patients who underwent levothyroxine replacement for CAT-induced hypothyroidism were randomly assigned to receive either 10 sessions of LLLT (L group, n = 23) or 10 sessions of a placebo treatment (P group, n = 20). Color Doppler ultrasounds were performed before and 30 days after interventions. To verify the vascularity of the thyroid parenchyma, power Doppler was performed. The systolic peak velocity (SPV) and resistance index (RI) in the superior (STA) and inferior thyroid arteries (ITAs) were measured by pulsed Doppler. Results. The frequency of normal vascularization of the thyroid lobes observed in the postintervention power Doppler examination was significantly higher in the L than in the P group (P = 0.023). The pulsed Doppler examination revealed an increase in the SPV of the ITA in the L group compared with the P group (P = 0.016). No significant differences in the SPV of the STA and in the RI were found between the groups. Conclusion. These results suggest that LLLT can ameliorate thyroid parenchyma vascularization and increase the SPV of the ITA of patients with hypothyroidism caused by CAT. PMID:23316383

  9. [Lasers].

    PubMed

    Passeron, T

    2012-11-01

    Lasers are a very effective approach for treating many hyperpigmented lesions. They are the gold standard treatment for actinic lentigos and dermal hypermelanocytosis, such as Ota nevus. Becker nevus, hyperpigmented mosaicisms, and lentigines can also be successfully treated with lasers, but they could be less effective and relapses can be observed. However, lasers cannot be proposed for all types of hyperpigmentation. Thus, freckles and café-au-lait macules should not be treated as the relapses are nearly constant. Due to its complex pathophysiology, melasma has a special place in hyperpigmented dermatoses. Q-switched lasers (using standard parameters or low fluency) should not be used because of consistent relapses and the high risk of post-inflammatory hyperpigmentation. Paradoxically, targeting the vascular component of the melasma lesion with lasers could have a beneficial effect. However, these results have yet to be confirmed. In all cases, a precise diagnosis of the type of hyperpigmentation is mandatory before any laser treatment, and the limits and the potential side effects of the treatment must be clearly explained to patients.

  10. Lasers.

    PubMed

    Passeron, T

    2012-12-01

    Lasers are a very effective approach for treating many hyperpigmented lesions. They are the gold standard treatment for actinic lentigos and dermal hypermelanocytosis, such as Ota nevus. Becker nevus, hyperpigmented mosaicisms, and lentigines can also be successfully treated with lasers, but they could be less effective and relapses can be observed. However, lasers cannot be proposed for all types of hyperpigmentation. Thus, freckles and café-au-lait macules should not be treated as the relapses are nearly constant. Due to its complex pathophysiology, melasma has a special place in hyperpigmented dermatoses. Q-switched lasers (using standard parameters or low fluency) should not be used because of consistent relapses and the high risk of post-inflammatory hyperpigmentation. Paradoxically, targeting the vascular component of the melasma lesion with lasers could have a beneficial effect. However, these results have yet to be confirmed. In all cases, a precise diagnosis of the type of hyperpigmentation is mandatory before any laser treatment, and the limits and the potential side effects of the treatment must be clearly explained to patients.

  11. Quantification of sympathetic vascular responses in skin by laser Doppler flowmetry.

    PubMed

    Khan, F; Spence, V A; Wilson, S B; Abbot, N C

    1991-05-01

    An improved physiological test of focal sympathetic nervous function using a laser Doppler flowmeter is presented. The test evaluates rapid reflex changes in skin blood flow at the finger tip where there are abundant arteriovenous anastomoses with dense sympathetic innervation. Indirect body heating was employed in all subjects to induce central vasodilation and to obtain stable comparable finger tip blood flows prior to stimulus. The reflex vasoconstriction which occurs following inspiratory gasp and contralateral hand cold challenge was quantified and its reproducibility investigated on three separate occasions in 20 young subjects. The variability in responses both within and between young subjects was small. The test was applied to 10 diabetic patients with autonomic neuropathy and to 10 age-matched control subjects. Vasoconstrictor reflexes were significantly lower in the diabetic group (p less than 0.005) with responses lower than 2 SD from the mean for age-matched controls. In conclusion, the test provides an assessment of focal autonomic damage which can be applied to other regions of the body rich in arteriovenous anastomoses and may have application in clinical studies investigating autonomic activity. PMID:2060997

  12. Identification of specific protein markers in microdissected hepatocellular carcinoma.

    PubMed

    Melle, Christian; Ernst, Günther; Scheibner, Olaf; Kaufmann, Roland; Schimmel, Bettina; Bleul, Annett; Settmacher, Utz; Hommann, Merten; Claussen, Uwe; von Eggeling, Ferdinand

    2007-01-01

    At present, the molecular mechanisms of hepatocellular carcinogenesis are not well-understood, and hepatocellular carcinoma (HCC) stays one of the most frequent and high-risk metastatic visceral neoplasms worldwide. For the identification of tumor-relevant proteins, we analyzed microdissected cells from nontumorous liver tissue (n = 28) and tissue derived from hepatic tumor center (n = 25), as well as tumor margin (n = 23). We unequivocally identified 53 proteins from hepatic tumor tissues by peptide fingerprint mapping and SELDI mass spectrometry that were separated using two-dimensional gel electrophoresis. Among a number of signals that were detected as significantly different in the protein profiling analysis, we identified for the first time ferritin light subunit (FLS) and adenylate kinase 3 alpha-like 1 (AK3), showing decreased expressions in hepatic tumor, as well as biliverdin reductase B (BVRB) that was upregulated in HCC. The use of ProteinChip technology in combination with tissue microdissection gives insight of the complex changes occurring at the protein level in hepatocellular cancer associated with tumor development and progression and resulted in three new potential diagnostically useful markers. PMID:17203974

  13. A Smart Haptic Hand-Held Device for Neurosurgical Microdissection.

    PubMed

    Payne, Christopher J; Marcus, Hani J; Yang, Guang-Zhong

    2015-09-01

    Microneurosurgery requires dexterity, precision and delicate force application in order to be carried out safely and effectively. Neurosurgeons must apply sufficient force in order to carry out microsurgical procedures effectively but not excessive force such that iatrogenic injury occurs. This paper presents a smart hand-held microsurgical instrument that indicates to the surgeon when a force-threshold has been exceeded by providing vibrotactile feedback. Many existing haptic-feedback systems, particularly master-slave robotic platforms, are large, highly complex, and costly. By comparison, the proposed device is compact, fail-safe and low cost. Two psychophysical user studies were carried out to assess the proposed vibrotactile force-threshold feedback system. A cadaveric pilot study was carried out to evaluate the device in a microdissection task. In all the studies performed, the haptic dissector device has shown to be effective in providing real-time feedback in terms of force application during microsurgical tasks. PMID:25631207

  14. Analysis of microdissected neurons by 18O mass spectrometry reveals altered protein expression in Alzheimer's disease

    PubMed Central

    Hashimoto, Masakazu; Bogdanovic, Nenad; Nakagawa, Hiroyuki; Volkmann, Inga; Aoki, Mikio; Winblad, Bengt; Sakai, Jun; Tjernberg, Lars O

    2012-01-01

    Abstract It is evident that the symptoms of Alzheimer's disease (AD) are derived from severe neuronal damage, and especially pyramidal neurons in the hippocampus are affected pathologically. Here, we analysed the proteome of hippocampal neurons, isolated from post-mortem brains by laser capture microdissection. By using 18O labelling and mass spectrometry, the relative expression levels of 150 proteins in AD and controls were estimated. Many of the identified proteins are involved in transcription and nucleotide binding, glycolysis, heat-shock response, microtubule stabilization, axonal transport or inflammation. The proteins showing the most altered expression in AD were selected for immunohistochemical analysis. These analyses confirmed the altered expression levels, and showed in many AD cases a pathological pattern. For comparison, we also analysed hippocampal sections by Western blot. The expression levels found by this method showed poor correlation with the neuron-specific analysis. Hence, we conclude that cell-specific proteome analysis reveals differences in the proteome that cannot be detected by bulk analysis. PMID:21883897

  15. Laser Vascular Lesion Treatment

    MedlinePlus

    ... 2006-2013 Logical Images, Inc. All rights reserved. Advertising Notice This Site and third parties who place ... would like to obtain more information about these advertising practices and to make choices about online behavioral ...

  16. Vascular Lesions.

    PubMed

    Jahnke, Marla N

    2016-08-01

    Vascular lesions in childhood are comprised of vascular tumors and vascular malformations. Vascular tumors encompass neoplasms of the vascular system, of which infantile hemangiomas (IHs) are the most common. Vascular malformations, on the other hand, consist of lesions due to anomalous development of the vascular system, including the capillary, venous, arterial, and lymphatic systems. Capillary malformations represent the most frequent type of vascular malformation. IHs and vascular malformations tend to follow relatively predictable growth patterns in that IHs grow then involute during early childhood, whereas vascular malformations tend to exhibit little change. Both vascular tumors and vascular malformations can demonstrate a wide range of severity and potential associated complications necessitating specialist intervention when appropriate. Evaluation and treatment of the most common types of vascular lesions are discussed in this article. [Pediatr Ann. 2016;45(8):e299-e305.]. PMID:27517358

  17. Spatial gene expression analysis in tomato hypocotyls suggests cysteine as key precursor of vascular sulfur accumulation implicated in Verticillium dahliae defense.

    PubMed

    Klug, Katharina; Hogekamp, Claudia; Specht, André; Myint, San Shwe; Blöink, Dominik; Küster, Helge; Horst, Walter J

    2015-02-01

    Verticillium dahliae is a prominent generator of plant vascular wilting disease and sulfur (S)-enhanced defense (SED) mechanisms contribute to its in-planta elimination. The accumulation of S-containing defense compounds (SDCs) including elemental S (S(0) ) has been described based on the comparison of two near-isogenic tomato (Solanum lycopersicum) lines differing in fungal susceptibility. To better understand the effect of S nutrition on V. dahliae resistance both lines were supplied with low, optimal or supraoptimal sulfate-S. An absolute quantification demonstrated a most effective fungal elimination due to luxury plant S nutrition. High-pressure liquid chromatography (HPLC) showed a strong regulation of Cys levels and an S-responsive GSH pool rise in the bulk hypocotyl. High-frequency S peak accumulations were detected in vascular bundles of resistant tomato plants after fungal colonization by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS). Global transcriptomic analysis suggested that early steps of the primary S metabolism did not promote the SDCs synthesis in the whole hypocotyl as gene expression was downregulated after infection. Enhanced S fertilization mostly alleviated the repressive fungal effect but did not reverse it. Upregulation of glutathione (GSH)-associated genes in bulk hypocotyls but not in vascular bundles indicated a global antioxidative role of GSH. To finally assign the contribution of S metabolism-associated genes to high S(0) accumulations exclusively found in the resistant tomato line, a spatial gene expression approach was applied. Laser microdissection of infected vascular bundles revealed a switch toward transcription of genes connected with cysteine (Cys) synthesis. The upregulation of LeOASTLp1 suggests a role for Cys as key precursor for local S accumulations (possibly S(0) ) in the vascular bundles of the V. dahliae-resistant tomato line.

  18. Lasers.

    ERIC Educational Resources Information Center

    Schewe, Phillip F.

    1981-01-01

    Examines the nature of laser light. Topics include: (1) production and characteristics of laser light; (2) nine types of lasers; (3) five laser techniques including holography; (4) laser spectroscopy; and (5) laser fusion and other applications. (SK)

  19. High-Throughput Microdissection for Next-Generation Sequencing

    PubMed Central

    Rosenberg, Avi Z.; Armani, Michael D.; Fetsch, Patricia A.; Xi, Liqiang; Pham, Tina Thu; Raffeld, Mark; Chen, Yun; O’Flaherty, Neil; Stussman, Rebecca; Blackler, Adele R.; Du, Qiang; Hanson, Jeffrey C.; Roth, Mark J.; Filie, Armando C.; Roh, Michael H.; Emmert-Buck, Michael R.; Hipp, Jason D.; Tangrea, Michael A.

    2016-01-01

    Precision medicine promises to enhance patient treatment through the use of emerging molecular technologies, including genomics, transcriptomics, and proteomics. However, current tools in surgical pathology lack the capability to efficiently isolate specific cell populations in complex tissues/tumors, which can confound molecular results. Expression microdissection (xMD) is an immuno-based cell/subcellular isolation tool that procures targets of interest from a cytological or histological specimen. In this study, we demonstrate the accuracy and precision of xMD by rapidly isolating immunostained targets, including cytokeratin AE1/AE3, p53, and estrogen receptor (ER) positive cells and nuclei from tissue sections. Other targets procured included green fluorescent protein (GFP) expressing fibroblasts, in situ hybridization positive Epstein-Barr virus nuclei, and silver stained fungi. In order to assess the effect on molecular data, xMD was utilized to isolate specific targets from a mixed population of cells where the targets constituted only 5% of the sample. Target enrichment from this admixed cell population prior to next-generation sequencing (NGS) produced a minimum 13-fold increase in mutation allele frequency detection. These data suggest a role for xMD in a wide range of molecular pathology studies, as well as in the clinical workflow for samples where tumor cell enrichment is needed, or for those with a relative paucity of target cells. PMID:26999048

  20. Vascular Cures

    MedlinePlus

    ... Malformation Atherosclerosis Buerger's Disease Carotid Artery Disease Chronic Venous Insufficiency Congenital Vascular Malformation Critical Limb Ischemia (CLI) Deep Vein Thrombosis (DVT) Diabetes and Vascular Disease Fibromuscular Dysplasia High ...

  1. Rapid generation of whole chromosome painting probes (WCPs) by chromosome microdissection

    SciTech Connect

    Guan, X.Y.; Meltzer, P.S.; Trent, J.M.

    1994-07-01

    A strategy for rapid construction of whole chromosome painting probes (WCPs) by chromosome microdissection has recently been developed. WCPs were prepared from 20 copies of each target chromosome microdissected from normal human metaphase chromosomes and then directly amplified by PCR using a universal primer. Fifteen WCPs, including chromosomes 1, 3, 6, 7, 9, 12, 13, 14, 15, 17, 19, 20, 21, 22, and X, have been generated using this strategy. The probe complexity and hybridization specificity of these WCPs have been characterized by gel electrophoresis and fluorescence in situ hybridization. Analysis of WCPs constructed by chromosome microdissection indicated that microdissected WCPs invariably provide strong and uniform signal intensity with no cytologically apparent cross-hybridization. To demonstrate the application of WCPs generated from microdissection, the authors have used these probes to detect complex chromosome rearrangements in a melanoma cell line, UM93-007. Two different translocations involving three chromosomes [t(1;3;13) and t(1;7;13)] have been identified, both of which were undetectable by conventional banding analysis. Further application of these WCPs (including generation of WCPs from mouse and other species) should greatly facilitate the cytogenetic analysis of complex chromosome rearrangements. 35 refs., 4 figs.

  2. [Vascular parkinsonism].

    PubMed

    Marxreiter, F; Winkler, J

    2016-07-01

    Parkinsonism may result from cerebral vascular disorders that feature white matter lesions and small vessel pathology. Vascular Parkinsonism typically presents as lower body Parkinsonism with predominant gait impairment. Urinary incontinence and cognitive decline are additional features of the disease. There is a considerable overlap between vascular Parkinsonism and vascular dementia. We review the clinical characteristics of vascular Parkinsonism and discuss the current treatment approaches, as well as the role of brain imaging for the diagnostic workup. . PMID:27299942

  3. Microdissection of black widow spider silk-producing glands.

    PubMed

    Jeffery, Felicia; La Mattina, Coby; Tuton-Blasingame, Tiffany; Hsia, Yang; Gnesa, Eric; Zhao, Liang; Franz, Andreas; Vierra, Craig

    2011-01-01

    case threads] and pyriform [produces attachment disc silk]. This approach is based upon anesthetizing the spider with carbon dioxide gas, subsequent separation of the cephalothorax from the abdomen, and microdissection of the abdomen to obtain the silk-producing glands. Following the separation of the different silk-producing glands, these tissues can be used to retrieve different macromolecules for distinct biochemical analyses, including quantitative real-time PCR, northern- and western blotting, mass spectrometry (MS or MS/MS) analyses to identify new silk protein sequences, search for proteins that participate in the silk assembly pathway, or use the intact tissue for cell culture or histological experiments. PMID:21248709

  4. Vascular ring

    MedlinePlus

    ... with aberrant subclavian and left ligamentum ateriosus; Congenital heart defect - vascular ring; Birth defect heart - vascular ring ... accounts for less than 1% of all congenital heart problems. The condition occurs as often in males ...

  5. Gene recovery microdissection (GRM) a process for producing chromosome region-specific libraries of expressed genes

    SciTech Connect

    Christian, A T; Coleman, M A; Tucker, J D

    2001-02-08

    Gene Recovery Microdissection (GRM) is a unique and cost-effective process for producing chromosome region-specific libraries of expressed genes. It accelerates the pace, reduces the cost, and extends the capabilities of functional genomic research, the means by which scientists will put to life-saving, life-enhancing use their knowledge of any plant or animal genome.

  6. PUNCTATE VASCULAR EXPRESSION1 Is a Novel Maize Gene Required for Leaf Pattern Formation That Functions Downstream of the Trans-Acting Small Interfering RNA Pathway1[C][W][OA

    PubMed Central

    Zhang, Xiaolan; Douglas, Ryan N.; Strable, Josh; Lee, Michelle; Buckner, Brent; Janick-Buckner, Diane; Schnable, Patrick S.; Timmermans, Marja C.P.; Scanlon, Michael J.

    2012-01-01

    The maize (Zea mays) gene RAGGED SEEDLING2-R (RGD2-R) encodes an ARGONAUTE7-like protein required for the biogenesis of trans-acting small interfering RNA, which regulates the accumulation of AUXIN RESPONSE FACTOR3A transcripts in shoots. Although dorsiventral polarity is established in the narrow and cylindrical leaves of rgd2-R mutant plants, swapping of adaxial/abaxial epidermal identity occurs and suggests a model wherein RGD2 is required to coordinate dorsiventral and mediolateral patterning in maize leaves. Laser microdissection-microarray analyses of the rgd2-R mutant shoot apical meristem identified a novel gene, PUNCTATE VASCULAR EXPRESSION1 (PVE1), that is down-regulated in rgd2-R mutant apices. Transcripts of PVE1 provide an early molecular marker for vascular morphogenesis. Reverse genetic analyses suggest that PVE1 functions during vascular development and in mediolateral and dorsiventral patterning of maize leaves. Molecular genetic analyses of PVE1 and of rgd2-R;pve1-M2 double mutants suggest a model wherein PVE1 functions downstream of RGD2 in a pathway that intersects and interacts with the trans-acting small interfering RNA pathway. PMID:22669891

  7. Femtosecond laser photodisruption of vitelline vessels of avian embryos as a technique to study embryonic vascular remodeling.

    PubMed

    Yalcin, Huseyin C

    2014-12-01

    During cardiogenesis, congenital heart defects (CHDs), generally start as local tissue abnormalities without underlying genetic causes, suggesting abnormal hemodynamics may be an important source. Due to the scarcity of experimental techniques that permits the formation of minimally-invasive and well-controlled cardiac perturbations, experimental investigation of embryonic development of CHD via in-vivo models is difficult. In this study, in order to investigate the relationship between abnormal mechanical signaling and embryonic CHD development, a previously developed laser-based technique was adopted to alter chicken embryonic cardiovascular development. The technique incorporates two-photon fluorescence microscopy to visualize deep tissue while femtosecond-pulsed laser photodisruption is used to ablate targeted tissue. Vitelline vessel remodeling under abnormal hemodynamics was the prime concern of the study. In order to alter the hemodynamics, blood flowing inside 50-300 µm diameter Hamburger-Hamilton 24 embryonic vessels was selectively ablated. Red blood cells in the blood and endothelial cells of the vessel walls were damaged as a result of ablation. Cellular injuries led to micro-occlusions in the vessels. Several micro-occlusions formed stable clots, resulting in a complete cessation of blood flow in the targeted vessels. By measuring blood velocities in the surrounding vessels via line scanning technique, the subsequent redistribution of blood flow in the immediate upstream and downstream vessels was revealed. The network was analyzed after 24 h, and it was found to be degraded. Degradation of the entire network can be attributed to the abnormalities in hemodynamics within the vessels. For studying embryonic development of heart defects under disturbed flow conditions, the present study can be extended to clot a blood vessel inside the embryo or a vitelline vessel in the vicinity of the heart. These results demonstrate that, laser-based noninvasive

  8. Comparative Tissue Proteomics of Microdissected Specimens Reveals Novel Candidate Biomarkers of Bladder Cancer*

    PubMed Central

    Chen, Chien-Lun; Chung, Ting; Wu, Chih-Ching; Ng, Kwai-Fong; Yu, Jau-Song; Tsai, Cheng-Han; Chang, Yu-Sun; Liang, Ying; Tsui, Ke-Hung; Chen, Yi-Ting

    2015-01-01

    More than 380,000 new cases of bladder cancer are diagnosed worldwide, accounting for ∼150,200 deaths each year. To discover potential biomarkers of bladder cancer, we employed a strategy combining laser microdissection, isobaric tags for relative and absolute quantitation labeling, and liquid chromatography-tandem MS (LC-MS/MS) analysis to profile proteomic changes in fresh-frozen bladder tumor specimens. Cellular proteins from four pairs of surgically resected primary bladder cancer tumor and adjacent nontumorous tissue were extracted for use in two batches of isobaric tags for relative and absolute quantitation experiments, which identified a total of 3220 proteins. A DAVID (database for annotation, visualization and integrated discovery) analysis of dysregulated proteins revealed that the three top-ranking biological processes were extracellular matrix organization, extracellular structure organization, and oxidation-reduction. Biological processes including response to organic substances, response to metal ions, and response to inorganic substances were highlighted by up-expressed proteins in bladder cancer. Seven differentially expressed proteins were selected as potential bladder cancer biomarkers for further verification. Immunohistochemical analyses showed significantly elevated levels of three proteins—SLC3A2, STMN1, and TAGLN2—in tumor cells compared with noncancerous bladder epithelial cells, and suggested that TAGLN2 could be a useful tumor tissue marker for diagnosis (AUC = 0.999) and evaluating lymph node metastasis in bladder cancer patients. ELISA results revealed significantly increased urinary levels of both STMN1 and TAGLN2 in bladder cancer subgroups compared with control groups. In comparisons with age-matched hernia urine specimens, urinary TAGLN2 in bladder cancer samples showed the largest fold change (7.13-fold), with an area-under-the-curve value of 0.70 (p < 0.001, n = 205). Overall, TAGLN2 showed the most significant

  9. Measurement of Inflammatory Chemokines in Micro-dissected Tissue Biopsy Samples by Chip-Based Immunoaffinity Capillary Electrophoresis.

    PubMed

    Phillips, Terry M; Wellner, Edward; McMohan, Shane; Kalish, Heather

    2016-01-01

    To aid in the biochemical analysis of human skin biopsies, a chip-based immunoaffinity capillary electrophoresis (ICE) system has been developed for measuring inflammatory chemokines in micro-dissected areas of the biopsy. Following isolation of the areas of interest, the tissue was solubilized and the analytes of interest were isolated by the immunoaffinity disk within the chip. The captured analytes were labeled in situ with a 635 nm light-emitting laser dye and electro-eluted into the chip separation channel. Electrophoretic separation of all of the analytes was achieved in 2.5 min with quantification of each peak being performed by online LIF detection and integration of each peak area. The degree of accuracy and precision achieved by the chip-based system is comparable to conventional immunoassays and the system is robust enough to be applied to the analysis of clinical samples. Further, with the expanding array of antibodies that are commercially available, this chip-based system can be applied to a wide variety of different biomedical and clinical analyses. PMID:27473486

  10. Microdissection and Chromosome Painting of the Alien Chromosome in an Addition Line of Wheat - Thinopyrum intermedium

    PubMed Central

    Yin, Weibo; Zhang, Yingxin; Chen, Yuhong; Wang, Richard R.-C.; Zhang, Xiangqi; Han, Fangpu; Hu, Zanmin

    2013-01-01

    In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat - Thinopyrumintermedium addition line, and the chromosomes of the three different genomes of Th. Intermedium. The smallest alien chromosome of TAi-27 was microdissected and its DNA amplified by DOP-PCR was used as a probe to hybridize with metaphase chromosomes of TAi-27 and Th. intermedium. Results showed that hybridization signals were observed in all regions of a pair of the smallest alien chromosomes and the pericentromeric area of another pair of alien chromosomes in TAi-27, indicating that the probe from microdissected chromosome is species specific. In Th. intermedium, 14 chromosomes had wide and strong hybridization signals distributed mainly on the pericentromere area and 9 chromosomes with narrow and weak signals on the pericentromere area. The remaining chromosomes displayed a very weak or no signal. Sequential FISH/GISH on Th. intermedium chromosomes using the DNAs of microdissected chromosome, Pseudoroegneriaspicata (St genome) and pDbH12 (a Js genome specific probe) as the probes indicated that the microdissected chromosome belonged to the St genome, three genomes (Js, J and St) in Th. intermedium could be distinguished, in which there is no hybridization signal on J genome that is similar to the genome of Th. bessarabicum. Our results showed that the smallest alien chromosomes may represent a truncated chromosome and the repetitive sequence distribution might be similar in different chromosomes within the St genome. However, the repetitive sequence distributions are different within the Js genome, within a single chromosome, and among different genomes in Th. intermedium. Our results suggested that chromosome painting could be feasible in some plants and useful in detecting chromosome variation and repetitive sequence distribution in different genomes of polyploidy plants, which is helpful for understanding the evolution of different genomes in

  11. Can the rapid identification of mature spermatozoa during microdissection testicular sperm extraction guide operative planning?

    PubMed

    Alrabeeah, K; Doucet, R; Boulet, E; Phillips, S; Al-Hathal, N; Bissonnette, F; Kadoch, I J; Zini, A

    2015-05-01

    The minimum sperm count and quality that must be identified during microdissection testicular sperm extraction (micro-TESE) to deem the procedure successful remains to be established. We conducted a retrospective study of 81 consecutive men with non-obstructive azoospermia who underwent a primary (first) micro-TESE between March 2007 and October 2013. Final assessment of sperm recovery [reported on the day of (intracytoplasmic sperm injection) ICSI] was recorded as (i) successful (available spermatozoa for ICSI) or (ii) unsuccessful (no spermatozoa for ICSI). The decision to perform a unilateral (with limited or complete microdissection) or bilateral micro-TESE was guided by the intra-operative identification of sperm recovery (≥5 motile or non-motile sperm) from the first testicle. Overall, sperm recovery was successful in 56% (45/81) of the men. A unilateral micro-TESE was performed in 47% (38/81) of the men (based on intra-operative identification of sperm) and in 100% (38/38) of these men, spermatozoa was found on final assessment. In 42% (16/38) of the unilateral cases, a limited microdissection was performed (owing to the rapid intra-operative identification of sperm). The remaining 43 men underwent a bilateral micro-TESE and 16% (7/43) of these men had sperm identified on final assessment. The cumulative ICSI pregnancy rates (per cycle started and per embryo transfer) were 47% (21/45) and 60% (21/35), respectively, with a mean (±SD) of 1.9 ± 1.0 embryos transferred. The data demonstrate that intra-operative assessment of sperm recovery can correctly identify those men that require a unilateral micro-TESE. Moreover, the rapid identification of sperm recovery can allow some men to undergo a limited unilateral micro-TESE and avoid the need for complete testicular microdissection.

  12. Microdissection and visualization of individual hair follicles for lineage tracing studies.

    PubMed

    Sequeira, Inês; Legué, Emilie; Capgras, Suzanne; Nicolas, Jean-François

    2014-01-01

    In vivo lineage tracing is a valuable technique to study cellular behavior. Our lab developed a lineage tracing method, based on the Cre/lox system, to genetically induce clonal labelling of cells and follow their progeny. Here we describe a protocol for temporally controlled clonal labelling and for microdissection of individual mouse hair follicles. We further present staining and visualization techniques used in our lab to analyze clones issued from genetically induced labelling. PMID:24281870

  13. Promoter methylation analysis on microdissected paraffin-embedded tissues using bisulfite treatment and PCR-SSCP.

    PubMed

    Bian, Y S; Yan, P; Osterheld, M C; Fontolliet, C; Benhattar, J

    2001-01-01

    Methylation-sensitive single-strand conformation analysis (MS-SSCA) is a new method of screening for DNA methylation changes. The combination of bisulfite modification and PCR results in the conversion of unmethylated cytosines to thymines, whereas methylated cytosines remain unchanged. This sequence conversion can lead to methylation-dependent alterations of single-strand conformation, which can be detected by SSCA. An analysis of mixtures of methylated and unmethylated DNA at known ratios revealed that the relative intensities of the corresponding bands following MS-SSCA were maintained. MS-SSCA was applied for methylation analysis of human p16 promoter region using genomic DNA obtained from either frozen, fixed, or microdissected fixed tissue sections. MS-SSCA is a rapid, specific, and semiquantitative approach that allows the detection of methylation of the p16 gene promoter. In reconstruction experiments, the method permits the detection of 10% or less of cells harboring a methylated p16 promoter. We have been successful in analyzing by MS-SSCA almost all (96%) tumor samples microdissected from archival paraffin-embedded fixed tissue sections and obtaining reproducible results. In addition, when microdissection was performed, the clonality of this genetic alteration could be identified.

  14. Microdissection and chromosome painting of X and B chromosomes in Locusta migratoria.

    PubMed

    Teruel, María; Cabrero, Josefa; Montiel, Eugenia E; Acosta, Manuel J; Sánchez, Antonio; Camacho, Juan Pedro M

    2009-01-01

    Acquisition of knowledge of the nature and DNA content of B chromosomes has been triggered by a collection of molecular techniques, one of which, microdissection, has provided interesting results in a number of B chromosome systems. Here we provide the first data on the molecular composition of B chromosomes in Locusta migratoria, after microdissection of the B and X chromosomes, DNA amplification by one (B) or two (X) different methods, and chromosome painting. The results showed that B chromosomes share at least two types of repetitive DNA sequences with the A chromosomes, suggesting that Bs in this species most likely arose intraspecifically. One of these repetitive DNAs is located on the heterochromatic distal half of the B chromosome and in the pericentromeric regions of about half of the A chromosomes, including the X. The other type of repetitive DNA is located interspersedly over the non-centromeric euchromatic regions of all A chromosomes and in an interstitial part of the proximal euchromatic half of the B chromosome. Chromosome painting, however, did not provide results sufficiently reliable to determine, in this species, which A chromosome gave rise to the B; this might be done by detailed analysis of the microdissected DNA sequences.

  15. Vascular Diseases

    MedlinePlus

    ... heart and blood vessels, such as diabetes or high cholesterol Smoking Obesity Losing weight, eating healthy foods, being active and not smoking can help vascular disease. Other treatments include medicines and surgery.

  16. Vascular anomalies: classification, imaging characteristics and implications for interventional radiology treatment approaches

    PubMed Central

    Prajapati, H J S; Martin, L G; Patel, T H

    2014-01-01

    The term vascular anomaly represents a broad spectrum of vascular pathology, including proliferating vascular tumours and vascular malformations. While the treatment of most vascular anomalies is multifactorial, interventional radiology procedures, including embolic therapy, sclerotherapy and laser coagulation among others, are playing an increasingly important role in vascular anomaly management. This review discusses the diagnosis and treatment of common vascular malformations, with emphasis on the technique, efficacy and complications of different interventional radiology procedures. PMID:24588666

  17. [Vascular dementia].

    PubMed

    Peters, N; Dichgans, M

    2010-10-01

    Vascular dementia (VaD) constitutes the second most frequent cause of dementia following Alzheimer's disease (AD). In contrast to AD, VaD encompasses a variety of conditions and dementia mechanisms including multiple and strategic infarcts, widespread white matter lesions and hemorrhages. The diagnosis of VaD is based on the patient history, the clinical evaluation and neuroimaging. Treatment of VaD should account for the underlying vascular condition and is directed towards the control of vascular risk factors and stroke prevention. The need for early diagnosis and preventive treatment has promoted the concept of vascular cognitive impairment (VCI). Harmonization standards for the description and study of VCI have recently been published. A common and distinct subtype of VaD is subcortical ischemic vascular dementia (SIVD) which is related to cerebral small vessel disease. SIVD is clinically characterized by impairment of executive functions and processing speed with relatively preserved memory. Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a genetic variant of SIVD, represents an important differential diagnosis and may serve as a model of SIVD.

  18. Transmyocardial Laser Revascularization

    MedlinePlus

    ... Vascular Access for Hemodialysis Ventricular Assist Devices Transmyocardial Laser Revascularization Like every other organ or tissue in ... bypass surgery, there is a procedure called transmyocardial laser revascularization, also called TMLR or TMR. TMLR cannot ...

  19. Peripheral vascular imaging and intervention

    SciTech Connect

    Kim, D. ); Orron, D.E. )

    1990-01-01

    This reference addresses the entire clinical approach to the vascular system from the diagnosis of pathology to surgery or interventional radiological management. All diagnostic imaging modalities currently available are included with specific information on how to interpret various results. It features discussions of the latest therapeutic techniques, including laser angioplasty, intravascular stents, and transluminal embolization.

  20. LASER CAPTURE MICRODISSECTION AND GENE ARRAY ANALYSIS OF PALATAL EPITHELIAL AND MESENCHYMAL CELLS EXPOSED TO TCDD

    EPA Science Inventory

    Palatal shelves from embryos exposed on gestation day (GD) 12 to either retinoic acid (RA) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) contact but fail to fuse. It is of interest to know if diverse agents that induce clefting via the same etiology also activate the same biochem...

  1. Vascular emergencies.

    PubMed

    Semashko, D C

    1997-01-01

    This article reviews the initial assessment and emergent management of several common as well as uncommon vascular emergencies. Aortic dissection, aneurysms, and arterial occlusive disease are familiar but challenging clinical entities. Less frequently encountered conditions are also discussed including an aortic enteric fistula, mesenteric venous thrombosis, phlegmasia alba dolens, and subclavian vein thrombosis.

  2. PCR artifacts in LOH and MSI analysis of microdissected tumor cells.

    PubMed

    Sieben, N L; ter Haar, N T; Cornelisse, C J; Fleuren, G J; Cleton-Jansen, A M

    2000-11-01

    Polymerase chain reaction (PCR) analysis to study loss of heterozygosity (LOH) and microsatellite instability (MSI) in tumors is widely used. Microdissection techniques are applied to obtain tumor-specific tissue cells. By microdissection, however, the amount of template DNA extracted may vary considerably and interfere with optimal PCR amplification. To circumvent LOH and MSI misinterpretations due to low DNA input, we have assessed the critical level of DNA input for reliable PCR analysis. PCR analysis was performed by using 18 polymorphic markers (mono-, di-, tri-, and tetranucleotide) on DNA derived from both paraffin-embedded, formalin-fixed, and fresh frozen tumor specimens at template input levels ranging from 0.05 to 25.0 ng. We show a highly significant relation between DNA input and the occurrence of LOH and MSI artifacts. Furthermore, for DNA extracted from paraffin-embedded material, the percentage of LOH artifacts is significantly higher compared with DNA extracted from frozen tissue. For reliable PCR analyses using a mono-, di-, tri-, or tetranucleotide marker, a minimum of 10.0 ng DNA is required when DNA is isolated from formalin-fixed, paraffin-embedded tissue and 5.0 ng when isolated from fresh frozen tissue. HUM PATHOL 31:1414-1419.

  3. In situ phosphorylation of proteins in MCTs microdissected from rat kidney: Effect of AVP

    SciTech Connect

    Homma, S.; Gapstur, S.M.; Yusufi, N.K.; Dousa, T.P. )

    1988-04-01

    Adenosine 3{prime},5{prime}-cyclic monophosphate (cAMP)-dependent protein phosphorylation is considered a key step in the cellular action of vasopressin (AVP) to regulate water permeability in collecting tubules. However, the proteins serving as a substrate(s) for phosphorylation in undisrupted cells have not yet been identified. In the present study, the authors developed a method for investigation of in situ phosphorylation of microdissected segments of medullary collecting tubules (MCT) from rat kidney. Incubation of microdissected MCT segments with low concentrations of saponin, semipermeabilization, increased permeability of the membrane for ATP but did not allow leakage of macromolecules such as lactate dehydrogenase. This treatment also did not cause major disruption of cell structure, or impairment of AVP-sensitive adenylate cyclase. Incubation of semipermeabilized MCT with {gamma}-({sup 32}P)ATP resulted in corporation of {sup 32}P{sub i} into two major protein bands detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis and subsequent autoradiography. Similar incubation of tubules disrupted by hyposmotic solutions and a stronger detergent Triton X-100 resulted in {sup 32}P{sub i} incorporation into multiple protein bands. These findings demonstrate a novel method for identification of endogenous protein substrate(s) for cAMP-dependent protein kinase and other protein kinases and phosphatases that are probably involved in post-cAMP steps in the cellular action of AVP in the intact cells of collecting tubules.

  4. Region-specific cosmids and STRPs identified by chromosome microdissection and FISH

    SciTech Connect

    Flejter, W.L.; Bennett-Baker, P.; Barcroft, C.L.

    1995-01-20

    A strategy for identifying short tandem repeat (STR)-containing cosmid clones from a specific chromosomal region is described. The approach is based an the use of uncloned, PCR-amplified DNA derived from chromosome microdissection and pooled groups of STR sequences as hybridization probes to screen a cosmid library. Cosmid clones that display a positive signal common to both hybridizations are then characterized for repeat length polymorphisms. This method has been applied to chromosome bands 17q12-q21, a region that includes a gene (BRCA1) involved in early onset familial breast and ovarian cancer. Of 1536 chromosome 17-specific cosmid clones tested, 38 were identified by the dual screening procedure. Fluorescence in situ hybridization revealed that 19 cosmids originated from the microdissected target region. Thirteen of the 19 cosmids were mapped between markers flanking the BRCA1 region and selected for further characterization. Tetranucleotide repeats were identified in 10 of these 13 cosmids. Primers designed for each marker were tested on a panel of 80 CEPH parents for allele sizes, frequencies, and observed heterozygosities. From these studies six polymorphic and one nonpolymorphic STRs were identified. A similar approach should be applicable for screening whole genomic or chromosome-specific cosmid libraries in efforts to isolate new polymorphic markers from any chromosomal region of interest. 32 refs., 3 figs., 2 tabs.

  5. Subcellular analysis by laser ablation electrospray ionization mass spectrometry

    DOEpatents

    Vertes, Akos; Stolee, Jessica A; Shrestha, Bindesh

    2014-12-02

    In various embodiments, a method of laser ablation electrospray ionization mass spectrometry (LAESI-MS) may generally comprise micro-dissecting a cell comprising at least one of a cell wall and a cell membrane to expose at least one subcellular component therein, ablating the at least one subcellular component by an infrared laser pulse to form an ablation plume, intercepting the ablation plume by an electrospray plume to form ions, and detecting the ions by mass spectrometry.

  6. Diagnosis of four chromosome abnormalities of unknown origin by chromosome microdissection and subsequent reverse and forward painting

    SciTech Connect

    Coelho, K.E.F.A. de; Egashira, M.; Kato, R.

    1996-06-14

    A molecular cytogenetic method consisting of chromosome microdissection and subsequent reverse/forward chromosome painting is a powerful tool to identify chromosome abnormalities of unknown origin. We present 4 cases of chromosome structural abnormalities whose origins were ascertained by this method. In one MCA/MR patient with an add(5q)chromosome, fluorescence in situ hybridization (FISH), using probes generated from a microdissected additional segment of the add(5q) chromosome and then from a distal region of normal chromosome 5, confirmed that the patient had a tandem duplication for a 5q35-qter segment. Similarly, we ascertained that an additional segment of an add(3p) chromosome in another MCA/MR patient had been derived from a 7q32-qter segment. In a woman with a history of successive spontaneous abortions and with a minute marker chromosome, painting using microdissected probes from the whole marker chromosome revealed that it was i(15)(p10) or psu dic(15;15)(q11;q11). Likewise, a marker observed in a fetus was a ring chromosome derived from the paracentromeric region of chromosome 19. We emphasize the value of the microdissection-based chromosome painting method in the identification of unknown chromosomes, especially for marker chromosomes. The method may contribute to a collection of data among patients with similar or identical chromosome abnormalities, which may lead to a better clinical syndrome delineation. 15 refs., 2 figs.

  7. Effects of the Combined PDL/Nd:YAG Laser on Surgical Scars: Vascularity and Collagen Changes Evaluated by In Vivo Confocal Microscopy

    PubMed Central

    Vas, Krisztina; Gaál, Magdolna; Varga, Erika; Kovács, Réka; Bende, Balázs; Kocsis, Ádám; Kemény, Lajos

    2014-01-01

    The aim of this study was to investigate the efficacy of the sequential combined 585 nm PDL and the 1064 nm neodymium:yttrium-aluminium-garnet laser (PDL/Nd:YAG) in the treatment of surgical scars and to evaluate the short-term effects by in vivo confocal microscopy (RCM) and the long-term effects by clinical assessment of the scars. Twenty-five patients were enrolled with 39 postoperative linear scars; each scar was divided into two fields. One half was treated with the combined PDL/Nd:YAG laser, whereas the other half remained untreated. Each scar was treated three times at monthly intervals. Scars were evaluated by an independent examiner, using the Vancouver Scar Scale. The combined PDL/Nd:YAG laser significantly improved the appearance of the scars. In order to study the short-term effects of combined laser treatment, six additional patients were enrolled with 7 postoperative linear scars. One half of scars was treated once with the combined PDL/Nd:YAG laser. One week after this laser treatment, both the treated and the nontreated parts of the scars were examined by dermoscopy and RCM. The dermoscopic pictures revealed improvements even in treated areas. In conclusion, the combined PDL/Nd:YAG laser was found to be effective in improving the quality and appearance of the surgical scars. PMID:25276770

  8. The Y chromosome of the Atelidae family (Platyrrhini): study by chromosome microdissection.

    PubMed

    Gifalli-Iughetti, C; Koiffmann, C P

    2009-01-01

    In order to study the intergeneric variability of the Y chromosome, we describe the hybridization of the Y chromosome of Brachytelesarachnoides, obtained by microdissection, to metaphases of Atelesbelzebuthmarginatus, Lagothrixlagothricha, and Alouatta male specimens. Brachytelesarachnoides (Atelinae) has 62 chromosomes and a very small Y chromosome. Our results showed that the Brachytelesarachnoides Y chromosome probe hybridized to Lagothrixlagothricha metaphases yielding one hybridization signal on only the tiny Y chromosome, and when hybridized with Atelesbelzebuthmarginatus metaphases it yielded one hybridization signal on two thirds of the small acrocentric Y chromosome. However, no hybridization signal was observed in Alouatta metaphases (subfamily Alouattinae), a closely related genus in the Atelidae family. Furthermore, our data support a close phylogenetic relationship among Brachyteles, Ateles, and Lagothrix and their placement in the Atelinae subfamily, but exclude Alouatta from this group indicating its placement as basal to this group.

  9. [New lasers in dermatologic surgery].

    PubMed

    Morvay, M

    1995-04-30

    Developments in the fields of laser technology and application have significantly broadened its clinical use over the past two decades. As lasers became more smaller, more reliable and less expensive, dermatology will benefit new laser-based therapeutic and diagnostic methods. Relevant laser systems and their clinical applications are presented, as are investigational laser systems, which may be of importance for the future in dermatology. We review advances in the use of pulsed lasers for treating vascular and non-vascular, pigmented epidermal and dermal lesions, for precise cutting of tissue, for photodynamic therapy and the future role of diode lasers in dermatological laser surgery is also discussed.

  10. SCOMP is superior to degenerated oligonucleotide primed-polymerase chain reaction for global amplification of minute amounts of DNA from microdissected archival tissue samples.

    PubMed

    Stoecklein, Nikolas H; Erbersdobler, Andreas; Schmidt-Kittler, Oleg; Diebold, Joachim; Schardt, Julian A; Izbicki, Jakob R; Klein, Christoph A

    2002-07-01

    Global genome amplification from formalin-fixed tissues is still problematic when performed with low cell numbers. Here, we tested a recently developed method for whole genome amplification termed "SCOMP" (single cell comparative genomic hybridization) on archival tissues of different ages. We show that the method is very well suited for formalin-fixed paraffin-embedded samples obtained by nuclei extraction or laser microdissection. The polymerase chain reaction (PCR) products can be used for subsequent comparative genomic hybridization, loss of heterozygosity studies, and DNA sequencing. To control for PCR-induced artifacts we amplified genomic DNA isolated from 20 nuclei of archival formalin-fixed, paraffin-embedded nonpathological lymph nodes. Subsequent comparative genomic hybridization revealed the expected balanced profiles. For loss of heterozygosity analysis by microsatellite PCR 60 to 160 cells were sufficient. In comparative experiments the approach turned out to be superior to published degenerated oligonucleotide-primed-PCR protocols. The method provides a robust and valuable tool to study very small cell samples, such as the genomes of dysplastic cells or the clonal evolution within heterogeneous tumors.

  11. Laser physics and laser-tissue interaction.

    PubMed

    Welch, A J; Torres, J H; Cheong, W F

    1989-01-01

    Within the last few years, lasers have gained increasing use in the management of cardiovascular disease, and laser angioplasty has become a widely performed procedure. For this reason, a basic knowledge of lasers and their applications is essential to vascular surgeons, cardiologists, and interventional radiologists. To elucidate some fundamental concepts regarding laser physics, we describe how laser light is generated and review the properties that make lasers useful in medicine. We also discuss beam profile and spotsize, as well as dosimetric specifications for laser angioplasty. After considering laser-tissue interaction and light propagation in tissue, we explain how the aforementioned concepts apply to direct laser angioplasty and laser-balloon angioplasty. An understanding of these issues should prove useful not only in performing laser angioplasty but in comparing the reported results of various laser applications.

  12. A simple method for fixation and microdissection of frozen fresh tissue sections for molecular cytogenetic analysis of cancers.

    PubMed

    Huang, Q; Sacks, P G; Mo, J; McCormick, S A; Iacob, C E; Guo, L; Schaefer, S; Schantz, S P

    2005-01-01

    Microdissection has been widely used for procuring DNA from specific microscopic regions of formalin fixed, paraffin embedded tissue sections. We have developed a method for fixation and microdissection of frozen fresh biopsy tissue sections. Five micrometer frozen fresh tissue sections were fixed with ethanol and stored at room temperature. Well defined regions from hematoxylin and eosin (H & E) stained or unstained sections were briefly steamed and microdissected using a needle. The dissected tissue was digested with proteinase K and DNA was isolated. Whole genome amplifications were obtained by degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR) from these samples. The reliability of this technique was demonstrated by comparing conventional comparative genomic hybridization (CGH) with DOP-PCR-CGH. The advantages of this method are that frozen fresh sections can be fixed easily and stored for more than 4 years, it is easy to microdissect and pick-up very minute regions (0.1 mm(2)), and it is rapid; microdissection and purification can be accomplished within 3 h. Using DNA from microdissected sections, DOP-PCR-CGH revealed genetic abnormalities more accurately than conventional CGH. Although this novel method was demonstrated using DOP-PCR-CGH, we believe that it will be useful for other genetic analyses of specific small regions and cell populations. We also observed whether storage time, H & E staining and crude DNA extracts affected the quality of amplified DNA. DNA integrity was maintained for at least 49 months in ethanol fixed sections that were stored at room temperature, but DNA was gradually degraded after one month if the ethanol fixed sections had been H & E stained and stored. When crude DNA extracts from H & E stained sections were used, the size of the DOP-PCR product was reduced. Our study suggests that ethanol fixed tissue sections may be stored at room temperature for at least 4 years without DNA degradation, the H & E stains may

  13. Vascular effect of photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Fyodorov, Svyatoslav N.; Kopayeva, V. G.; Andreev, J. B.; Ponomarev, Gelii V.; Stranadko, Eugeny P.; Suchin, H. M.

    1996-01-01

    Vascular effect of PDT has been studied in patients with corneal vascularized leucomas (10 patients) and in patients with corneal neovascularized transplant (3 patients). For vascularized leucomas the method of photodynamic therapy consisted of the local injection of dimegin (deiteroporphyrin derivative) into the space of the newly-formed vessels under operating microscope (opton) with the microneedle (diameter 200 microns) and corneal irradiation by the operating microscope light. For corneal neovascularized transplant the injection of photogem (hematoporphyrin derivative) intravenously were made with subsequent irradiation by light of dye laser (5 hours after the injection) with light density of 150 mW/cm2 for 15 minutes. In all the cases at the time of irradiation the aggregated blood flow was appeared, followed by blood flow stasis. In postoperative period the vessels disintegrated into separate fragments which disappeared completely after 10 - 15 days. Taking into account the data of light microscopy, the disappearance of the vessels took place as a result of the vascular endothelium lisis along the vascular walls. Neovascularized cornea and newly-formed vessels in tumor stroms have much in common. The vessel alterations study presented in this paper, may serve to specify the mechanism of photodynamic destruction of neovascularized stroma of tumor.

  14. Genetic association between chromosome 8 microsatellite (MS8-134) and Werner syndrome (WRN): Chromosome microdissection and homozygosity mapping

    SciTech Connect

    Ye, Lin; Nakura, Jun; Mitsuda, Noriaki; Miki, Tetsuro

    1995-08-10

    Werner syndrome (WRN) is an autosomal recessive disorder characterized by premature aging that has been mapped to the short arm of chromosome 8, 8p11.2-p12. To refine the genetic map around the WRN region, we have isolated eight microsatellites for this region from a microdissection library. We typed members of Japanese families with WRN on the basis of homozygosity mapping analysis. There was no obligate recombination between the WRN locus and microsatellite clone, MS8-134 (D8S1055). The maximum lod score was 20.28 at {theta} = 0.00. Alleles for MS8-134 showed association with WRN in a case-control study (OR = 3.55, 95% CI 1.56-8.07, P < 0.01). Such microsatellites from a microdissection library of the definite chromosome region may be useful for positional cloning of the WRN gene. 23 refs., 1 figs., 1 tab.

  15. Directed isolation and mapping of microsatellites from swine Chromosome 1q telomeric region through microdissection and RH mapping.

    PubMed

    Sarker, N; Hawken, R J; Takahashi, S; Alexander, L J; Awata, T; Schook, L B; Yasue, H

    2001-07-01

    Several quantitative trait loci (QTLs) (vertebrate number, birth weight, age at puberty, growth rate, gestation length, and backfat depth) have been independently mapped to the distal region of swine Chromosome (SSC) 1q in several resource populations. In order to improve the map resolution and refine these QTLs more precisely on SSC1q, we have isolated and mapped additional microsatellites (ms), using chromosome microdissection and radiation hybrid (RH) mapping. Five copies of the telomeric region of SSC1q were microdissected from metaphase spreads and pooled. The chromosomal fragment DNA was randomly amplified by using degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR), enriched for ms, and subcloned into a PCR vector. Screening of subsequent clones with ms probes identified 23 unique ms sequences. Fifteen of these (65%) were subjected to radiation hybrid (RH) mapping by using the INRA-University of Minnesota porcine RH panel (IMpRH); and the remaining eight were not suited for the RH mapping. Twelve microsatellites were assigned to SSC1q telomeric region of IMpRH map (LOD >6), and three remain unlinked (LOD <6). Out of the 15 microsatellite markers, 9 were polymorphic in NIAI reference population based on the Meishan and Göttingen miniature pig. In summary, we have used microdissection and radiation hybrid mapping to clone and map 12 new microsatellites to the swine gene map to increase the resolution of SSC1q in the region of known QTLs.

  16. Rapid generation of region-specific probes by chromosome microdissection: Application to the identification of chromosomal rearrangements

    SciTech Connect

    Trent, J.M.; Guan, X.Y.; Zang, J.; Meltzer, P.S. )

    1993-01-01

    The authors present results using a novel strategy for chromosome microdissection and direct in vitro amplification of specific chromosomal regions, to identify cryptic chromosome alterations, and to rapidly generate region-specific genomic probes. First, banded chromosomes are microdissected and directly PCR amplified by a procedure which eliminates microchemistry (Meltzer, et al., Nature Genetics, 1:24, 1992). The resulting PCR product can be used for several applications including direct labeling for fluorescent in situ hybridization (FISH) to normal metaphase chromosomes. A second application of this procedure is the extremely rapid generation of chromosome region-specific probes. This approach has been successfully used to determine the derivation of chromosome segments unidentifiable by standard chromosome banding analysis. In selected instances these probes have also been used on interphase nuclei and provides the potential for assessing chromosome abnormalities in a variety of cell lineages. The microdissection probes (which can be generated in <24 hours) have also been utilized in direct library screening and provide the possibility of acquiring a significant number of region-specific probes for any chromosome band. This procedure extends the limits of conventional cytogenetic analysis by providing an extremely rapid source of numerous band-specific probes, and by enabling the direct analysis of essentially any unknown chromosome region.

  17. Microdissection and painting of the Y chromosome in spinach (Spinacia oleracea).

    PubMed

    Deng, Chuan-Liang; Qin, Rui-Yun; Cao, Ying; Gao, Jun; Li, Shu-Fen; Gao, Wu-Jun; Lu, Long-Dou

    2013-07-01

    Spinach has long been used as a model for genetic and physiological studies of sex determination and expression. Although trisomic analysis from a cross between diploid and triploid plants identified the XY chromosome as the largest chromosome, no direct evidence has been provided to support this at the molecular level. In this study, the largest chromosomes of spinach from mitotic metaphase spreads were microdissected using glass needles. Degenerate oligonucleotide primed polymerase chain reaction was used to amplify the dissected chromosomes. The amplified products from the Y chromosome were identified using the male-specific marker T11A. For the first time, the largest spinach chromosome was confirmed to be a sex chromosome at the molecular level. PCR products from the isolated chromosomes were used in an in situ probe mixture for painting the Y chromosome. The fluorescence signals were mainly distributed on all chromosomes and four pair of weaker punctate fluorescence signal sites were observed on the terminal region of two pair of autosomes. These findings provide a foundation for the study of sex chromosome evolution in spinach.

  18. Altered p53 in microdissected, metachronous, premalignant and malignant oral lesions from the same patients

    PubMed Central

    Li, Y-Q; Pavelic, Z P; Wang, L-J; McDonald, J S; Gleich, L; Munck-Wikland, E; Dacic, S; Danilovic, Z; Pavelic, L J; Wilson, K M; Gluckman, J L; Stambrook, P J

    1995-01-01

    Aims—To determine whether mutant p53 alleles harboured by malignant tumours of the oral cavity were also present in previous premalignant lesions at the same site. Methods—Paraffin embedded tumour specimens along with their premalignant counterparts were analysed for p53 alterations using immunohistochemistry, microdissection, polymerase chain reaction amplification, and DNA sequencing. Results—Malignant lesions from five of eight patients showed overexpression of p53 protein by immunohistochemistry. Upon DNA sequencing, two of these five specimens had p53 mutations. Of the five patients whose cancers showed p53 overexpression by immunohistochemistry, three had previous premalignant lesions that also had immunohistochemically detectable p53 protein. However, DNA sequencing showed that none of these three had mutations in the p53 gene. The remaining five premalignant lesions had no immunohistochemically detectable p53 protein. Conclusions—Some premalignant lesions have increased p53 protein which can be detected by staining with antibody to p53. This staining is not caused by mutations in p53 that are found in subsequent tumours at the same site. Images PMID:16696020

  19. Molecular Analysis of the B Microchromosome in Steindachnerina insculpta (Characiformes: Curimatidae) by Microdissection.

    PubMed

    Sampaio, Tatiane R; Gouveia, Juceli G; da Silva, Carlos R M; Dias, Ana L; da Rosa, Renata

    2015-01-01

    B chromosomes are additional elements to standard karyotypes observed in different species of fishes, especially in Curimatidae. However, despite studies demonstrating the occurrence of Bs, little is known about their origin and evolution. To better understand the genomic composition and evolutionary processes involving B chromosomes, microdissection of B microchromosomes in Steindachnerina insculpta was conducted. Chromosome painting revealed the totally hybridized B and markings on A chromosomes both in S. in sculpta and in Cyphocharax spilotus, demonstrating a strong homology between these different species. In specimens of C. modestus, which do not have Bs, the signals were observed on A chromosomes. Cloning and sequencing of some B fragments revealed that the B microchromosome in S. insculpta is composed of repetitive elements, homologous to the DIRS-4 LTR retrotransposon of Xenopus (Silurana) tropicalis. FISH with clone pSi48 with the DIRS-4 retroelement revealed signals on all A chromosomes in the 2 species and also on the B, suggesting the insertion of repetitive elements in these species. PMID:25999244

  20. Cupula-receptor cell relationships with evidence provided by SEM microdissection.

    PubMed

    Barber, V C; Emerson, C J

    1979-01-01

    Ampullae from the inner ear of the skate, Raja ocellata, were examined by SEM to further elucidate the relationships of the sensory hairs of the receptor cells to the overlying cupula. Specimens were prepared by a variety of methods and were fractured to visualize these relationships. Critical point drying resulted in good cellular and sensory hair preservation, but the cupulae were grossly shrunken. Freeze-dried cupulae, in contrast, more closely approximated the in vivo condition. Although tissues suffered freezing damage, sensory hair bundles were readily discerned. In both critical point dried and freeze-dried ampullae, the peripheral hair cell bundles and the kinocilia of most of the central hair cells, extended across the subcupular space and contacted the cupular material. Some of the specimens were further investigated by dissection in the SEM, the process being viewed in real time stereo. Microdissection confirmed that most kinocilia were attached to the cupula. When the cupula was displaced, sensory hair bundles remained attached to it and broke preferentially at their base from the receptor cells.

  1. Microdissection, microchip arrays, and molecular analysis of tumor cells (primary and metastases).

    PubMed

    Pappalardo, P A; Bonner, R; Krizman, D B; Emmert-Buck, M R; Liotta, L A

    1998-07-01

    Advances in biotechnology and bioinformatics are offering promise for new breakthroughs in gene discovery and elucidation of gene function. At present, many candidate genes related to cancer pathogenesis have been identified in several types of human cancer, yet frequently their function remains elusive. This is particularly true as it relates to the progression of human cancer. This landscape could change dramatically, however, as technological innovations and improvements continue to revolutionize these fields. High-throughput molecular approaches are emerging, which may become accurate, automated, and cost-effective. For example, DNA arrays on microchips are under development with numerous applications, including the ability to screen genes rapidly for mutations and to study patterns of gene expression on a large scale. Automated systems for microdissection and sequencing are also in their implementation stages. Commensurate with their integration and evolution, these information and technological tools have the potential to offer a more comprehensive understanding of multiple genetic and cellular alterations occurring during cancer initiation, development, and progression. Ultimately, this fundamental knowledge can provide strategies for intervention, prevention, and early diagnosis. This is a US government work. There are no restrictions on its use.

  2. Plant Vascular Biology 2013: vascular trafficking.

    PubMed

    Ursache, Robertas; Heo, Jung-Ok; Helariutta, Ykä

    2014-04-01

    About 200 researchers from around the world attended the Third International Conference on Plant Vascular Biology (PVB 2013) held in July 2013 at the Rantapuisto Conference Center, in Helsinki, Finland (http://www.pvb2013.org). The plant vascular system, which connects every organ in the mature plant, continues to attract the interest of researchers representing a wide range of disciplines, including development, physiology, systems biology, and computational biology. At the meeting, participants discussed the latest research advances in vascular development, long- and short-distance vascular transport and long-distance signalling in plant defence, in addition to providing a context for how these studies intersect with each other. The meeting provided an opportunity for researchers working across a broad range of fields to share ideas and to discuss future directions in the expanding field of vascular biology. In this report, the latest advances in understanding the mechanism of vascular trafficking presented at the meeting have been summarized.

  3. Integrating Scleral Buckling, Transscleral Drainage of Subretinal Fluid, Intravitreal Anti-Vascular Endothelial Growth Factor, and Laser Photocoagulation in Stage 3B Coats' Total Retinal Detachment.

    PubMed

    Huang, Ying-Chen; Lai, Chi-Chun; Wu, Wei-Chi

    2016-09-01

    The integration of quadruple therapy in a 13-year-old boy with stage 3B Coats' disease achieved retinal reattachment and visual improvement. Scleral buckling might play a role in retinal detachment in Coats' disease, although it has previously been considered insignificant. Instead of performing vitrectomy and internal drainage with a drainage hole in the retina, less-invasive procedures that do not require retinotomy appear to be beneficial in cases of advanced Coats' disease. [Ophthalmic Surg Lasers Imaging Retina. 2016;47:865-868.]. PMID:27631484

  4. Branding of vascular surgery.

    PubMed

    Perler, Bruce A

    2008-03-01

    The Society for Vascular Surgery surveyed primary care physicians (PCPs) to understand how PCPs make referral decisions for their patients with peripheral vascular disease. Responses were received from 250 PCPs in 44 states. More than 80% of the respondents characterized their experiences with vascular surgeons as positive or very positive. PCPs perceive that vascular surgeons perform "invasive" procedures and refer patients with the most severe vascular disease to vascular surgeons but were more than twice as likely to refer patients to cardiologists, believing they are better able to perform minimally invasive procedures. Nevertheless, PCPs are receptive to the notion of increasing referrals to vascular surgeons. A successful branding campaign will require considerable education of referring physicians about the totality of traditional vascular and endovascular care increasingly provided by the contemporary vascular surgical practice and will be most effective at the local grassroots level.

  5. Alteration with dietary state of the activity and zonal distribution of adenylate cyclase stimulated by glucagon, fluoride and forskolin in microdissected rat liver tissue.

    PubMed

    Zierz, S; Jungermann, K

    1984-12-17

    Adenylate cyclase activated by glucagon, fluoride and forskolin was measured in liver homogenates and microdissected periportal and perivenous tissue of fed and fasted rats. A radiochemical microtest, more sensitive by 2-3 orders of magnitude as compared with the usual assay, was established for the determination of the activity in liver samples corresponding to 200-600 ng dry weight. In liver homogenates from fasted as compared to fed animals the glucagon-stimulated and fluoride-stimulated activity was increased by 1.65-fold, while the basal and the forskolin-stimulated activity remained the same. In microdissected tissue of both fed and fasted animals the activity was stimulated in about 60% of the samples by glucagon, fluoride and forskolin (responsive samples). However, in about 40% of the microdissected tissue samples the activity could not be stimulated by any of the above activators (non-responsive samples). In responsive microdissected tissue of fasted as compared to fed animals, the glucagon-stimulated and fluoride stimulated activity but not the basal and the forskolin-activated activity was increased by 2-3-fold. In responsive microdissected samples of fed animals neither the basal nor the stimulated activities showed a significant periportal to perivenous gradient. In samples of fasted animals, however, a zonal gradient was observed for the glucagon-stimulated activity exhibiting a 1.5-fold higher rate in the perivenous zone.

  6. Different Hormonal Regulation of Cellular Differentiation and Function in Nucellar Projection and Endosperm Transfer Cells: A Microdissection-Based Transcriptome Study of Young Barley Grains1[W

    PubMed Central

    Thiel, Johannes; Weier, Diana; Sreenivasulu, Nese; Strickert, Marc; Weichert, Nicola; Melzer, Michael; Czauderna, Tobias; Wobus, Ulrich; Weber, Hans; Weschke, Winfriede

    2008-01-01

    Nucellar projection (NP) and endosperm transfer cells (ETC) are essential tissues in growing barley (Hordeum vulgare) grains, responsible for nutrient transfer from maternal to filial tissues, endosperm/embryo nutrition, and grain development. A laser microdissection pressure catapulting-based transcriptome analysis was established to study NP and ETC separately using a barley 12K macroarray. A major challenge was to isolate high-quality mRNA from preembedded, fixed tissue while maintaining tissue integrity. We show that probes generated from fixed and embedded tissue sections represent largely the transcriptome (>70%) of nonchemically treated and nonamplified references. In NP, the top-down gradient of cellular differentiation is reflected by the expression of C3HC4-type ubiquitin ligases and different histone genes, cell wall biosynthesis and expansin/extensin genes, as well as genes involved in programmed cell death-related proteolysis coupled to nitrogen remobilization, indicating distinct areas simultaneously undergoing mitosis, cell elongation, and disintegration. Activated gene expression related to gibberellin synthesis and function suggests a regulatory role for gibberellins in establishment of the differentiation gradient. Up-regulation of plasmalemma-intrinsic protein and tonoplast-intrinsic protein genes indicates involvement in nutrient transfer and/or unloading. In ETC, AP2/EREBP-like transcription factors and ethylene functions are transcriptionally activated, a response possibly coupled to activated defense mechanisms. Transcriptional activation of nucleotide sugar metabolism may be attributed to ascorbate synthesis and/or cell wall biosynthesis. These processes are potentially controlled by trehalose-6-P synthase/phosphatase, as suggested by expression of their respective genes. Up-regulation of amino acid permeases in ETC indicates important roles in active nutrient uptake from the apoplastic space into the endosperm. PMID:18784282

  7. Contractile properties of isolated vascular smooth muscle after photoradiation

    SciTech Connect

    Freas, W.; Hart, J.L.; Golightly, D.; McClure, H.; Muldoon, S.M.

    1989-03-01

    The purpose of this study was to characterize the responses of various types of vascular smooth muscle to conditions that would be encountered during photodynamic therapy, namely laser illumination of photosensitizer-pretreated tissue. Vascular smooth muscle obtained from representative canine, rodent, and rabbit vascular beds was cut into rings and placed in organ baths (37 degrees C, aerated with 95% O2-5% CO2). These vessels were pretreated for 30 min with the photosensitizer hematoporphyrin derivative (HpD, 3-30 micrograms/ml) washed, and then exposed to red laser light (633 nm, 1-3.5 mW) for up to 20 min. Under basal tension conditions laser illumination of HpD-pretreated vessels resulted in an increase in tension, whereas laser illumination of vessels not exposed to HpD did not contract. This sustained contraction was not reversed by washing the tissue with fresh Krebs-Ringer solution. Responses to norepinephrine, transmural electrical stimulation, and elevated concentrations of KCl were reduced in blood vessels tested after HpD laser illumination. Laser-induced contractions of canine carotid arteries did not require the presence of an intact vascular endothelium. Vascular effect of these photosensitizers appears to involve the formation of oxygen-derived radicals. This preparation could provide a good model for examining the effects of free radicals on vascular physiology.

  8. Isolation, characterization, and regional mapping of microclones from a human chromosome 21 microdissection library

    SciTech Connect

    Yu, J.; Hartz, J.; Yisheng Xu; Gemmill, R.M.; Patterson, D.; Kao, Faten ); Gemmill, R.M.; Patterson, D.; Kao, Fa-Ten ); Korenberg, J.R. )

    1992-08-01

    Thirty-four unique-sequence microclones were isolated from a previously described microdissection library of human chromosome 21 and were regionally mapped using a cell hybrid mapping panel which consists of six cell hybrids and divides chromosome 21 into eight regions. The mapping results showed that the microclones were unevenly distributed along chromosome 21, with the majority of microclones located in the distal half portion of the long arm, between 21q21.3 and 21qter. The number of unique-sequence clones began to decrease significantly from 21q21.2 to centromere and extending to the short arm. This finding is consistent with those reported in other chromosome 21 libraries. Thus, it may be inferred that the proximal portion of the long arm of chromosome 21 contains higher proportions of repetitive sequences, rather than unique sequences of genes. The microclones were also characterized for insert size and were used to identify the corresponding genomic fragments generated by HindIII. In addition, the authors demonstrated that the microclones with short inserts can be efficiently used to identify YAC (yeast artificial chromosome) clones with large inserts, for increased genomic coverage for high-resolution physical mapping. They also used 200 unique-sequence microclones to screen a human liver cDNA library and identified two cDNA clones which were regionally assigned to the 21q21.3-q22.1 region. Thus, generation of unique-sequence microclones from chromosome 21 appears to be useful to isolate and regionally map many cDNA clones, among which will be candidate genes for important diseases on chromosome 21, including Down syndrome, Alzheimer disease, amyotrophic lateral sclerosis, and one form of epilepsy.

  9. A molecular map of G protein alpha chains in microdissected rat nephron segments.

    PubMed Central

    Senkfor, S I; Johnson, G L; Berl, T

    1993-01-01

    Membrane-associated guanine nucleotide binding proteins regulate many receptor-mediated signals. Heterogeneity of biochemical and functional properties in nephron segments could be due to differences in G protein expression. To ascertain whether such heterogeneity of G proteins is present in various nephron segments, this study examines the distribution and relative abundance of G protein alpha chains in microdissected medullary thick ascending limb, cortical collecting tubules, outer medullary collecting tubules, proximal inner medullary tubules, and distal inner medullary tubules. Reverse transcription and polymerase chain reactions were employed using oligonucleotides encoding highly conserved regions of all known alpha chains. The cDNA was sequenced for alpha chain identification. The alpha i2 versus alpha s distribution was different in the outer medullary collecting tubules, when compared with the medullary thick ascending limb (P < 0.001) or the cortical collecting tubule, the proximal inner medullary tubules, and the distal inner medullary tubules (P < 0.05). These latter four segments did not significantly differ from each other. A similar analysis was applied to the frequently used line of kidney cells, LLC-PK1, whose exact cellular origin remains unclear. Interestingly, we detected both alpha i2 and alpha i3, while only alpha i2 was detected in the rat distal nephron. No alpha o or alpha z reverse transcription PCR products were detected. In contrast alpha 11 and alpha 14 members of the more recently described alpha q family were detected in the outer medullary collecting tubules and the proximal inner medullary tubules, respectively. We conclude that the majority of nephron segments have a relatively constant distribution of G protein alpha chains. Images PMID:8349818

  10. In vivo molecular imaging of colorectal cancer using quantum dots targeted to vascular endothelial growth factor receptor 2 and optical coherence tomography/laser-induced fluorescence dual-modality imaging

    PubMed Central

    Carbary-Ganz, Jordan L.; Welge, Weston A.; Barton, Jennifer K.; Utzinger, Urs

    2015-01-01

    Abstract. Optical coherence tomography/laser induced fluorescence (OCT/LIF) dual-modality imaging allows for minimally invasive, nondestructive endoscopic visualization of colorectal cancer in mice. This technology enables simultaneous longitudinal tracking of morphological (OCT) and biochemical (fluorescence) changes as colorectal cancer develops, compared to current methods of colorectal cancer screening in humans that rely on morphological changes alone. We have shown that QDot655 targeted to vascular endothelial growth factor receptor 2 (QD655-VEGFR2) can be applied to the colon of carcinogen-treated mice and provides significantly increased contrast between the diseased and undiseased tissue with high sensitivity and specificity ex vivo. QD655-VEGFR2 was used in a longitudinal in vivo study to investigate the ability to correlate fluorescence signal to tumor development. QD655-VEGFR2 was applied to the colon of azoxymethane (AOM-) or saline-treated control mice in vivo via lavage. OCT/LIF images of the distal colon were taken at five consecutive time points every three weeks after the final AOM injection. Difficulties in fully flushing unbound contrast agent from the colon led to variable background signal; however, a spatial correlation was found between tumors identified in OCT images, and high fluorescence intensity of the QD655 signal, demonstrating the ability to detect VEGFR2 expressing tumors in vivo. PMID:26397238

  11. In vivo molecular imaging of colorectal cancer using quantum dots targeted to vascular endothelial growth factor receptor 2 and optical coherence tomography/laser-induced fluorescence dual-modality imaging

    NASA Astrophysics Data System (ADS)

    Carbary-Ganz, Jordan L.; Welge, Weston A.; Barton, Jennifer K.; Utzinger, Urs

    2015-09-01

    Optical coherence tomography/laser induced fluorescence (OCT/LIF) dual-modality imaging allows for minimally invasive, nondestructive endoscopic visualization of colorectal cancer in mice. This technology enables simultaneous longitudinal tracking of morphological (OCT) and biochemical (fluorescence) changes as colorectal cancer develops, compared to current methods of colorectal cancer screening in humans that rely on morphological changes alone. We have shown that QDot655 targeted to vascular endothelial growth factor receptor 2 (QD655-VEGFR2) can be applied to the colon of carcinogen-treated mice and provides significantly increased contrast between the diseased and undiseased tissue with high sensitivity and specificity ex vivo. QD655-VEGFR2 was used in a longitudinal in vivo study to investigate the ability to correlate fluorescence signal to tumor development. QD655-VEGFR2 was applied to the colon of azoxymethane (AOM-) or saline-treated control mice in vivo via lavage. OCT/LIF images of the distal colon were taken at five consecutive time points every three weeks after the final AOM injection. Difficulties in fully flushing unbound contrast agent from the colon led to variable background signal; however, a spatial correlation was found between tumors identified in OCT images, and high fluorescence intensity of the QD655 signal, demonstrating the ability to detect VEGFR2 expressing tumors in vivo.

  12. Collagen vascular disease

    MedlinePlus

    ... this page: //medlineplus.gov/ency/article/001223.htm Collagen vascular disease To use the sharing features on ... were previously said to have "connective tissue" or "collagen vascular" disease. We now have names for many ...

  13. [The use of lasers in dermatology].

    PubMed

    Lecocq, C; Pirard, D; del Marmol, V; Berlingin, E

    2013-01-01

    Albert Einstein is undoubtedly the father of lasers. But it is not until 1964 that the first dermatological lasers were introduced. The Nd-YAG laser, the CO2 laser were developed by Kumar Patel. In a 40 year period lasers not only were diversified but have also become safer and miniaturized. This article hopes to strengthen general practionners' and specialist's knowledge of the different categories of available lasers. The most frequently used ones are ablative lasers (CO2-Erbium), vascular lasers (Nd-YAG, KTP, pulsed dye laser) and the pigment lasers (Q-Switched Nd-YAG, Alexandrite). A description of these lasers and their indications in dermatology will be discussed.

  14. Vascular restoration therapy and bioresorbable vascular scaffold

    PubMed Central

    Wang, Yunbing; Zhang, Xingdong

    2014-01-01

    This article describes the evolution of minimally invasive intervention technologies for vascular restoration therapy from early-stage balloon angioplasty in 1970s, metallic bare metal stent and metallic drug-eluting stent technologies in 1990s and 2000s, to bioresorbable vascular scaffold (BVS) technology in large-scale development in recent years. The history, the current stage, the challenges and the future of BVS development are discussed in detail as the best available approach for vascular restoration therapy. The criteria of materials selection, design and processing principles of BVS, and the corresponding clinical trial results are also summarized in this article. PMID:26816624

  15. Multifocal vascular lesions.

    PubMed

    Levin, Laura E; Lauren, Christine T

    2016-03-01

    Multifocal vascular lesions are important to recognize and appropriately diagnose. Generally first noticed on the skin, multifocal vascular lesions may have systemic involvement. Distinguishing among the different types of multifocal vascular lesions is often based on clinical features; however, radiological imaging and/or biopsy are frequently needed to identify distinct features and guide treatment. Knowledge of the systemic associations that can occur with different vascular anomalies may reduce life-threatening complications, such as coagulopathy, bleeding, cardiac compromise, and neurologic sequelae. This review provides a synopsis of the epidemiology, pathogenesis, presentation, workup, and treatment of several well-recognized multifocal vascular tumors and malformations. PMID:27607324

  16. Initiation of vascular development.

    PubMed

    Ohashi-Ito, Kyoko; Fukuda, Hiroo

    2014-06-01

    The initiation of vascular development occurs during embryogenesis and the development of lateral organs, such as lateral roots and leaves. Understanding the mechanism underlying the initiation of vascular development has been an important goal of plant biologists. Auxin flow is a crucial factor involved in the initiation of vascular development. In addition, recent studies have identified key factors that regulate the establishment of vascular initial cells in embryos and roots. In this review, we summarize the recent findings in this field and discuss the initiation of vascular development.

  17. [Novel mechanism for retinal vascular diseases].

    PubMed

    Suzuma, Kiyoshi

    2015-03-01

    I. A new therapeutic target for diabetic retinopathy. Recent reports state that succinate may be an independent retinal angiogenic factor. We evaluated concentrations in vitreous from proliferative diabetic retinopathy (PDR), and found that succinate increased significantly in PDR. Interestingly, levels of succinate from bevacizumab-pre-injected PDR were normal, suggesting that vascular endothelial growth factor (VEGF) had a positive feedback mechanism for succinate since succinate was previously reported to induce VEGF. II. A new understanding of central retinal vein occlusion (CRVO). We evaluated retinal blood flow velocity with laser speckle flowgraphy (LSFG) made in Japan, and found that cases in which both macular edema and retinal blood flow velocity improved after anti-VEGF therapy had better prognosis. In ischemic CRVO at final visit, mean retinal blood velocity was less than 50% of fellow eyes after 1st anti-VEGF therapy, suggesting that those cases might have poor prognosis. LSFG is useful for evaluation and decision in CRVO treatment. III. From exploration for mechanism in retinal vascular diseases to re-vascularization therapy. The standard treatment for retinal non-perfusion area is scatter laser photocoagulation, which is both invasive of the peripheral retina and may prove destructive. Re-vascularization is an ideal strategy for treatment of retinal non-perfusion area. To develop a new methods for re-vascularization in retinal non-perfusion area, we have designed experiments using a retina without vasculature differentiated from induced pluripotent stem(iPS) cells.

  18. Cutaneous laser therapy.

    PubMed

    Dixon, J A; Gilbertson, J J

    1985-12-01

    The carbon dioxide laser is useful for vaporizing lesions and applying incisions, the argon laser coagulates superficial vascular lesions and the neodymium-YAG laser is used for large vascular and more deeply situated lesions. Many patients with port-wine stains have been treated with excellent to poor results, major problems consisting of incomplete color removal and hypertrophic scarring (occurring in 4% to 23% of cases). While results are imperfect, patients are satisfied with the improvement in 86% of cases. Lasers have been used with good results for treating patients with strawberry angioma of infancy, pyogenic granuloma, telangiectasia of the face, decorative tattoos, genital condylomata and warts. The results of laser treatment of essential telangiectasia of the lower extremities have generally been poor. The CO(2) laser has been effective in excising small lesions and elevating skin flaps. PMID:4090490

  19. Tissue Microdissection and Degenerate Oligonucleotide Primed-Polymerase Chain Reaction (DOP-PCR) Is an Effective Method to Analyze Genetic Aberrations in Invasive Tumors

    PubMed Central

    Hirose, Yuichi; Aldape, Kenneth; Takahashi, Michelle; Berger, Mitchel S.; Feuerstein, Burt G.

    2001-01-01

    We amplified various amounts of DNA derived from frozen SF210 and U251NCI human glioblastoma cells, carried out comparative genomic hybridization (CGH) using degenerate oligonucleotide primed-PCR (DOP-PCR) products as test probes, and compared results to analyses performed with probes prepared by standard nick translation. Next we extracted DNA from hematoxylin-eosin (HE)- and methyl green (MG)-stained, microdissected sections of formalin-fixed and paraffin-embedded U251NCI cells, amplified and labeled it by DOP-PCR, and subjected it to CGH. Finally, we used the same methods in multiple samples from a single human mixed glioma tissue. DOP-PCR products from 50 pg to 250 ng of DNA were equally effective in generating the same CGH profiles as the standard method. DOP-PCR products from microdissected pieces of MG-stained cells were effective probes for CGH, but HE-stained samples were not desirable. As the proportion of HE-stained sample increased, CGH profiles deteriorated. DOP-PCR products from microdissected pieces of MG-stained paraffin sections of glioma tissue produced CGH profiles compatible with their histological features. CGH performed with DOP-PCR products from microdissected paraffin blocks allows for the accurate investigation of the cytogenetic characteristics from invasive tumors and of cytogenetic heterogeneity within neoplastic tissue. PMID:11333301

  20. Imaging Pediatric Vascular Lesions

    PubMed Central

    Nguyen, Tuyet A.; Krakowski, Andrew C.; Naheedy, John H.; Kruk, Peter G.

    2015-01-01

    Vascular anomalies are commonly encountered in pediatric and dermatology practices. Most of these lesions are benign and easy to diagnose based on history and clinical exam alone. However, in some cases the diagnosis may not be clear. This may be of particular concern given that vascular anomalies may occasionally be associated with an underlying syndrome, congenital disease, or serious, life-threatening condition. Defining the type of vascular lesion early and correctly is particularly important to determine the optimal approach to management and treatment of each patient. The care of pediatric patients often requires collaboration from a multitude of specialties including pediatrics, dermatology, plastic surgery, radiology, ophthalmology, and neurology. Although early characterization of vascular lesions is important, consensus guidelines regarding the evaluation and imaging of vascular anomalies does not exist to date. Here, the authors provide an overview of pediatric vascular lesions, current classification systems for characterizing these lesions, the various imaging modalities available, and recommendations for appropriate imaging evaluation. PMID:26705446

  1. Mechanisms of Laser-Induced Dissection and Transport of Histologic Specimens

    PubMed Central

    Vogel, Alfred; Lorenz, Kathrin; Horneffer, Verena; Hüttmann, Gereon; von Smolinski, Dorthe; Gebert, Andreas

    2007-01-01

    Rapid contact- and contamination-free procurement of histologic material for proteomic and genomic analysis can be achieved by laser microdissection of the sample of interest followed by laser-induced transport (laser pressure catapulting). The dynamics of laser microdissection and laser pressure catapulting of histologic samples of 80 μm diameter was investigated by means of time-resolved photography. The working mechanism of microdissection was found to be plasma-mediated ablation initiated by linear absorption. Catapulting was driven by plasma formation when tightly focused pulses were used, and by photothermal ablation at the bottom of the sample when defocused pulses producing laser spot diameters larger than 35 μm were used. With focused pulses, driving pressures of several hundred MPa accelerated the specimen to initial velocities of 100–300 m/s before they were rapidly slowed down by air friction. When the laser spot was increased to a size comparable to or larger than the sample diameter, both driving pressure and flight velocity decreased considerably. Based on a characterization of the thermal and optical properties of the histologic specimens and supporting materials used, we calculated the evolution of the heat distribution in the sample. Selected catapulted samples were examined by scanning electron microscopy or analyzed by real-time reverse-transcriptase polymerase chain reaction. We found that catapulting of dissected samples results in little collateral damage when the laser pulses are either tightly focused or when the laser spot size is comparable to the specimen size. By contrast, moderate defocusing with spot sizes up to one-third of the specimen diameter may involve significant heat and ultraviolet exposure. Potential side effects are maximal when samples are catapulted directly from a glass slide without a supporting polymer foil. PMID:17766336

  2. Application of array CGH on archival formalin-fixed paraffin-embedded tissues including small numbers of microdissected cells.

    PubMed

    Johnson, Nicola A; Hamoudi, Rifat A; Ichimura, Koichi; Liu, Lu; Pearson, Danita M; Collins, V Peter; Du, Ming-Qing

    2006-09-01

    Array-based comparative genomic hybridisation (aCGH) has diverse applications in cancer gene discovery and translational research. Currently, aCGH is performed primarily using high molecular weight DNA samples and its application to formalin-fixed and paraffin-embedded (FFPE) tissues remains to be established. To explore how aCGH can be reliably applied to archival FFPE tissues and whether it is possible to apply aCGH to small numbers of cells microdissected from FFPE tissue sections, we have systematically performed aCGH on 15 pairs of matched frozen and FFPE astrocytic tumour tissues using a well-established in-house human 1 Mb BAC/PAC genomic array. By spiking tumour DNA with normal DNA, we demonstrated that at least 70% of tumour DNA was required for reliable aCGH analysis. Using aCGH data from frozen tissue as a reference, it was found that only FFPE astrocytic tumour tissues that supported PCR amplification of >300 bp DNA fragment provided high quality, reproducible aCGH data. The presence of necrosis in a tissue specimen had an adverse effect on the quality of aCGH, while fixation in formalin for up to 96 h of fresh tissue did not appear to affect the quality of the result. As little as 10-20 ng DNA from frozen or FFPE tissues could be readily used for aCGH analysis following whole genome amplification (WGA). Furthermore, as few as 2000 microdissected cells from haematoxylin-stained slides of archival FFPE tissues could be successfully used for aCGH investigations when WGA was used. By careful assessment of DNA integrity and review of histology, to exclude necrosis and select specimens with a high proportion of tumour cells, it is feasible to preselect archival FFPE tissues adequate for aCGH analysis. With the help of microdissection and WGA, it is also possible to apply aCGH to histologically defined lesions, such as carcinoma in situ.

  3. Microdissected double-minute DNA detects variable patterns of chromosomal localizations and multiple abundantly expressed transcripts in normal and leukemic cells

    SciTech Connect

    Sen, S.; Zhou, Hongyi; Stass, S.A.; Sen, P. ); Mulac-Jericevic, B.; Pirrotta, V. )

    1994-02-01

    Double-minute (dm) chromosomes are cytogenetically resolvable DNA amplification-mediating acentric extrachromosomal structures that are commonly seen in primary tumors, tumor cell lines, and drug-resistant cells grown in vitro. Selective isolation of dm DNAs with standard molecular biological techniques is difficult, and thus, detailed studies to elucidate their structure, site of chromosomal origin, and chromosomal reintegration patterns have been limited. In those instances in which a gene has been localized on dms, characterization of the remainder of the DNA, which far exceeds the size of the gene identified, has remained inconclusive. dms seen in the acute myeloid leukemia cell line HL-60 have been shown to harbor the c-myc protooncogene. In this paper, the authors report the successful isolation of the dm-specific DNAs from these cells by the microdissection/polymerase chain reaction technique and demonstrate that the dm DNAs derived from a single discrete normal chromosome segment 8q24.1-q24.2 reintegrate at various specific locations in the leukemic cells. The microdissected dm DNA detects multiple abundantly expressed transcripts distinct from c-myc mRNA on Northern blots. By devising a [open quotes]transcript selection[close quotes] strategy, they cloned the partial genomic sequence of a gene from the microdissected DNA that encodes two of these RNAs. This strategy will be generally applicable for rapid cloning of unknown amplified genes harbored on dms. With DNA from 20 microdissected dms, they constructed a genomic library of about 20,000 recombinant microclones with an average insert size of about 450 bp. The microclones should help in isolating corresponding yeast artificial chromosome clones for high-resolution physical mapping of dms in HL-60 cells. Furthermore, application of the microdissection technique appears to be an extremely feasible approach to characterization of dms in other cell types. 42 refs., 6 figs., 1 tab.

  4. Digital holographic microscopy for quantitative cell dynamic evaluation during laser microsurgery

    PubMed Central

    Yu, Lingfeng; Mohanty, Samarendra; Zhang, Jun; Genc, Suzanne; Kim, Myung K.; Berns, Michael W.; Chen, Zhongping

    2010-01-01

    Digital holographic microscopy allows determination of dynamic changes in the optical thickness profile of a transparent object with subwavelength accuracy. Here, we report a quantitative phase laser microsurgery system for evaluation of cellular/ sub-cellular dynamic changes during laser micro-dissection. The proposed method takes advantage of the precise optical manipulation by the laser microbeam and quantitative phase imaging by digital holographic microscopy with high spatial and temporal resolution. This system will permit quantitative evaluation of the damage and/or the repair of the cell or cell organelles in real time. PMID:19582118

  5. Microdissection specimens of connective, chondrous, or Bone Tissue of human osteosarcomas and chondrosarcomas transplanted to athymic nude mice.

    PubMed

    Nilsson, O S; Lindholm, T S; Nilsonne, U

    1982-01-01

    Five osteosarcomas and two chondrosarcomas were microdissected to separate tumor compartments of calcified, chondrous, and connective tissue. The compartments were lyophilized separately and transplanted subcutaneously or intramuscularly into nude mice for three, four, and five weeks, respectively. In three of the osteosarcomas and in one of the chrondrosarcomas, calcified tissue induced ectopic bone formation by the host, while cartilaginous tissue induced ectopic bone formation in one of the osteosarcomas and in one of the chondrosarcomas. The tumor-derived connective tissue did not induce osteogenic response in the host tissue. Thus, the ability to develop an osteoinductive response and to produce bone morphogenetic protein seems to be restricted to the population of cells that eventually will, or have, differentiated into bone or cartilage.

  6. Society for Vascular Medicine

    MedlinePlus

    ... Sessions June 14-17, 2017 Sheraton New Orleans New Orleans, LA USA Board Review Course June 16-18, 2017 SVM in the Vascular Lab June 17, 2017 Learn more Patient Information Pages from Vascular Medicine October 2016 Smoking Cessation More info for patients. SVM Case ...

  7. [Vascular graft prosthesis].

    PubMed

    Chakfé, N; Dieval, F; Thaveau, F; Rinckenbach, S; Hassani, O; Camelot, G; Durand, B; Kretz, J-G

    2004-06-01

    Performed since the 1950s, vascular grafting has opened modern era of vascular surgery. Autologous venous grafts are of first choice for revascularisation of small arteries. Synthetic grafts are mainly modelled using microporous polytetrafluoroethylene or terephtalate polyethylene. These prosthesis are mainly used for revascularization of medium and large size arteries. PMID:15220107

  8. [Vascular factors in glaucoma].

    PubMed

    Mottet, B; Aptel, F; Geiser, M; Romanet, J P; Chiquet, C

    2015-12-01

    The exact pathophysiology of glaucoma is not fully understood. Understanding of the vascular pathophysiology of glaucoma requires: knowing the techniques for measuring ocular blood flow and characterizing the topography of vascular disease and the mechanisms involved in this neuropathy. A decreased mean ocular perfusion pressure and a loss of vascular autoregulation are implicated in glaucomatous disease. Early decrease in ocular blood flow has been identified in primary open-angle glaucoma and normal pressure glaucoma, contributing to the progression of optic neuropathy. The vascular damage associated with glaucoma is present in various vascular territories within the eye (from the ophthalmic artery to the retina) and is characterized by a decrease in basal blood flow associated with a dysfunction of vasoregulation.

  9. [Vascular factors in glaucoma].

    PubMed

    Mottet, B; Aptel, F; Geiser, M; Romanet, J P; Chiquet, C

    2015-12-01

    The exact pathophysiology of glaucoma is not fully understood. Understanding of the vascular pathophysiology of glaucoma requires: knowing the techniques for measuring ocular blood flow and characterizing the topography of vascular disease and the mechanisms involved in this neuropathy. A decreased mean ocular perfusion pressure and a loss of vascular autoregulation are implicated in glaucomatous disease. Early decrease in ocular blood flow has been identified in primary open-angle glaucoma and normal pressure glaucoma, contributing to the progression of optic neuropathy. The vascular damage associated with glaucoma is present in various vascular territories within the eye (from the ophthalmic artery to the retina) and is characterized by a decrease in basal blood flow associated with a dysfunction of vasoregulation. PMID:26597554

  10. Laser photochemotherapy of psoriasis

    NASA Astrophysics Data System (ADS)

    Tuchin, Valery V.; Utz, Sergei R.; Barabanov, Alexander Y.; Dovzansky, S. I.; Ulyanov, A. N.; Aravin, Vladislav A.; Khomutova, T. G.

    1991-06-01

    Over the last seven years, I and several co-investigators have studied the vascular welding potential of the milliwatt CO2 laser. It has become clear that there are definite limitations to this method of vascular repair which has hindered its advancement into the clinical arena. The thrust of this review is to highlight these limitations and thereby clarify what future direction might be considered.

  11. Vascular Access in Children

    SciTech Connect

    Krishnamurthy, Ganesh Keller, Marc S.

    2011-02-15

    Establishment of stable vascular access is one of the essential and most challenging procedures in a pediatric hospital. Many clinical specialties provide vascular service in a pediatric hospital. At the top of the 'expert procedural pyramid' is the pediatric interventional radiologist, who is best suited and trained to deliver this service. Growing awareness regarding the safety and high success rate of vascular access using image guidance has led to increased demand from clinicians to provide around-the-clock vascular access service by pediatric interventional radiologists. Hence, the success of a vascular access program, with the pediatric interventional radiologist as the key provider, is challenging, and a coordinated multidisciplinary team effort is essential for success. However, there are few dedicated pediatric interventional radiologists across the globe, and also only a couple of training programs exist for pediatric interventions. This article gives an overview of the technical aspects of pediatric vascular access and provides useful tips for obtaining vascular access in children safely and successfully using image guidance.

  12. Laser micromanipulation systems as universal tools in cellular and molecular biology and in medicine.

    PubMed

    Schütze, K; Pösl, H; Lahr, G

    1998-07-01

    The UV-laser microbeam has been established as a valuable tool in a wide area of molecular biology as well as in medical research and applications. This system allows to cut or fuse microscopically small specimen. An important application of the cutting laser is laser microbeam microdissection (LMM) combined with laser pressure catapulting (LPC), which allows to procure single cells or small homogeneous cell areas for subsequent molecular analysis in an entirely "non-contact" manner. With LMM minute tissue areas, single cells or chromosomes are microdissected and separated from their surroundings. Subsequently, LPC ejects the dissectates directly into the cap of a sample tube without any mechanical contact. This enables the rapid procurement of homogeneous specimen from less than one up to several hundreds of micrometers in diameter without encroachment of the adjacent region. The mRNA information of the selected specimen as well as of the remaining probe are well preserved, as demonstrated with laser isolated samples from a routinely prepared tissue section of a differentiated colorectal adenocarcinoma. Reverse transcription of specific mRNA coding for cytoplasmic beta-actin and subsequent hemi-nested PCR amplification was not impaired. Any kind of tissue, as well as single cells from different sources and even subcellular structures can be captured using this laser method. Wherever homogeneous samples are required to analyze cell or chromosome-specific genetic alterations such as in cancer research or prenatal diagnosis this unique and rapid laser micropreparation method will become a key technology of great value.

  13. [Vascular factors in dementia].

    PubMed

    Bidzan, Leszek

    2005-01-01

    Cerebrovascular factors are a common cause of dementia or contribute to cognitive decline in other dementias. Studies showing that cerebrovascular factors are the risk factors for neurodegenerative dementias, especially Alzheimer's disease. Practically all neurodegenerative dementias have a vascular component that reduces cerebral perfusion and has great impact on the clinical picture. Recent data support the view that the neurodegenerative process is caused by cerebrovascular mechanisms. The results showed that patients with vascular cognitive impairment have a typical clinical picture. Various important non-cognitive features are caused by cerebrovascular factors and are associated with a more rapid course of illness. On the other hand the term vascular diseases or cerebrovascular factors include a variety of vascular pathologies. PMID:16358596

  14. Vascular ring (image)

    MedlinePlus

    Vascular ring is a term used to describe a number of abnormal formations of the aorta, the large artery ... the pulmonary artery. The abnormal vessel(s) forms a ring, which encircles and may press down on the ...

  15. Heart and vascular services

    MedlinePlus

    ... scan of the heart Stress tests (many different types of stress tests exist) Vascular ultrasound, such as carotid ultrasound Venous ultrasound of the arms and legs SURGERIES AND INTERVENTIONS ... these types of procedures, a catheter is inserted through the ...

  16. Vascular Disease Foundation

    MedlinePlus

    ... or 911 immediately. @ 2016 Vascular Cures is a tax-exempt, nonprofit organization tax ID#: 94-2825216 as described in the Section ... 3) of the Internal Revenue Code. Donations are tax deductible. 555 Price Ave., Suite 180, Redwood City, ...

  17. Implications of Vascular Aging

    PubMed Central

    Barodka, Viachaslau M.; Joshi, Brijen L.; Berkowitz, Dan E.; Hogue, Charles W.; Nyhan, Daniel

    2011-01-01

    Chronological age is a well established risk factor for the development of cardiovascular diseases. The changes that accumulate in the vasculature with age, though, are highly variable. It is now increasingly recognized that indices of vascular health are more reliable than age per se in predicting adverse cardiovascular outcomes. The variation in the accrual of these age-related vascular changes is a function of multiple genetic and environmental factors. In this review, we highlight some of the pathophysiological mechanisms that characterize the vascular aging phenotype. Furthermore, we provide an overview of the key outcome studies that address the value of these vascular health indices in general and discuss potential effects on perioperative cardiovascular outcomes. PMID:21474663

  18. Sinuosities in vascular structures

    NASA Astrophysics Data System (ADS)

    Masson, J.-B.; Martin, J.-L.

    2007-12-01

    In most organs, depending on the scale, the nature of the heart pump, the geometry and topology of the organ, some of the blood vessels tend to exhibit sinuous trajectories. We describe a part of this sinuous behavior, including partial biological and strong physical effects in a global physical framework. We will voluntarily focus on physical and topological effects. This study is performed on the vitelline membrane of the chicken embryo. Crossing angles, sinuosity, and the oscillation amplitude of the vascular system are analyzed. Surprisingly, the equation of river meandering dynamics is found to model the sinuosities in the vascular system, and an extension of this equation to non planar case is able to explain the effect of tissue global curvature on the vascular system. Results of this study could lead to a new understanding of the interplay between biological signaling and physical effects in determining the vascular pattern in different tissues.

  19. What Is Vascular Disease?

    MedlinePlus

    ... or 911 immediately. @ 2016 Vascular Cures is a tax-exempt, nonprofit organization tax ID#: 94-2825216 as described in the Section ... 3) of the Internal Revenue Code. Donations are tax deductible. 555 Price Ave., Suite 180, Redwood City, ...

  20. Women and Vascular Disease

    MedlinePlus

    ... Search Patient information Membership Directory (SIR login) Interventional Radiology Women and Vascular Disease Early Warning Symptom for ... major public health issue, the Society of Interventional Radiology recommends greater screening efforts by the medical community ...

  1. Vascular structures in dermoscopy*

    PubMed Central

    Ayhan, Erhan; Ucmak, Derya; Akkurt, ZeynepMeltem

    2015-01-01

    Dermoscopy is an aiding method in the visualization of the epidermis and dermis. It is usually used to diagnose melanocytic lesions. In recent years, dermoscopy has increasingly been used to diagnose non-melanocytic lesions. Certain vascular structures, their patterns of arrangement and additional criteria may demonstrate lesion-specific characteristics. In this review, vascular structures and their arrangements are discussed separately in the light of conflicting views and an overview of recent literature. PMID:26375224

  2. Vascular Hyperpermeability and Aging

    PubMed Central

    Oakley, Ryan; Tharakan, Binu

    2014-01-01

    Vascular hyperpermeability, the excessive leakage of fluid and proteins from blood vessels to the interstitial space, commonly occurs in traumatic and ischemic injuries. This hyperpermeability causes tissue vasogenic edema, which often leads to multiple organ failure resulting in patient death. Vascular hyperpermeability occurs most readily in small blood vessels as their more delicate physical constitution makes them an easy target for barrier dysfunction. A single layer of endothelial cells, linked to one another by cell adhesion molecules, covers the interior surface of each blood vessel. The cell adhesion molecules play a key role in maintaining barrier functions like the regulation of permeability. Aging is a major risk factor for microvascular dysfunction and hyperpermeability. Apart from age-related remodeling of the vascular wall, endothelial barrier integrity and function declines with the advancement of age. Studies that address the physiological and molecular basis of vascular permeability regulation in aging are currently very limited. There have been many cellular and molecular mechanisms proposed to explain aging-related endothelial dysfunction but their true relationship to barrier dysfunction and hyperpermeability is not clearly known. Among the several mechanisms that promote vascular dysfunction and hyperpermeability, the following are considered major contributors: oxidative stress, inflammation, and the activation of apoptotic signaling pathways. In this review we highlighted (a) the physiological, cellular and molecular changes that occur in the vascular system as a product of aging; (b) the potential mechanisms by which aging leads to barrier dysfunction and vascular hyperpermeability in the peripheral and the blood-brain barrier; (c) the mechanisms by which the age-related increases in oxidative stress, inflammatory markers and apoptotic signaling etc. cause endothelial dysfunction and their relationship to hyperpermeability; and (d) the

  3. [Lasers and aesthetic dermatology].

    PubMed

    Stratigos, A J; Dover, J S; Arndt, K A

    2003-07-01

    The improved understanding of laser-tissue interaction along with the latest advances of laser technology have led to the development of sophisticated, safe, and user-friendly laser systems that provide effective treatment for a variety of aesthetic skin conditions. The use of lasers and their tissue-specific capabilities in the treatment of pigmented and vascular lesions has been greatly expanded to include rhytides, photoaged skin, atrophic scars, and unwanted hair. In addition, laser techniques have been employed in traditional "rejuvenating" procedures of aged skin, e.g., face-lifting, blepharoplasty, and hair transplantation, decreasing the intra-operative time and limiting the recovery period. These advances have led to a wide acceptance of cutaneous laser surgery by the dermatologic community and have created an increasing popularity among the public. The purpose of this article is to review the applications of lasers in aesthetic dermatology and discuss their limitations and potential side effects. PMID:12835862

  4. Assessing vascular dementia.

    PubMed

    Forette, F; Rigaud, A S; Morin, M; Gisselbrecht, M; Bert, P

    1995-10-01

    Vascular dementia is the most common cause of dementia in the elderly after Alzheimer's disease. Many forms of vascular dementia have been described: multi-infarct dementia, lacunar dementia, Binswanger's subcortical encephalopathy, cerebral amyloid angiopathy, white matter lesions associated with dementias, single infarct dementia, dementia linked to hypoperfusion and haemorrhagic dementia. The difficulty of diagnosing vascular dementia must not be underestimated and an international consensus is needed for epidemiological studies. The NINCDS-AIREN group has recently published diagnostic criteria. The State of California Alzheimer's Disease Diagnostic and Treatment Centers also proposed some which differ from the NINCDS-AIREN criteria in considering only ischaemic vascular dementia and not other mechanisms such as haemorrhagic or hypoxic lesions. Most studies stress hypertension as the most powerful risk factor for all forms of vascular dementia. The incidence rate ranges from 7 per 1000 person-years in normal volunteers to 16 per 1000 person-years in hypertensive patients. No therapeutic attempt has influenced the course of the disease once the dementing condition is established. The only effective approach is preventive treatment. The objective of the SYST-EUR Vascular Dementia project is to confirm that the treatment of isolated systolic hypertension is able to reduce its incidence.

  5. Isolation of region-specific cosmids by hybridization with microdissection clones from human chromosome 10q11. 1-q21. 1

    SciTech Connect

    Karakawa, Katsu; Takami, Koji; Fujita, Shoichi Osaka Univ. Medical School, Fukushima-ku, Osaka ); Nakamura, Tsutomu; Takai, Shin-ichiro; Nishisho, Isamu ); Jones, C. ); Ohta, Tohru; Jinno, Yoshihiro; Niikawa, Norio )

    1993-08-01

    A region-specific plasmid library composed of 20,000 recombinants was constructed by microdissection of human chromosome 10 (10q11.2-q21.1) and subsequent amplification with the primer-linker method of polymerase chain reaction (PCR). Hybridization with total human DNA showed that 32 of 217 microclones studied contained highly repetitive sequences. Further analysis of the remaining 185 microclones proved that 43 microclones, each having an insert longer than 200 bp, contained unique sequences of human chromosome 10 origin. Twenty-five microclones randomly selected from the 43 were used directly as probes to isolate corresponding cosmid clones, resulting in 32 cosmids corresponding to 14 microclones. Of the 25 cosmids that could be mapped by fluorescence in situ hybridization, 24 proved to originate from the microdissected or adjacent region (10p11.2-q22.3)and 1 from a rather distal region (10q24.3-q25.1). In addition, 15 of the 32 cosmids revealed restriction fragment length polymorphisms, including 1 with a variable number of tandem repeats marker. The microdissection library and the obtained cosmids are valuable resources for constructing high-resolution physical and linkage maps of the pericentromeric region of chromosome 10, where the gene predisposing to multiple endocrine neoplasia type 2A (MEN2A) has been mapped. 30 refs., 3 figs., 3 tabs.

  6. Antioxidants and vascular health.

    PubMed

    Bielli, Alessandra; Scioli, Maria Giovanna; Mazzaglia, Donatella; Doldo, Elena; Orlandi, Augusto

    2015-12-15

    Oxygen free radicals and other reactive oxygen species (ROS) are common products of normal aerobic cellular metabolism, but high levels of ROS lead to oxidative stress and cellular damage. Increased production of ROS favors vascular dysfunction, inducing altered vascular permeability and inflammation, accompanied by the loss of vascular modulatory function, the imbalance between vasorelaxation and vasoconstriction, and the aberrant expression of inflammatory adhesion molecules. Inflammatory stimuli promote oxidative stress generated from the increased activity of mitochondrial nicotinamide adenine dinucleotide phosphate oxidase, particularly of the Nox4 isoform, with the consequent impairment of mitochondrial β-oxidation. Vascular dysfunction due to the increase in Nox4 activity and ROS overproduction leads to the progression of cardiovascular diseases, diabetes, inflammatory bowel disease, and neurological disorders. Considerable research into the development of effective antioxidant therapies using natural derivatives or new synthetic molecules has been conducted. Antioxidants may prevent cellular damage by reducing ROS overproduction or interfering in reactions that involve ROS. Vitamin E and ascorbic acid are well known as natural antioxidants that counteract lipid peroxidative damage by scavenging oxygen-derived free radicals, thus restoring vascular function. Recently, preliminary studies on natural antioxidants such as goji berries, thymus, rosemary, green tea ginseng, and garlic have been conducted for their efficacy in preventing vascular damage. N-acetyl-cysteine and propionyl-L-carnitine are synthetic compounds that regulate ROS production by replacing endogenous antioxidants in both endothelial and smooth muscle cells. In this review, we consider the molecular mechanisms underlying the generation of oxidative stress-induced vascular dysfunction as well as the beneficial effects of antioxidant therapies.

  7. Nanoengineering of therapeutics for retinal vascular disease.

    PubMed

    Gahlaut, Nivriti; Suarez, Sandra; Uddin, Md Imam; Gordon, Andrew Y; Evans, Stephanie M; Jayagopal, Ashwath

    2015-09-01

    Retinal vascular diseases, including diabetic retinopathy, neovascular age related macular degeneration, and retinal vein occlusion, are leading causes of blindness in the Western world. These diseases share several common disease mechanisms, including vascular endothelial growth factor (VEGF) signaling, hypoxia, and inflammation, which provide opportunities for common therapeutic strategies. Treatment of these diseases using laser therapy, anti-VEGF injections, and/or steroids has significantly improved clinical outcomes. However, these strategies do not address the underlying root causes of pathology, and may have deleterious side effects. Furthermore, many patients continue to progress toward legal blindness despite receiving regular therapy. Nanomedicine, the engineering of therapeutics at the 1-100 nm scale, is a promising approach for improving clinical management of retinal vascular diseases. Nanomedicine-based technologies have the potential to revolutionize the treatment of ophthalmology, through enabling sustained release of drugs over several months, reducing side effects due to specific targeting of dysfunctional cells, and interfacing with currently "undruggable" targets. We will discuss emerging nanomedicine-based applications for the treatment of complications associated with retinal vascular diseases, including angiogenesis and inflammation.

  8. Swept-Source Optical Coherence Tomography Angiography and Vascular Perfusion Map Findings in Obstructive Sleep Apnea.

    PubMed

    Li, Daniel Q; Golding, John; Choudhry, Netan

    2016-09-01

    Obstructive sleep apnea (OSA) is a highly prevalent chronic sleep disorder associated with considerable systemic and ophthalmic consequences. The authors present the retinal vascular findings of a visually asymptomatic 56-year-old man clinically diagnosed with OSA using swept-source optical coherence tomography and vascular perfusion mapping. [Ophthalmic Surg Lasers Imaging Retina. 2016;47:880-884.]. PMID:27631487

  9. In vivo microdissection and live embryo imaging by two-photon microscopy to study Drosophila melanogaster early development

    NASA Astrophysics Data System (ADS)

    Supatto, Willy; Brouzes, Eric; Farge, Emmanuel; Beaurepaire, Emmanuel

    2004-09-01

    Animal embryo development exhibits a complex choreography of cell movements highly regulated both in time and space. This sequence of morphogenetic movements is initiated at gastrulation and is tightly controlled by a cascade of developmental gene expression. We have recently reported that developmental gene expression can in turn be mechanically regulated by morphogenetic movements during Drosophila melanogaster early development. In order to study this phenomenon of mechanically induced gene expression, it is necessary to develop new techniques of in vivo investigation. We show that the combination of femtosecond pulse intratissue surgery and two-photon-excitation fluorescence (2PEF) microscopy is a powerful tool for (i) disrupting natural morphogenetic movements and (ii) imaging native and disrupted morphogenetic movements during Drosophila development. (i) First, non-linear-absorption-mediated photo-disruption makes it possible to perform controlled intra-vital micro-dissections resulting in the modulation of morphogenetic movements and subsequent mechano-sensitive gene expression. (ii) Second, in vivo 2PEF microscopy of transgenic GFP systems appears to be an excellent technique for long-term in vivo imaging of the complex morphogenetic movements involved in normal or perturbed Drosophila gastrulation. Together, these two techniques provide a powerful novel approach to study embryo development.

  10. Micro-dissected tumor tissues on chip: an ex vivo method for drug testing and personalized therapy.

    PubMed

    Astolfi, M; Péant, B; Lateef, M A; Rousset, N; Kendall-Dupont, J; Carmona, E; Monet, F; Saad, F; Provencher, D; Mes-Masson, A-M; Gervais, T

    2016-01-21

    In cancer research and personalized medicine, new tissue culture models are needed to better predict the response of patients to therapies. With a concern for the small volume of tissue typically obtained through a biopsy, we describe a method to reproducibly section live tumor tissue to submillimeter sizes. These micro-dissected tissues (MDTs) share with spheroids the advantages of being easily manipulated on-chip and kept alive for periods extending over one week, while being biologically relevant for numerous assays. At dimensions below ~420 μm in diameter, as suggested by a simple metabolite transport model and confirmed experimentally, continuous perfusion is not required to keep samples alive, considerably simplifying the technical challenges. For the long-term culture of MDTs, we describe a simple microfluidic platform that can reliably trap samples in a low shear stress environment. We report the analysis of MDT viability for eight different types of tissues (four mouse xenografts derived from human cancer cell lines, three from ovarian and prostate cancer patients, and one from a patient with benign prostatic hyperplasia) analyzed by both confocal microscopy and flow cytometry over an 8-day incubation period. Finally, we provide a proof of principle for chemosensitivity testing of human tissue from a cancer patient performed using the described MDT chip method. This technology has the potential to improve treatment success rates by identifying potential responders earlier during the course of treatment and providing opportunities for direct drug testing on patient tissues in early drug development stages.

  11. Building Vascular Networks

    PubMed Central

    Bae, Hojae; Puranik, Amey S.; Gauvin, Robert; Edalat, Faramarz; Carrillo-Conde, Brenda; Peppas, Nicholas A.; Khademhosseini, Ali

    2013-01-01

    Only a few engineered tissues—skin, cartilage, bladder—have achieved clinical success, and biomaterials designed to replace more complex organs are still far from commercial availability. This gap exists in part because biomaterials lack a vascular network to transfer the oxygen and nutrients necessary for survival and integration after transplantation. Thus, generation of a functional vasculature is essential to the clinical success of engineered tissue constructs and remains a key challenge for regenerative medicine. In this Perspective, we discuss recent advances in vascularization of biomaterials through the use of biochemical modification, exogenous cells, or microengineering technology. PMID:23152325

  12. Vascular wall extracellular matrix proteins and vascular diseases

    PubMed Central

    Xu, Junyan; Shi, Guo-Ping

    2014-01-01

    Extracellular matrix proteins form the basic structure of blood vessels. Along with providing basic structural support to blood vessels, matrix proteins interact with different sets of vascular cells via cell surface integrin or non-integrin receptors. Such interactions induce vascular cell de novo synthesis of new matrix proteins during blood vessel development or remodeling. Under pathological conditions, vascular matrix proteins undergo proteolytic processing, yielding bioactive fragments to influence vascular wall matrix remodeling. Vascular cells also produce alternatively spliced variants that induce vascular cell production of different matrix proteins to interrupt matrix homeostasis, leading to increased blood vessel stiffness; vascular cell migration, proliferation, or death; or vascular wall leakage and rupture. Destruction of vascular matrix proteins leads to vascular cell or blood-borne leukocyte accumulation, proliferation, and neointima formation within the vascular wall; blood vessels prone to uncontrolled enlargement during blood flow diastole; tortuous vein development; and neovascularization from existing pathological tissue microvessels. Here we summarize discoveries related to blood vessel matrix proteins within the past decade from basic and clinical studies in humans and animals — from expression to cross-linking, assembly, and degradation under physiological and vascular pathological conditions, including atherosclerosis, aortic aneurysms, varicose veins, and hypertension. PMID:25045854

  13. Vascular air embolism

    PubMed Central

    Gordy, Stephanie; Rowell, Susan

    2013-01-01

    Vascular air embolism is a rare but potentially fatal event. It may occur in a variety of procedures and surgeries but is most often associated as an iatrogenic complication of central line catheter insertion. This article reviews the incidence, pathophysiology, diagnosis, treatment, and prevention of this phenomenon. PMID:23724390

  14. Adhesion in vascular biology

    PubMed Central

    de Rooij, Johan

    2014-01-01

    The vasculature delivers vital support for all other tissues by supplying oxygen and nutrients for growth and by transporting the immune cells that protect and cure them. Therefore, the microvasculature developed a special barrier that is permissive for gasses like oxygen and carbon dioxide, while fluids are kept inside and pathogens are kept out. While maintaining this tight barrier, the vascular wall also allows immune cells to exit at sites of inflammation or damage, a process that is called transmigration. The endothelial cell layer that forms the inner lining of the vasculature is crucial for the vascular barrier function as well as the regulation of transmigration. Therefore, adhesions between vascular endothelial cells are both tight and dynamic and the mechanisms by which they are established, and the mechanisms by which they are controlled have been extensively studied over the past decades. Because of our fundamental strive to understand biology, but also because defects in vascular barrier control cause a variety of clinical problems and treatment strategies may evolve from our detailed understanding of its mechanisms. This special focus issue features a collection of articles that review key components of the development and control of the endothelial cell-cell junction that is central to endothelial barrier function. PMID:25422845

  15. Engineered Vascularized Muscle Flap.

    PubMed

    Egozi, Dana; Shandalov, Yulia; Freiman, Alina; Rosenfeld, Dekel; Ben-Shimol, David; Levenberg, Shulamit

    2016-01-01

    One of the main factors limiting the thickness of a tissue construct and its consequential viability and applicability in vivo, is the control of oxygen supply to the cell microenvironment, as passive diffusion is limited to a very thin layer. Although various materials have been described to restore the integrity of full-thickness defects of the abdominal wall, no material has yet proved to be optimal, due to low graft vascularization, tissue rejection, infection, or inadequate mechanical properties. This protocol describes a means of engineering a fully vascularized flap, with a thickness relevant for muscle tissue reconstruction. Cell-embedded poly L-lactic acid/poly lactic-co-glycolic acid constructs are implanted around the mouse femoral artery and vein and maintained in vivo for a period of one or two weeks. The vascularized graft is then transferred as a flap towards a full thickness defect made in the abdomen. This technique replaces the need for autologous tissue sacrifications and may enable the use of in vitro engineered vascularized flaps in many surgical applications. PMID:26779840

  16. Calcium efflux activity of plasma membrane Ca2+ ATPase-4 (PMCA4) mediates cell cycle progression in vascular smooth muscle cells.

    PubMed

    Afroze, Talat; Yang, Ge; Khoshbin, Amir; Tanwir, Mansoor; Tabish, Taha; Momen, Abdul; Husain, Mansoor

    2014-03-01

    We explored the role played by plasma membrane calcium ATPase-4 (PMCA4) and its alternative splice variants in the cell cycle of vascular smooth muscle cells (VSMC). A novel variant (PMCA4e) was discovered. Quantitative real-time-PCR-quantified PMCA4 splice variant proportions differed in specific organs. The PMCA4a:4b ratio in uninjured carotid arteries (∼1:1) was significantly reduced by wire denudation injury (to ∼1:3) by modulation of alternative splicing, as confirmed by novel antibodies against PMCA4a/e and PMCA4b. Laser capture microdissection localized this shift to the media and adventitia. Primary carotid VSMC from PMCA4 knock-out (P4KO) mice showed impaired [(3)H]thymidine incorporation and G1 phase arrest as compared with wild type (P4WT). Electroporation of expression constructs encoding PMCA4a, PMCA4b, and a PMCA4b mutant lacking PDZ binding rescued this phenotype of P4KO cells, whereas a mutant with only 10% of normal Ca(2+) efflux activity could not. Microarray of early G1-synchronized VSMC showed 39-fold higher Rgs16 (NFAT (nuclear factor of activated T-cells) target; MAPK inhibitor) and 69-fold higher Decorin (G1 arrest marker) expression in P4KO versus P4WT. Validation by Western blot also revealed decreased levels of Cyclin D1 and NFATc3 in P4KO. Microarrays of P4KO VSMC rescued by PMCA4a or PMCA4b expression showed reversal of perturbed Rgs16, Decorin, and NFATc3 expression levels. However, PMCA4a rescue caused a 44-fold reduction in AP-2β, a known anti-proliferative transcription factor, whereas PMCA4b rescue resulted in a 50-fold reduction in p15 (Cyclin D1/Cdk4 inhibitor). We conclude that Ca(2+) efflux activity of PMCA4 underlies G1 progression in VSMC and that PMCA4a and PMCA4b differentially regulate specific downstream mediators. PMID:24448801

  17. Vascular Anomalies and Airway Concerns

    PubMed Central

    Clarke, Caroline; Lee, Edward I.; Edmonds, Joseph

    2014-01-01

    Vascular anomalies, both tumors and malformations, can occur anywhere in the body, including the airway, often without any external manifestations. However, vascular anomalies involving the airway deserve special consideration as proper recognition and management can be lifesaving. In this article, the authors discuss vascular anomalies as they pertains to the airway, focusing on proper diagnosis, diagnostic modalities, and therapeutic options. PMID:25045336

  18. Distribution of cyclic AMP phosphodiesterase in microdissected periportal and perivenous rat liver tissue with different dietary states.

    PubMed

    Runge, D; Jungermann, K

    1991-01-01

    Cyclic AMP phosphodiesterase was measured in liver homogenates and microdissected periportal and perivenous liver tissue from rats in different dietary states under different conditions of substrate saturation and effector stimulation. A radiochemical microtest, more sensitive by 2-3 orders of magnitude than the usual assay, was established for the determination of the activity in liver samples corresponding to 200-800 ng dry weight. At saturating cyclic AMP concentrations (46 microM) phosphodiesterase was homogeneously distributed within the liver acinus of fed rats. Starvation for 48 h led to a decrease in the overall activity and to a heterogenous distribution with slightly higher activities in the perivenous zone. At physiological cyclic AMP concentrations (1.8 microM) phosphodiesterase showed a flat zonal gradient in livers of fed rats with higher levels in the periportal zone; after 48 h starvation it was homogeneously distributed. In the presence of cyclic GMP (2 microM) the basal activity at physiological substrate concentrations was stimulated to a greater extent in the perivenous zone. This led to a homogeneous activity distribution in the fed state and to a heterogenous pattern with a slight perivenous maximum in the fasted state. Thus there was no or only a small zonal heterogeneity of signal transmitting enzymes such as cyclic AMP phosphodiesterase and glucagon-stimulated adenylate cyclase (Zierz and Jungermann 1984). This similar signal transducing capacity in the periportal and the perivenous area will contribute to maintain the zonation of signal input due to the hormone concentration gradients across the liver acinus.

  19. Lasers and laser-like devices: part two.

    PubMed

    Sebaratnam, Deshan F; Lim, Adrian C; Lowe, Patricia M; Goodman, Greg J; Bekhor, Philip; Richards, Shawn

    2014-02-01

    Part two of this review series evaluates the use of lasers and laser-like devices in dermatology based on published evidence and the collective experience of the senior authors. Dermatologists can laser-treat a wide range of dermatoses, including vascular, pigmentary, textural, benign proliferative and premalignant conditions. Some of these conditions include vascular malformation, haemangioma, facial telangiectases, café-au-lait macules, naevi of Ota, lentigines, acne scarring, rhytides, rhinophyma and miscellaneous skin lesions. Photodynamic therapy with lasers and intense pulsed light is addressed, with particular reference to actinic keratosis and actinic cheilitis. A treatment algorithm for acne scarring based on scar morphology and severity is comprehensively outlined. Following from part one, the various devices are matched to the corresponding dermatological conditions with representative pictorial case vignettes illustrating likely clinical outcomes as well as limitations and potential complications of the various laser and light therapies. PMID:24433372

  20. Lasers and laser-like devices: part two.

    PubMed

    Sebaratnam, Deshan F; Lim, Adrian C; Lowe, Patricia M; Goodman, Greg J; Bekhor, Philip; Richards, Shawn

    2014-02-01

    Part two of this review series evaluates the use of lasers and laser-like devices in dermatology based on published evidence and the collective experience of the senior authors. Dermatologists can laser-treat a wide range of dermatoses, including vascular, pigmentary, textural, benign proliferative and premalignant conditions. Some of these conditions include vascular malformation, haemangioma, facial telangiectases, café-au-lait macules, naevi of Ota, lentigines, acne scarring, rhytides, rhinophyma and miscellaneous skin lesions. Photodynamic therapy with lasers and intense pulsed light is addressed, with particular reference to actinic keratosis and actinic cheilitis. A treatment algorithm for acne scarring based on scar morphology and severity is comprehensively outlined. Following from part one, the various devices are matched to the corresponding dermatological conditions with representative pictorial case vignettes illustrating likely clinical outcomes as well as limitations and potential complications of the various laser and light therapies.

  1. The Vascular Depression Hypothesis: Mechanisms Linking Vascular Disease with Depression

    PubMed Central

    Taylor, Warren D.; Aizenstein, Howard J.; Alexopoulos, George S.

    2013-01-01

    The ‘Vascular Depression’ hypothesis posits that cerebrovascular disease may predispose, precipitate, or perpetuate some geriatric depressive syndromes. This hypothesis stimulated much research that has improved our understanding of the complex relationships between late-life depression (LLD), vascular risk factors, and cognition. Succinctly, there are well-established relationships between late-life depression, vascular risk factors, and cerebral hyperintensities, the radiological hallmark of vascular depression. Cognitive dysfunction is common in late-life depression, particularly executive dysfunction, a finding predictive of poor antidepressant response. Over time, progression of hyperintensities and cognitive deficits predicts a poor course of depression and may reflect underlying worsening of vascular disease. This work laid the foundation for examining the mechanisms by which vascular disease influences brain circuits and influences the development and course of depression. We review data testing the vascular depression hypothesis with a focus on identifying potential underlying vascular mechanisms. We propose a disconnection hypothesis, wherein focal vascular damage and white matter lesion location is a crucial factor influencing neural connectivity that contributes to clinical symptomatology. We also propose inflammatory and hypoperfusion hypotheses, concepts that link underlying vascular processes with adverse effects on brain function that influence the development of depression. Testing such hypotheses will not only inform the relationship between vascular disease and depression but also provide guidance on the potential repurposing of pharmacological agents that may improve late-life depression outcomes. PMID:23439482

  2. Extracranial vascular malformations (hemangiomas and vascular malformations) in children and adolescents – diagnosis, clinic, and therapy

    PubMed Central

    Eivazi, Behfar; Werner, Jochen A.

    2014-01-01

    The field of extracranial vascular anomalies is considered as special focus of pediatric otolaryngology and it has shown a rapid development during the last years. The reason for this interest is finally also due to the global acceptance of the classification introduced by the ISSVA (International Society for the Study of Vascular Anomalies). Hemangiomas are the most frequently observed vascular tumors. Today the systemic propranolol therapy is mostly used for therapy of hemangiomas requiring treatment. Increasingly, the topical application of beta blocker is discussed while the benefit in the head and neck seems to be limited. Vascular malformations are classified according to the morphology of the affected part of the vascular system in arterial, venous, arterio-venous, lymphatic, capillary, and combined vascular malformations. Conventional surgery, sclerosing therapy, and laser treatment are invasive options for the treatment of lymphatic malformations. The options for the treatment of venous malformations could be significantly improved during the last years. In this context, the use of Nd:YAG laser, the conservative treatment of the localized disseminated intravascular coagulation with low-molecular weight heparin, the re-discovery of bleomycin as effective sclerosing agent, and the improvement of alcohol-based embolization agents must be mentioned. Today the treatment with dye laser is the preferred therapy for capillary malformations and it is superior to other therapeutic options as for example photodynamic therapy. Arterio-venous malformations as representatives for high-flow lesions are the high-risk lesions. Frequently they are compared to malignant head and neck tumors, in particular when a curative treatment can no longer be assured because of diffuse or multifocal extent and when the disease shows a progressive course. The combined treatment of embolization and surgical resection and if necessary consecutive defect reconstruction have turned out to be

  3. Sensitive method for determination of picogram amounts of epinephrine and other catecholamines in microdissected samples of rat brain using liquid chromatography with electrochemical detection.

    PubMed

    Opacka-Juffry, J; Tacconelli, F; Coen, C W

    1988-12-01

    Liquid chromatography with high-sensitivity electrochemical detection has been employed to measure picogram amounts of epinephrine and other catecholamines in microdissected samples of the rat hypothalamus. Tissue catecholamines are purified by solvent extraction; this provides better selectivity and recovery than methods involving alumina. The solvent extraction technique has been modified in order to eliminate its major disadvantage, the presence of electroactive substances separating with catecholamines. Detection limits of below 1 pg allow for analysis of catecholamines including epinephrine in very small brain samples such as micropunches.

  4. Vascular cognitive impairment and dementia.

    PubMed

    Gorelick, Philip B; Counts, Scott E; Nyenhuis, David

    2016-05-01

    Vascular contributions to cognitive impairment are receiving heightened attention as potentially modifiable factors for dementias of later life. These factors have now been linked not only to vascular cognitive disorders but also Alzheimer's disease. In this chapter we review 3 related topics that address vascular contributions to cognitive impairment: 1. vascular pathogenesis and mechanisms; 2. neuropsychological and neuroimaging phenotypic manifestations of cerebrovascular disease; and 3. prospects for prevention of cognitive impairment of later life based on cardiovascular and stroke risk modification. This article is part of a Special Issue entitled: Vascular Contributions to Cognitive Impairment and Dementia edited by M. Paul Murphy, Roderick A. Corriveau and Donna M. Wilcock. PMID:26704177

  5. Neurobiology of Vascular Dementia

    PubMed Central

    Enciu, Ana-Maria; Constantinescu, Stefan N.; Popescu, Laurenţiu M.; Mureşanu, Dafin F.; Popescu, Bogdan O.

    2011-01-01

    Vascular dementia is, in its current conceptual form, a distinct type of dementia with a spectrum of specific clinical and pathophysiological features. However, in a very large majority of cases, these alterations occur in an already aged brain, characterized by a milieu of cellular and molecular events common for different neurodegenerative diseases. The cell signaling defects and molecular dyshomeostasis might lead to neuronal malfunction prior to the death of neurons and the alteration of neuronal networks. In the present paper, we explore some of the molecular mechanisms underlying brain malfunction triggered by cerebrovascular disease and risk factors. We suggest that, in the age of genetic investigation and molecular diagnosis, the concept of vascular dementia needs a new approach. PMID:21876809

  6. Plant Vascular Biology 2010

    SciTech Connect

    Ding, Biao

    2014-11-17

    This grant supported the Second International Conference on Plant Vascular Biology (PVB 2010) held July 24-28, 2010 on the campus of Ohio State University, Columbus, Ohio. Biao Ding (Ohio State University; OSU) and David Hannapel (Iowa State University; ISU) served as co-chairs of this conference. Biao Ding served as the local organizer. PVB is defined broadly here to include studies on the biogenesis, structure and function of transport systems in plants, under conditions of normal plant growth and development as well as of plant interactions with pathogens. The transport systems cover broadly the xylem, phloem, plasmodesmata and vascular cell membranes. The PVB concept has emerged in recent years to emphasize the integrative nature of the transport systems and approaches to investigate them.

  7. [Vascular variability syndromes].

    PubMed

    Otsuka, Kuniaki; Okajima, Kiyotaka; Yamanaka, Takashi; Cornelissen, Germaine

    2014-08-01

    Analytical global and local methods applied to human blood pressure (BP) records of around-the-clock measurements. The chronobiological interpretation of ambulatory BP monitoring records in the light of time-specified reference values derived from healthy peers matched by sex and age identify vascular variability disorders (VVDs) for an assessment of cardio-, cerebro-, and renovascular disease risk. VVD includes circadian BP over-swinging (CHAT, short for circadian hyper-amplitude tension), deficient heart rate variability, MESOR (midline-estimating statistic of rhythm) hypertension, excessively elevated pulse pressure over 60 mmHg, BP ecphasia (an odd timing of the circadian rhythms in BP but not in that of heart rate) and frequency alteration. The term MESOR-hypertension indicates only one of several VVDs that can combine to for sets of 2, 3 and n-component vascular variability syndromes. PMID:25167758

  8. [Vascular endothelial Barrier Function].

    PubMed

    Ivanov, A N; Puchinyan, D M; Norkin, I A

    2015-01-01

    Endothelium is an important regulator of selective permeability of the vascular wall for different molecules and cells. This review summarizes current data on endothelial barrier function. Endothelial glycocalyx structure, its function and role in the molecular transport and leukocytes migration across the endothelial barrier are discussed. The mechanisms of transcellular transport of macromolecules and cell migration through endothelial cells are reviewed. Special section of this article addresses the structure and function of tight and adherens endothelial junction, as well as their importance for the regulation of paracellular transport across the endothelial barrier. Particular attention is paid to the signaling mechanism of endothelial barrier function regulation and the factors that influence on the vascular permeability.

  9. Vascular Cambium Development

    PubMed Central

    Nieminen, Kaisa; Blomster, Tiina; Helariutta, Ykä; Mähönen, Ari Pekka

    2015-01-01

    Secondary phloem and xylem tissues are produced through the activity of vascular cambium, the cylindrical secondary meristem which arises among the primary plant tissues. Most dicotyledonous species undergo secondary development, among them Arabidopsis. Despite its small size and herbaceous nature, Arabidopsis displays prominent secondary growth in several organs, including the root, hypocotyl and shoot. Together with the vast genetic resources and molecular research methods available for it, this has made Arabidopsis a versatile and accessible model organism for studying cambial development and wood formation. In this review, we discuss and compare the development and function of the vascular cambium in the Arabidopsis root, hypocotyl, and shoot. We describe the current understanding of the molecular regulation of vascular cambium and compare it to the function of primary meristems. We conclude with a look at the future prospects of cambium research, including opportunities provided by phenotyping and modelling approaches, complemented by studies of natural variation and comparative genetic studies in perennial and woody plant species. PMID:26078728

  10. Congenital Vascular Anomalies.

    PubMed

    Gravereaux, Edwin C.; Nguyen, Louis L.; Cunningham, Leslie D.

    2004-04-01

    Congenital vascular anomalies are rare. The cardiovascular specialist should nevertheless be aware of the more common types of vascular anomalies and understand the implications for patient treatment and the likelihood of associated morbidity. The presentation of congenital arteriovenous malformations can range from asymptomatic or cosmetic lesions, to those causing ischemia, ulceration, hemorrhage, or high-output congestive heart failure. Treatment of large, symptomatic arteriovenous malformations often requires catheter-directed embolization prior to the attempt at complete surgical excision. Later recurrence, due to collateral recruitment, is frequent. Graded compression stockings and leg elevation are the mainstays of treatment for the predominantly venous congenital vascular anomalies. Most congenital central venous disorders are clinically silent. An exception is the retrocaval ureter. Retroaortic left renal vein, circumaortic venous ring, and absent, left-sided or duplicated inferior vena cava are relevant when aortic or inferior vena cava procedures are planned. The treatment of the venous disorders is directed at prevention or management of symptoms. Persistent sciatic artery, popliteal entrapment syndrome, and aberrant right subclavian artery origin are congenital anomalies that are typically symptomatic at presentation. Because they mimic more common diseases, diagnosis is frequently delayed. Delay can result in significant morbidity for the patient. Failure to make the diagnosis of persistent sciatic artery and popliteal entrapment can result in critical limb ischemia and subsequent amputation. Unrecognized aberrant right subclavian artery origin associated with aneurysmal degeneration can rupture and result in death. The treatment options for large-vessel arterial anomalies are surgical, sometimes in combination with endovascular techniques.

  11. [The use of lasers in dermatology].

    PubMed

    Lecocq, C; Pirard, D; del Marmol, V; Berlingin, E

    2013-01-01

    Albert Einstein is undoubtedly the father of lasers. But it is not until 1964 that the first dermatological lasers were introduced. The Nd-YAG laser, the CO2 laser were developed by Kumar Patel. In a 40 year period lasers not only were diversified but have also become safer and miniaturized. This article hopes to strengthen general practionners' and specialist's knowledge of the different categories of available lasers. The most frequently used ones are ablative lasers (CO2-Erbium), vascular lasers (Nd-YAG, KTP, pulsed dye laser) and the pigment lasers (Q-Switched Nd-YAG, Alexandrite). A description of these lasers and their indications in dermatology will be discussed. PMID:23534310

  12. Retinal vascular changes are a marker for cerebral vascular diseases

    PubMed Central

    Moss, Heather E.

    2016-01-01

    The retinal circulation is a potential marker of cerebral vascular disease because it shares origin and drainage with the intracranial circulation and because it can be directly visualized using ophthalmoscopy. Cross sectional and cohort studies have demonstrated associations between chronic retinal and cerebral vascular disease, acute retinal and cerebral vascular disease and chronic retinal vascular disease and acute cerebral vascular disease. In particular, certain qualitative features of retinopathy, retinal artery occlusion and increased retinal vein caliber are associated with concurrent and future cerebrovascular events. These associations persist after accounting for confounding variables known to be disease-causing in both circulations, which supports the potential use of retinal vasculature findings to stratify individuals with regards to cerebral vascular disease risk. PMID:26008809

  13. Laser Therapy For Arteriosclerosis: Current Review

    NASA Astrophysics Data System (ADS)

    Dries, David J.; Pollock, Marc E.; Eugene, John

    1987-03-01

    Shortly after the ruby laser was introduced, in 1959, a study for the use of this ruby laser for the in-vitro dissolution of arteriosclerotic plaque was performed.' With subsequent advances in laser technology and with refined delivery techniques, laser applications to the treatment of arteriosclerosis in the coronary arteries and peripheral vascular system is a reality. This report reviews the disease process, arteriosclerosis, and the ef-forts towards laser treatment of this disease. We conclude with a review of the technical barriers to the routine application of laser energy in arteriosclerotic cardiovascular disease and the progress being made to overcome these obstacles.

  14. Vascular effects of flavonoids.

    PubMed

    Almeida Rezende, Bruno; Pereira, Aline Carvalho; Cortes, Steyner F; Lemos, Virginia Soares

    2016-01-01

    Flavonoids are natural plant-derived polyphenolic compounds with various biological properties particularly in the cardiovascular system, including antiatherogenic, antioxidant, vasodilation, antihypertensive, and antiplatelet activities. These biological properties have been evaluated in several experimental and clinical studies. In addition, extensive reviews have discussed the antiatherogenic effect of these polyphenols. However, limited studies have investigated the potential therapeutic vascular effects of these compounds. This review brings together some recent studies, to establish the different signaling pathways involved in the molecular mechanisms that underlie the vasodilation induced by flavonoids.

  15. Medical management of vascular anomalies.

    PubMed

    Trenor, Cameron C

    2016-03-01

    We have entered an exciting era in the care of patients with vascular anomalies. These disorders require multidisciplinary care and coordination and dedicated centers have emerged to address this need. Vascular tumors have been treated with medical therapies for many years, while malformations have been historically treated with endovascular and operative procedures. The recent serendipitous discoveries of propranolol and sirolimus for vascular anomalies have revolutionized this field. In particular, sirolimus responses are challenging the dogma that vascular malformations are not biologically active. While initially explored for lymphatic anomalies, sirolimus is now being used broadly throughout the spectrum of vascular anomalies. Whether medical therapies are reserved for refractory patients or used first line is currently dependent on the experience and availability of alternative therapies at each institution. On the horizon, we anticipate new drugs targeting genes and pathways involved in vascular anomalies to be developed. Also, combinations of medications and protocols combining medical and procedural approaches are in development for refractory patients. PMID:27607327

  16. The pathobiology of vascular dementia

    PubMed Central

    Iadecola, Costantino

    2013-01-01

    Vascular cognitive impairment defines alterations in cognition, ranging from subtle deficits to full-blown dementia, attributable to cerebrovascular causes. Often coexisting with Alzheimer’s disease, mixed vascular and neurodegenerative dementia has emerged as the leading cause of age-related cognitive impairment. Central to the disease mechanism is the crucial role that cerebral blood vessels play in brain health, not only for the delivery of oxygen and nutrients, but also for the trophic signaling that links inextricably the well being of neurons and glia to that of cerebrovascular cells. This review will examine how vascular damage disrupts these vital homeostatic interactions, focusing on the hemispheric white matter, a region at heightened risk for vascular damage, and on the interplay between vascular factors and Alzheimer’s disease. Finally, preventative and therapeutic prospects will be examined, highlighting the importance of midlife vascular risk factor control in the prevention of late-life dementia. PMID:24267647

  17. Vascular pattern formation in plants.

    PubMed

    Scarpella, Enrico; Helariutta, Ykä

    2010-01-01

    Reticulate tissue systems exist in most multicellular organisms, and the principles underlying the formation of cellular networks have fascinated philosophers, mathematicians, and biologists for centuries. In particular, the beautiful and varied arrangements of vascular tissues in plants have intrigued mankind since antiquity, yet the organizing signals have remained elusive. Plant vascular tissues form systems of interconnected cell files throughout the plant body. Vascular cells are aligned with one another along continuous lines, and vascular tissues differentiate at reproducible positions within organ environments. However, neither the precise path of vascular differentiation nor the exact geometry of vascular networks is fixed or immutable. Several recent advances converge to reconcile the seemingly conflicting predictability and plasticity of vascular tissue patterns. A control mechanism in which an apical-basal flow of signal establishes a basic coordinate system for body axis formation and vascular strand differentiation, and in which a superimposed level of radial organizing cues elaborates cell patterns, would generate a reproducible tissue configuration in the context of an underlying robust, self-organizing structure, and account for the simultaneous regularity and flexibility of vascular tissue patterns.

  18. Vascular trauma in civilian practice.

    PubMed Central

    Golledge, J.; Scriven, M. W.; Fligelstone, L. J.; Lane, I. F.

    1995-01-01

    Vascular trauma is associated with major morbidity and mortality, but little is known about its incidence or nature in Britain. A retrospective study of 36 patients requiring operative intervention for vascular trauma under one vascular surgeon over a 6-year period was undertaken. Twenty-four patients suffered iatrogenic trauma (median age 61 years); including cardiological intervention (19), radiological intervention (2), varicose vein surgery (1), umbilical vein catherisation (1) and isolated hyperthermic limb perfusion (1). There were 23 arterial and three venous injuries. Twelve patients had accidental trauma (median age 23 years). Three of the ten patients with blunt trauma were referred for vascular assessment before orthopaedic intervention, two after an on-table angiogram and five only after an initial orthopaedic procedure (range of delay 6 h to 10 days). Injuries were arterial in nine, venous in two and combined in one. Angiography was obtained in six patients, and in two patients with multiple upper limb fractures identified the site of injury when clinical localisation was difficult. A variety of vascular techniques were used to treat the injuries. Two patients died postoperatively and one underwent major limb amputation. Thirty-two (89%) remain free of vascular sequelae after a median follow-up of 48 months (range 3-72 months). Vascular trauma is uncommon in the United Kingdom. To repair the injuries a limited repertoire of vascular surgery techniques is needed. Therefore, vascular surgical assessment should be sought at an early stage to prevent major limb loss. PMID:8540659

  19. Vascularization of engineered teeth.

    PubMed

    Nait Lechguer, A; Kuchler-Bopp, S; Hu, B; Haïkel, Y; Lesot, H

    2008-12-01

    The implantation of cultured dental cell-cell re-associations allows for the reproduction of fully formed teeth, crown morphogenesis, epithelial histogenesis, mineralized dentin and enamel deposition, and root-periodontium development. Since vascularization is critical for organogenesis and tissue engineering, this work aimed to study: (a) blood vessel formation during tooth development, (b) the fate of blood vessels in cultured teeth and re-associations, and (c) vascularization after in vivo implantation. Ex vivo, blood vessels developed in the dental mesenchyme from the cap to bell stages and in the enamel organ, shortly before ameloblast differentiation. In cultured teeth and re-associations, blood-vessel-like structures remained in the peridental mesenchyme, but never developed into dental tissues. After implantation, both teeth and re-associations became revascularized, although later in the case of the re-associations. In implanted re-associations, newly formed blood vessels originated from the host, allowing for their survival, and affording conditions organ growth, mineralization, and enamel secretion.

  20. Vascular Distribution of Nanomaterials

    PubMed Central

    Stapleton, Phoebe A.; Nurkiewicz, Timothy R.

    2014-01-01

    Once considered primarily occupational, novel nanotechnology innovation and application has led to widespread domestic use and intentional biomedical exposures. With these exciting advances, the breadth and depth of toxicological considerations must also be expanded. The vascular system interacts with every tissue in the body, striving to homeostasis. Engineered nanomaterials (ENM) have been reported to distribute in many different organs and tissues. However, these observations have tended to use approaches requiring tissue homogenization and/or gross organ analyses. These techniques, while effective in establishing presence, preclude an exact determination of where ENM are deposited within a tissue. It is necessary to identify this exact distribution and deposition of ENM throughout the cardiovascular system, with respect to vascular hemodynamics and in vivo/ in vitro ENM modifications taken into account if nanotechnology is to achieve its full potential. Distinct levels of the vasculature will first be described as individual compartments. Then the vasculature will be considered as a whole. These unique compartments and biophysical conditions will be discussed in terms of their propensity to favor ENM deposition. Understanding levels of the vasculature will also be discussed. Ultimately, future studies must verify the mechanisms speculated on and presented herein. PMID:24777845

  1. Vascular graft infections.

    PubMed

    Hasse, Barbara; Husmann, Lars; Zinkernagel, Annelies; Weber, Rainer; Lachat, Mario; Mayer, Dieter

    2013-01-01

    Vascular procedures are rarely complicated by infection, but if prosthetic vascular graft infection (PVGI) occurs, morbidity and mortality are high. Several patient-related, surgery-related and postoperative risk factors are reported, but they are not well validated. PVGI is due to bacterial colonisation of the wound and the underlying prosthetic graft, generally as a result of direct contamination during the operative procedure, mainly from the patient's skin or adjacent bowel. There is no consensus on diagnostic criteria or on the best management of PVGI. On the basis of reported clinical studies and our own experience, we advocate a surgical approach combining repeated radical local debridement, with graft preservation whenever possible or partial excision of the infected graft, depending on its condition, plus simultaneous negative-pressure wound therapy (NPWT). In addition, antimicrobial therapy is recommended, but there is no consensus on which classes of agent are adequate for the treatment of PVGI and whether certain infections may be treated by means of NPWT alone. Since staphylococci and Gram-negative rods are likely to be isolated, empirical treatment might include a penicillinase-resistant beta-lactam or a glycopeptide, plus an aminoglycoside, the latter for Gram-negative coverage and synergistic treatment of Gram-positive cocci. Additionally, empirical treatment might include rifampicin since it penetrates well into biofilms.

  2. Use of high-power lasers in oral surgery

    NASA Astrophysics Data System (ADS)

    Gaspar, Lajos

    1993-12-01

    The treatment of 2989 patients with different type of lasers was described. The argon laser beam was used in 57 cases (portwine stains, telangiectasias, angiofibromas and other vascular lesions) and 84 operations were performed by Nd:YAG laser (leukoplakia, hemangioma etc.) furthermore 53 operations by combined laser beam. 2795 operations by carbon dioxide laser were performed in precancerous states and other white lesions, benign tumors and tumor-like states, malignant tumors and other lesions.

  3. Non-invasive optical modulation of local vascular permeability

    NASA Astrophysics Data System (ADS)

    Choi, Myunghwan; Choi, Chulhee

    2011-03-01

    For a systemically administered drug to act, it first needs to cross the vascular wall. This step represents a bottleneck for drug development, especially in the brain or retina, where tight junctions between endothelial cells form physiological barriers. Here, we demonstrate that femtosecond pulsed laser irradiation focused on the blood vessel wall induces transient permeabilization of plasma. Nonlinear absorption of the pulsed laser enabled the noninvasive modulation of vascular permeability with high spatial selectivity in three dimensions. By combining this method with systemic injection, we could locally deliver molecular probes in various tissues, such as brain cortex, meninges, ear, striated muscle, and bone. We suggest this method as a novel delivery tool for molecular probes or drugs.

  4. 219 vascular fellows' perception of the future of vascular surgery.

    PubMed

    Hingorani, Anil P; Ascher, Enrico; Marks, Natalie; Shiferson, Alexander; Puggioni, Alessandra; Tran, Victor; Patel, Nirav; Jacob, Theresa

    2009-01-01

    In an attempt to identify the fellows' concerns about the future of the field of vascular surgery, we conducted a survey consisting of 22 questions at an annual national meeting in March from 2004 to 2007. In order to obtain accurate data, all surveys were kept anonymous. The fellows were asked (1) what type of practice they anticipated they would be in, (2) what the new training paradigm for fellows should be, (3) to assess their expectation of the needed manpower with respect to the demand for vascular surgeons, (4) what were major threats to the future of vascular surgery, (5) whether they had heard of and were in favor of the American Board of Vascular Surgery (ABVS), (6) who should be able to obtain vascular privileges, and (7) about their interest in an association for vascular surgical trainees. Of 273 attendees, 219 (80%) completed the survey. Males made up 87% of those surveyed, and 60% were between the ages of 31 and 35 years. Second-year fellows made up 82% of those surveyed. Those expecting to join a private, academic, or mixed practice made up 35%, 28%, and 20% of the respondents, respectively, with 71% anticipating entering a 100% vascular practice. Forty percent felt that 5 years of general surgery with 2 years of vascular surgery should be the training paradigm, while 45% suggested 3 and 3 years, respectively. A majority, 79%, felt that future demand would exceed the available manpower, while 17% suggested that manpower would meet demand. The major challenges to the future of vascular surgery were felt to be competition from cardiology (82%) or radiology (30%) and lack of an independent board (29%). Seventeen percent were not aware of the ABVS, and only 2% were against it; 71% suggested that vascular privileges be restricted to board-certified vascular surgeons. Seventy-six percent were interested in forming an association for vascular trainees to address the issues of the future job market (67%), endovascular training during fellowship (56

  5. Clinical Characteristics and Treatment Options of Infantile Vascular Anomalies

    PubMed Central

    Yang, Bin; Li, Li; Zhang, Li-xin; Sun, Yu-juan; Ma, Lin

    2015-01-01

    Abstract To analyze the clinical characteristics and treatment outcomes of vascular anomalies, and determine which therapy is safe and effective. The data of vascular anomalies pediatric patients who arrived at Beijing children's Hospital from January 2001 to December 2014 were analyzed retrospectively, including the influence of gender, age, clinical manifestation, diagnosis, treatment options, and outcomes. As to infantile hemangiomas, the outcomes of different treatments and their adverse reactions were compared. As to spider angioma and cutaneous capillary malformation, the treatment effect of 595 nm pulsed dye laser (PDL) is analyzed. A total number of 6459 cases of vascular anomalies were reclassified according to the 2014 ISSVA classification system. Among them, the gender ratio is 1:1.69, head-and-neck involved is 53.3%, the onset age within the first month is 72.4%, the age of initial encounter that younger than 6 months is 60.1%. The most common anomalies were infantile hemangiomas (42.6%), congenital hemangiomas (14.1%), and capillary malformations (29.9%). In treating infantile hemangiomas, laser shows the lowest adverse reactions rate significantly. Propranolol shows a higher improvement rate than laser, glucocorticoids, glucocorticoids plus laser, and shows no significant difference with propranolol plus laser both in improvement rate and adverse reactions rate. The total improvement rate of 595 nm PDL is 89.8% in treating spider angioma and 46.7% in treating cutaneous capillary malformation. The improvement rate and excellent rate of laser in treating cutaneous capillary malformation are growing synchronously by increasing the treatment times, and shows no significant difference among different parts of lesion that located in a body. Vascular anomalies possess a female predominance, and are mostly occurred in faces. Definite diagnosis is very important before treatment. In treating infantile hemangioma, propranolol is recommended as the first

  6. Argon laser treatment of strawberry hemangioma in infancy.

    PubMed

    Achauer, B M; Vander Kam, V M

    1985-11-01

    Argon laser therapy is effective for removing port-wine stains and for reducing cutaneous vascular and pigmented lesions. Strawberry hemangiomas, being much thicker lesions than port-wine stains, were considered not appropriate for argon laser treatment. Using argon laser therapy in 13 cases of strawberry hemangioma, we achieved poor to dramatic results. PMID:4082569

  7. Holographic laser Doppler ophthalmoscopy.

    PubMed

    Simonutti, M; Paques, M; Sahel, J A; Gross, M; Samson, B; Magnain, C; Atlan, M

    2010-06-15

    We report laser Doppler ophthalmoscopic fundus imaging in the rat eye with near-IR heterodyne holography. Sequential sampling of the beat of the reflected radiation against a frequency-shifted optical local oscillator is made onto an array detector. Wide-field maps of fluctuation spectra in the 10 Hz to 25 kHz band exhibit angiographic contrasts in the retinal vascular tree without requirement of an exogenous marker.

  8. Action spectrum of vascular specific injury using pulsed irradiation

    SciTech Connect

    Tan, O.T.; Murray, S.; Kurban, A.K.

    1989-06-01

    It has been clearly demonstrated that cutaneous blood vessels will be selectively damaged by a laser whose wavelength matches one of the three absorption spectral peaks of the chromophore, oxyhemoglobin, for example, 577 nm. A restriction in the application of this wavelength for the treatment of benign cutaneous vascular tumors, such as portwine stains, has been the penetration depth of 577 nm irradiation of approximately 0.5 mm from the dermal epidermal junction (DEJ). This study was undertaken to establish whether it was possible to increase the penetration depth from 0.5 mm by changing the wavelength to beyond 577 nm in albino pig skin. Results from this study confirm that penetration depth increases from 0.5 to 1.2 mm by changing the wavelength from 577 to 585 nm at 4 J/cm2, while maintaining the same degree of vascular selectivity as that previously described after 577 nm irradiation. This occurred in spite of a mismatch in the wavelength between 585 nm and the oxyhemoglobin absorption peak of 577 nm. Unlike 585 nm irradiation and in contrast with theoretical predictions, 590 nm laser light did not penetrate as deeply as 585 nm. Not only was there a reduction in the penetration depth of the laser beam from 1.2 mm at 585 nm to 0.8 mm at 590 nm, at 4 J/cm2, but there was also a decrease in vascular selectivity in albino pig skin exposed to 590 nm irradiation.

  9. Vascular surgery: the European perspective.

    PubMed

    Harris, P

    1999-09-01

    Isaac Newton, among others, observed that 'we see so far because we are standing upon the shoulders of giants'. In vascular surgery most of the giants have been European, and this is a heritage which we as Europeans can take pride in and build upon if we chose to do so. As in other areas of life, commitment is essential in order to influence the future. For vascular surgeons in Europe this means active participation in the European scientific societies for vascular surgery and in the UEMS. The main value of the EBSQ.VASC assessments to date has been to expose the uneven standards of training in vascular surgery within the European Union. Only if action follows to address these inequalities will the tactics of the European Board of Vascular Surgery be vindicated.

  10. Caffeine's Vascular Mechanisms of Action

    PubMed Central

    Echeverri, Darío; Montes, Félix R.; Cabrera, Mariana; Galán, Angélica; Prieto, Angélica

    2010-01-01

    Caffeine is the most widely consumed stimulating substance in the world. It is found in coffee, tea, soft drinks, chocolate, and many medications. Caffeine is a xanthine with various effects and mechanisms of action in vascular tissue. In endothelial cells, it increases intracellular calcium stimulating the production of nitric oxide through the expression of the endothelial nitric oxide synthase enzyme. Nitric oxide is diffused to the vascular smooth muscle cell to produce vasodilation. In vascular smooth muscle cells its effect is predominantly a competitive inhibition of phosphodiesterase, producing an accumulation of cAMP and vasodilation. In addition, it blocks the adenosine receptors present in the vascular tissue to produce vasoconstriction. In this paper the main mechanisms of action of caffeine on the vascular tissue are described, in which it is shown that caffeine has some cardiovascular properties and effects which could be considered beneficial. PMID:21188209

  11. Study of the Structure, Oxygen-Transporting Functions, and Ionic Composition of Erythrocytes at Vascular Diseases

    PubMed Central

    Revin, Viktor V.; Gromova, Natalia V.; Revina, Elvira S.; Mel'nikova, Natalya A.; Balykova, Larisa A.; Solomadin, Ilia N.; Tychkov, Alexander Yu.; Revina, Nadezhda V.; Gromova, Oksana Yu.; Anashkina, Irina V.; Yakushkin, Viktor A.

    2015-01-01

    The present paper explores the role of erythrocytes in the pathogenesis of vascular diseases. The state of erythrocytes, their ionic composition and structure, and properties of erythrocytes hemoglobin were studied by using laser interference microscopy, Raman scattering spectroscopy, and capillary electrophoresis. In patients suffering from vascular disorders we identified statistically significant changes in the shape of erythrocytes, their ionic composition, and redistribution of hemoglobin throughout cells. PMID:26601112

  12. Connections Matter: Channeled Hydrogels to Improve Vascularization

    PubMed Central

    Muehleder, Severin; Ovsianikov, Aleksandr; Zipperle, Johannes; Redl, Heinz; Holnthoner, Wolfgang

    2014-01-01

    The use of cell-laden hydrogels to engineer soft tissue has been emerging within the past years. Despite, several newly developed and sophisticated techniques to encapsulate different cell types the importance of vascularization of the engineered constructs is often underestimated. As a result, cell death within a construct leads to impaired function and inclusion of the implant. Here, we discuss the fabrication of hollow channels within hydrogels as a promising strategy to facilitate vascularization. Furthermore, we present an overview on the feasible use of removable spacers, 3D laser-, and planar processing strategies to create channels within hydrogels. The implementation of these structures promotes control over cell distribution and increases oxygen transport and nutrient supply in vitro. However, many studies lack the use of endothelial cells in their approaches leaving out an important factor to enhance vessel ingrowth and anastomosis formation upon implantation. In addition, the adequate endothelial cell type needs to be considered to make these approaches bridge the gap to in vivo applications. PMID:25453032

  13. Geriatric syndromes--vascular disorders?

    PubMed

    Strandberg, Timo E; Pitkälä, Kaisu H; Tilvis, Reijo S; O'Neill, Desmond; Erkinjuntti, Timo J

    2013-05-01

    The term geriatric syndrome is used to characterize multifactorial clinical conditions among older people which are not subsumed readily into disease entities, but which nevertheless predispose older people to disability and death. Commonly included are frailty, dementia, delirium, incontinence, falls, and dizziness. Geriatric syndromes are common among older people: in a recent survey, 50% of those aged more than 65 had one or more of these conditions. Better methods for prevention and treatment are needed, but current strategies have lacked a coherent conceptual and diagnostic framework. Prevention and interventions need to be targeted at earlier ages, with geriatrics expertise needed in the definition and operationalization of these complex entities. In this review we consolidate evidence that vascular disorders, including vascular ageing and vascular diseases, are key etiological factors of geriatric syndromes. Identifying this vascular dimension would offer opportunities for more efficient preventive strategies and mandates earlier intervention, especially for women, among whom vascular disease is often expressed more insidiously than among men. This would entail a sensitization of the health care system to the systematic detection of the syndromes, which are currently underdiagnosed. Further disentangling of the mechanisms of vascular ageing may offer therapies for vascular diseases and geriatric syndromes alike.

  14. Overgrowth syndromes with vascular anomalies.

    PubMed

    Blei, Francine

    2015-04-01

    Overgrowth syndromes with vascular anomalies encompass entities with a vascular anomaly as the predominant feature vs those syndromes with predominant somatic overgrowth and a vascular anomaly as a more minor component. The focus of this article is to categorize these syndromes phenotypically, including updated clinical criteria, radiologic features, evaluation, management issues, pathophysiology, and genetic information. A literature review was conducted in PubMed using key words "overgrowth syndromes and vascular anomalies" as well as specific literature reviews for each entity and supportive genetic information (e.g., somatic mosaicism). Additional searches in OMIM and Gene Reviews were conducted for each syndrome. Disease entities were categorized by predominant clinical features, known genetic information, and putative affected signaling pathway. Overgrowth syndromes with vascular anomalies are a heterogeneous group of disorders, often with variable clinical expression, due to germline or somatic mutations. Overgrowth can be focal (e.g., macrocephaly) or generalized, often asymmetrically (and/or mosaically) distributed. All germ layers may be affected, and the abnormalities may be progressive. Patients with overgrowth syndromes may be at an increased risk for malignancies. Practitioners should be attentive to patients having syndromes with overgrowth and vascular defects. These patients require proactive evaluation, referral to appropriate specialists, and in some cases, early monitoring for potential malignancies. Progress in identifying vascular anomaly-related overgrowth syndromes and their genetic etiology has been robust in the past decade and is contributing to genetically based prenatal diagnosis and new therapies targeting the putative causative genetic mutations. PMID:25937473

  15. [The future of vascular medicine].

    PubMed

    Kroeger, K; Luther, B

    2014-10-01

    In the future vascular medicine will still have a great impact on health of people. It should be noted that the aging of the population does not lead to a dramatic increase in patient numbers, but will be associated with a changing spectrum of co-morbidities. In addition, vascular medical research has to include the intensive care special features of vascular patients, the involvement of vascular medicine in a holistic concept of fast-track surgery, a geriatric-oriented intensive monitoring and early geriatric rehabilitation. For the future acceptance of vascular medicine as a separate subject area under delimitation of cardiology and radiology is important. On the other hand, the subject is so complex and will become more complex in future specialisations that mixing of surgery and angiology is desirable, with the aim to preserve the vascular surgical knowledge and skills on par with the medical and interventional measures and further develop them. Only large, interdisciplinary guided vascular centres will be able to provide timely diagnosis and therapy, to deal with the growing multi-morbidity of the patient, to perform complex therapies even in an acute emergency and due to sufficient number of cases to present with well-trained and experienced teams. These requirements are mandatory to decrease patients' mortality step by step.

  16. Vascular Injuries: Trends in Management

    PubMed Central

    Wani, Mohd Lateef; Ahangar, Ab Gani; Ganie, Farooq Ahmad; Wani, Shadab Nabi; Wani, Nasir-ud-din

    2012-01-01

    Abstract Vascular injury presents a great challenge to the emergency resident because these injuries require urgent intervention to prevent loss of life or limb. Sometimes serious vascular injury presents with only subtle or occult signs or symptoms. The patient may present weeks or months after initial injury with symptoms of vascular insufficiency, embolization, pseudoaneurysm, arteriovenous fistula etc. Although the majority of vascular injuries are caused by penetrating trauma from gunshot wounds, stabbing or blast injury, the possibility of vascular injury needs to be considered in patients presenting with displaced long bone fractures, crush injury, prolonged immobilization in a fixed position by tight casts or bandages and various invasive procedures. iatrogenic vascular injuries constitute about 10% of cases in most series; however the incidence is an increasing trend because more endovascular procedures such as angioplasty and cardiac catheterization are being performed routinely. Civilian trauma is more frequently seen in young males. However, it can occur at any age due to road accidents, firearms, bomb blasts and diagnostic procedures. Most of the time, civilian trauma causes less tissue damage. There is an epidemic of vascular injuries in Kashmir valley because of problems in law and order in the past two decades. This review deals with the topic in detail. PMID:24350103

  17. Overgrowth syndromes with vascular anomalies.

    PubMed

    Blei, Francine

    2015-04-01

    Overgrowth syndromes with vascular anomalies encompass entities with a vascular anomaly as the predominant feature vs those syndromes with predominant somatic overgrowth and a vascular anomaly as a more minor component. The focus of this article is to categorize these syndromes phenotypically, including updated clinical criteria, radiologic features, evaluation, management issues, pathophysiology, and genetic information. A literature review was conducted in PubMed using key words "overgrowth syndromes and vascular anomalies" as well as specific literature reviews for each entity and supportive genetic information (e.g., somatic mosaicism). Additional searches in OMIM and Gene Reviews were conducted for each syndrome. Disease entities were categorized by predominant clinical features, known genetic information, and putative affected signaling pathway. Overgrowth syndromes with vascular anomalies are a heterogeneous group of disorders, often with variable clinical expression, due to germline or somatic mutations. Overgrowth can be focal (e.g., macrocephaly) or generalized, often asymmetrically (and/or mosaically) distributed. All germ layers may be affected, and the abnormalities may be progressive. Patients with overgrowth syndromes may be at an increased risk for malignancies. Practitioners should be attentive to patients having syndromes with overgrowth and vascular defects. These patients require proactive evaluation, referral to appropriate specialists, and in some cases, early monitoring for potential malignancies. Progress in identifying vascular anomaly-related overgrowth syndromes and their genetic etiology has been robust in the past decade and is contributing to genetically based prenatal diagnosis and new therapies targeting the putative causative genetic mutations.

  18. Vascular calcification: Mechanisms of vascular smooth muscle cell calcification.

    PubMed

    Leopold, Jane A

    2015-05-01

    Vascular calcification is highly prevalent and, when present, is associated with major adverse cardiovascular events. Vascular smooth muscle cells play an integral role in mediating vessel calcification by undergoing differentiation to osteoblast-like cells and generating matrix vesicles that serve as a nidus for calcium-phosphate deposition in the vessel wall. Once believed to be a passive process, it is now recognized that vascular calcification is a complex and highly regulated process that involves activation of cellular signaling pathways, circulating inhibitors of calcification, genetic factors, and hormones. This review will examine several of the key mechanisms linking vascular smooth muscle cells to vessel calcification that may be targeted to reduce vessel wall mineralization and, thereby, reduce cardiovascular risk.

  19. The Society for Vascular Surgery Vascular Quality Initiative.

    PubMed

    Cronenwett, Jack L; Kraiss, Larry W; Cambria, Richard P

    2012-05-01

    The Society for Vascular Surgery (SVS) Vascular Quality Initiative (VQI) is designed to improve the quality, safety, effectiveness, and cost of vascular health care. It uses the structure of a Patient Safety Organization to permit collection of patient-identified information but protect benchmarked comparisons from legal discovery. The SVS VQI is uniquely organized as a distributed network of regional quality groups to facilitate local translation of registry data into practice change while maintaining the power of a national registry. Detailed data specific to each commonly performed open and endovascular procedure are collected, both in-hospital and at ≥ 1 year of follow-up. Quality measures are reported to physicians and hospitals, which allow anonymous risk-adjusted benchmarking within regions or nationally. All specialties that perform vascular procedures are included, and international participation is encouraged. This review describes the current status of the SVS VQI.

  20. Lasers in digestive endoscopy

    NASA Astrophysics Data System (ADS)

    Brunetaud, Jean Marc; Maunoury, Vincent; Cochelard, Dominique

    1997-01-01

    Lasers were introduced in digestive endoscopy to stop active gastroduodenal hemorrhages. Their use spread progressively to the treatment of chronic hemorrhages from vascular malformations and sessile tumors. Laser face competition from other endoscopic techniques such as electrocoagulation, injection techniques, dilation, stents, and brachytherapy. Many series have reported the efficacy of lasers in digestive endoscopy used for their thermal or photochemical effects. However, they were gradually abandoned for the treatment of hemorrhages because of competition from nonlaser techniques. Lasers are still used for ablation of sessile tumors, but their true impact is difficult to evaluate. Modern methods of technology assessment did not allow gastroenterologists to clearly define the place of lasers among surgery, radio-chemotherapy, and other endoscopic techniques, and data on the daily use of lasers are not available. Therefore, the conclusion can only be subjective. The best current application of thermal lasers appears to be in the treatment of rectosigmoid villous adenomas in elderly patients. Small superficial rectal cancers may also become a good subject due to the impact of endoscopic ultrasonography. Early lesions with multifocal or diffuse disease such as early esophageal cancers could be the most promising subject of application for photodynamic therapy in the future.

  1. Defining excellence in vascular neurosurgery.

    PubMed

    Sanai, Nader; Spetzler, Robert F

    2010-01-01

    Success as a vascular neurosurgeon almost always begins with passion, an inherent love for the work that drives an insatiable desire for personal improvement. A personal definition of excellence in vascular neurosurgery includes several fundamental qualities: mastery of the basics, refinement of technique, advancement of technology, investigative study, advanced decision making, microsurgical innovation, a well-rounded surgical armamentarium, and a lifelong commitment to teaching. Ultimately, the reward for these efforts is the ability to influence generations to come, particularly as one follows the rising careers of former trainees, each redefining the term "excellence" in vascular neurosurgery.

  2. Retina vascular network recognition

    NASA Astrophysics Data System (ADS)

    Tascini, Guido; Passerini, Giorgio; Puliti, Paolo; Zingaretti, Primo

    1993-09-01

    The analysis of morphological and structural modifications of the retina vascular network is an interesting investigation method in the study of diabetes and hypertension. Normally this analysis is carried out by qualitative evaluations, according to standardized criteria, though medical research attaches great importance to quantitative analysis of vessel color, shape and dimensions. The paper describes a system which automatically segments and recognizes the ocular fundus circulation and micro circulation network, and extracts a set of features related to morphometric aspects of vessels. For this class of images the classical segmentation methods seem weak. We propose a computer vision system in which segmentation and recognition phases are strictly connected. The system is hierarchically organized in four modules. Firstly the Image Enhancement Module (IEM) operates a set of custom image enhancements to remove blur and to prepare data for subsequent segmentation and recognition processes. Secondly the Papilla Border Analysis Module (PBAM) automatically recognizes number, position and local diameter of blood vessels departing from optical papilla. Then the Vessel Tracking Module (VTM) analyses vessels comparing the results of body and edge tracking and detects branches and crossings. Finally the Feature Extraction Module evaluates PBAM and VTM output data and extracts some numerical indexes. Used algorithms appear to be robust and have been successfully tested on various ocular fundus images.

  3. Constructal vascularized structures

    NASA Astrophysics Data System (ADS)

    Cetkin, Erdal

    2015-06-01

    Smart features such as self-healing and selfcooling require bathing the entire volume with a coolant or/and healing agent. Bathing the entire volume is an example of point to area (or volume) flows. Point to area flows cover all the distributing and collecting kinds of flows, i.e. inhaling and exhaling, mining, river deltas, energy distribution, distribution of products on the landscape and so on. The flow resistances of a point to area flow can be decreased by changing the design with the guidance of the constructal law, which is the law of the design evolution in time. In this paper, how the flow resistances (heat, fluid and stress) can be decreased by using the constructal law is shown with examples. First, the validity of two assumptions is surveyed: using temperature independent Hess-Murray rule and using constant diameter ducts where the duct discharges fluid along its edge. Then, point to area types of flows are explained by illustrating the results of two examples: fluid networks and heating an area. Last, how the structures should be vascularized for cooling and mechanical strength is documented. This paper shows that flow resistances can be decreased by morphing the shape freely without any restrictions or generic algorithms.

  4. History of lasers in dermatology.

    PubMed

    Geiges, Michael L

    2011-01-01

    In the 1950s, based on the theory of stimulating radiant energy published by Albert Einstein in 1916, the collaboration of physicists and electrical engineers, searching for monochromatic radiation to study the spectra of molecules, led to the invention of the first laser in 1960. Ophthalmologists and dermatologists were the first to study the biological effects and therapeutic possibilities of laser beams. The construction of new laser systems emitting energy at different wavelengths or with different durations, as well as the development of new concepts of the biomedical effects, led to its broad use in surgery in the treatment of vascular and pigmented lesions as well as cosmetic applications.

  5. BMP signaling in vascular diseases.

    PubMed

    Cai, Jie; Pardali, Evangelia; Sánchez-Duffhues, Gonzalo; ten Dijke, Peter

    2012-07-01

    Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-β (TGF-β) family that signal via type I and type II serine/threonine kinase receptors and intracellular Smad transcription factors. BMPs are multifunctional regulators of development and tissue homeostasis and they were initially characterized as inducers of bone regeneration. Genetic studies in humans and mice showed that perturbations in BMP signaling lead to various diseases, such as skeletal diseases, vascular diseases and cancer. Mutations in BMP type II receptor and BMP type I receptor/activin receptor-like kinase 1 have been linked to pulmonary arterial hypertension and hereditary hemorrhagic telangiectasia, respectively. BMPs have also been implicated in promoting vascular calcification and tumor angiogenesis. In this review we discuss the role of BMP signaling in vascular diseases and the value of BMP signaling as a vascular disease marker or a therapeutic target. PMID:22710160

  6. Biomaterials for vascular tissue engineering

    PubMed Central

    Ravi, Swathi; Chaikof, Elliot L

    2010-01-01

    Cardiovascular disease is the leading cause of mortality in the USA. The limited availability of healthy autologous vessels for bypass grafting procedures has led to the fabrication of prosthetic vascular conduits. While synthetic polymers have been extensively studied as substitutes in vascular engineering, they fall short of meeting the biological challenges at the blood–material interface. Various tissue engineering strategies have emerged to address these flaws and increase long-term patency of vascular grafts. Vascular cell seeding of scaffolds and the design of bioactive polymers for in situ arterial regeneration have yielded promising results. This article describes the advances made in biomaterials design to generate suitable materials that not only match the mechanical properties of native vasculature, but also promote cell growth, facilitate extracellular matrix production and inhibit thrombogenicity. PMID:20017698

  7. Tissue engineering: Perfusable vascular networks

    NASA Astrophysics Data System (ADS)

    Forgacs, Gabor

    2012-09-01

    A rapid vascular casting approach that uses carbohydrate glass as a sacrificial template allows tissues to be built that can be kept alive for longer in the laboratory until needed for transplantation.

  8. Measuring Vascular Permeability In Vivo.

    PubMed

    Meijer, Eelco F J; Baish, James W; Padera, Timothy P; Fukumura, Dai

    2016-01-01

    Over the past decades, in vivo vascular permeability measurements have provided significant insight into vascular functions in physiological and pathophysiological conditions such as the response to pro- and anti-angiogenic signaling, abnormality of tumor vasculature and its normalization, and delivery and efficacy of therapeutic agents. Different approaches for vascular permeability measurements have been established. Here, we describe and discuss a conventional 2D imaging method to measure vascular permeability, which was originally documented by Gerlowski and Jain in 1986 (Microvasc Res 31:288-305, 1986) and further developed by Yuan et al. in the early 1990s (Microvasc Res 45:269-289, 1993; Cancer Res 54:352-3356, 1994), and our recently developed 3D imaging method, which advances the approach originally described by Brown et al. in 2001 (Nat Med 7:864-868, 2001). PMID:27581015

  9. How to Prevent Vascular Disease

    MedlinePlus

    ... or 911 immediately. @ 2016 Vascular Cures is a tax-exempt, nonprofit organization tax ID#: 94-2825216 as described in the Section ... 3) of the Internal Revenue Code. Donations are tax deductible. 555 Price Ave., Suite 180, Redwood City, ...

  10. Social media in vascular surgery.

    PubMed

    Indes, Jeffrey E; Gates, Lindsay; Mitchell, Erica L; Muhs, Bart E

    2013-04-01

    There has been a tremendous growth in the use of social media to expand the visibility of various specialties in medicine. The purpose of this paper is to describe the latest updates on some current applications of social media in the practice of vascular surgery as well as existing limitations of use. This investigation demonstrates that the use of social networking sites appears to have a positive impact on vascular practice, as is evident through the incorporation of this technology at the Cleveland Clinic and by the Society for Vascular Surgery into their approach to patient care and physician communication. Overall, integration of social networking technology has current and future potential to be used to promote goals, patient awareness, recruitment for clinical trials, and professionalism within the specialty of vascular surgery. PMID:23321344

  11. Laser Technology.

    ERIC Educational Resources Information Center

    Gauger, Robert

    1993-01-01

    Describes lasers and indicates that learning about laser technology and creating laser technology activities are among the teacher enhancement processes needed to strengthen technology education. (JOW)

  12. Vascular heterogeneity in the kidney.

    PubMed

    Molema, Grietje; Aird, William C

    2012-03-01

    Blood vessels and their endothelial lining are uniquely adapted to the needs of the underlying tissue. The structure and function of the vasculature varies both between and within different organs. In the kidney, the vascular architecture is designed to function both in oxygen/nutrient delivery and filtration of blood according to the homeostatic needs of the body. Here, we review spatial and temporal differences in renal vascular phenotypes in both health and disease.

  13. Multimodality imaging of vascular anomalies.

    PubMed

    Restrepo, Ricardo

    2013-03-01

    Vascular malformations and hemangiomas are common in children but remain a source of confusion during diagnosis, in part because of the lack of a uniform terminology. With the existing treatments for hemangiomas and vascular malformations, it is important to make the correct diagnosis initially to prevent adverse physical and emotional sequelae in not only the child but also the family. The diagnosis of vascular malformations is made primarily by the clinician and based on the physical exam. Imaging is carried out using predominantly ultrasound (US) and magnetic resonance imaging (MRI), which are complementary modalities. In most cases of vascular anomalies, US is the first line of imaging as it is readily available, less expensive, lacks ionizing radiation and does not require sedation. MRI is also of great help for further characterizing the lesions. Conventional arteriography is reserved for cases that require therapeutic intervention, more commonly for arteriovenous malformations. Radiographs usually play no role in diagnosing vascular anomalies in children. In this article, the author describes the terminology and types of hemangiomas and vascular malformations and their clinical, histological features, as well as the imaging approach and appearance.

  14. Laser treatment of periocular skin conditions.

    PubMed

    Yates, Breton; Que, Syril Keena T; D'Souza, Logan; Suchecki, Jeanine; Finch, Justin J

    2015-01-01

    Advances in laser technology in recent decades have increased the options for the treatment of dermatologic conditions of the eye and eyelid. Benign tumors can be laser-ablated with relative ease, and vascular and melanocytic lesions can be precisely targeted with modern lasers. In this contribution, we review treatment of periocular pigmented lesions, including melanocytic nevi and nevus of Ota; vascular lesions including telangiectasias, port wine stains, and infantile hemangiomas; hair removal; eyeliner tattoo removal; laser ablation of common benign periocular tumors, such as syringomas, xanthelasma, milia, and seborrheic keratoses; and laser resurfacing. The recent advent of fractionated laser technology has resulted in dramatically decreased healing times for periocular skin resurfacing and fewer adverse effects. Fractionated laser resurfacing has now nearly supplanted traditional full-field laser resurfacing, and safe treatment of rhytides on the thin skin of the eyelids is possible. Proper eye protection is, of course, essential when using lasers near the eye. Patient preparation, safety precautions, and risks--intraocular and extraocular--are discussed herein. As laser technology continues to advance, we are sure to see improvements in current treatments, as well as development of new applications of cutaneous lasers. PMID:25704939

  15. Observation and stimulation of biological processes using excimer lasers

    NASA Astrophysics Data System (ADS)

    Greulich, Karl-Otto; Wolfrum, Juergen M.

    1990-06-01

    Examples are given for the application o f high power excimer lasers in spectroscopy and processing of biological material. An excimer-laser pumped dye laser serves as light source for a pulsed UV Raman spectrometer which allows resonant Raman studies on nucleic acids. Experiments on the pH induced double helix formation of poly adenylic acid are described. By combining the excimer laser with a distributed feedback dye laser and a streak camera, a picosecond UV fluorescence spectrometer is built up .Tyrosine fluorescence lifetimes of selected tryptophan free peptides with up to 9 amino acids can be explained in a surprisingly simple way: only the directly neighbouring amino acid on the C-terminal side and only a few amino acids on the N-terminal side have an influence on the fluorecscence lifetime of these peptides. Besides spectroscopic applications, the excimer laser serves as light source for processing of biological material. For medical applications , high power UV Laser light has to be transmitted through light guides. A tapered light guide transmitting more than GW/cm2 is described. Microprocessing of biological material with accuracies of a few hundred nanometers can be performed when an excimer pumped dye laser is coupled into a microscope .The resulting UV laser microbeam can be used to introduce foreign genetic material into plant cells, tissues and subcellular organelles such as mitochondria and chioroplasts. Selected pairs of different cells can be fused in the UV laser microbeam under total microscopic control. Finally, one can microdissect human chromosomes and isolate DNA probes for the analysis of human disease.

  16. RNA Sequencing of Laser-Capture Microdissected Compartments of the Maize Kernel Identifies Regulatory Modules Associated with Endosperm Cell Differentiation[OPEN

    PubMed Central

    Zhan, Junpeng; Thakare, Dhiraj; Ma, Chuang; Lloyd, Alan; Nixon, Neesha M.; Arakaki, Angela M.; Burnett, William J.; Logan, Kyle O.; Wang, Dongfang; Wang, Xiangfeng; Drews, Gary N.; Yadegari, Ramin

    2015-01-01

    Endosperm is an absorptive structure that supports embryo development or seedling germination in angiosperms. The endosperm of cereals is a main source of food, feed, and industrial raw materials worldwide. However, the genetic networks that regulate endosperm cell differentiation remain largely unclear. As a first step toward characterizing these networks, we profiled the mRNAs in five major cell types of the differentiating endosperm and in the embryo and four maternal compartments of the maize (Zea mays) kernel. Comparisons of these mRNA populations revealed the diverged gene expression programs between filial and maternal compartments and an unexpected close correlation between embryo and the aleurone layer of endosperm. Gene coexpression network analysis identified coexpression modules associated with single or multiple kernel compartments including modules for the endosperm cell types, some of which showed enrichment of previously identified temporally activated and/or imprinted genes. Detailed analyses of a coexpression module highly correlated with the basal endosperm transfer layer (BETL) identified a regulatory module activated by MRP-1, a regulator of BETL differentiation and function. These results provide a high-resolution atlas of gene activity in the compartments of the maize kernel and help to uncover the regulatory modules associated with the differentiation of the major endosperm cell types. PMID:25783031

  17. Laser capture microdissection coupled with RNA-seq reveals that short cuboidal trophoblast cells of the porcine placenta possess a transcriptome consistent with a migratory cell type

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The porcine placenta is classified as epitheliochorial with a diffuse distribution of chorionic villi that do not invade maternal tissues. Thus, the maternal and fetal bloodstreams are separated by six distinct tissue layers. The porcine fetal-maternal interface (FMI) is substantially folded to incr...

  18. Temporal Expression of miRNAs in Laser Capture Microdissected Palate Medial Edge Epithelium from Tgfβ3(-/-) Mouse Fetuses.

    PubMed

    Warner, Dennis; Ding, Jixiang; Mukhopadhyay, Partha; Brock, Guy; Smolenkova, Irina A; Seelan, Ratnam S; Webb, Cindy L; Wittliff, James L; Greene, Robert M; Pisano, M Michele

    2015-01-01

    Clefting of the secondary palate is the most common birth defect in humans. Midline fusion of the bilateral palatal processes is thought to involve apoptosis, epithelial to mesenchymal transition, and cell migration of the medial edge epithelium (MEE), the specialized cells of the palate that mediate fusion of the palatal processes during fetal development. Data presented in this manuscript are the result of analyses designed to identify microRNAs that are expressed and regulated by TGFβ3 in developing palatal MEE. The expression of 7 microRNAs was downregulated and 1 upregulated in isolated MEE from wildtype murine fetuses on gestational day (GD) 13.5 to GD14.5 (prior to and during epithelial fusion of the palatal processes, respectively). Among this group were miRNAs linked to apoptosis (miR-378) and epithelial to mesenchymal transformation (miR-200b, miR-205, and miR-93). Tgfβ3(-/-) fetuses, which present with a complete and isolated cleft of the secondary palate, exhibited marked dysregulation of distinct miRNAs both in the palatal MEE and mesenchyme when compared to comparable wild-type tissue. These included, among others, miRNAs known to affect apoptosis (miR-206 and miR-186). Dysregulation of miRNAs in the mesenchyme underlying the palatal MEE of Tgfβ3(-/-) fetuses is also discussed in relation to epithelial-mesenchymal transformation of the MEE. These results are the first systematic analysis of the expression of microRNAs in isolated fetal palatal epithelium and mesenchyme. Moreover, analysis of the Tgfβ3 knockout mouse model has enabled identification of miRNAs with altered expression that may contribute to the cleft palate phenotype.

  19. Use of laser capture microdissection for the assessment of equine lamellar basal epithelial cell signalling in the early stages of laminitis

    PubMed Central

    Leise, B. S.; Watts, M.; Roy, S.; Yilmaz, S.; Alder, H.; Belknap, J. K.

    2016-01-01

    Summary Reason for performing study Dysadhesion of the laminar basal epithelial cells (LBEC) from the underlying dermis is the central event leading to structural failure in equine laminitis. Although many studies of sepsis-related laminitis have reported multiple events occurring throughout the lamellar tissue, there is minimal information regarding signalling events occurring specifically in the LBEC. Objectives To determine the signalling events in the LBECs during the early stages of carbohydrate induced laminitis. Study Design Experimental study. Methods Eight horses were given an overload of carbohydrate (corn starch mixture, CHO) via nasogastric tube. Prior to administration of CHO, lamellar biopsies were taken from the left fore foot (CON). Biopsies were taken from the left hind foot at the onset of fever (DEV) and from the right fore foot at the onset of lameness (OG1). Laminar basal epithelial cells (LBECs) were isolated from cryosections using a LCM microscope. Next generation sequencing (RNA-Seq) was used to identify transcripts expressed in the LBECs for each time point and bioinformatic analysis was performed with thresholds for between group comparisons set at a greater than 2-fold change and p-value ≤0.05. Results Forty genes (22 increased/18 decreased) were significantly different from DEV time vs. CON and 107 genes (57 increased/50 decreased) were significantly different from OG1 time vs. CON. Significant increases in inflammatory genes were present in addition to significantly altered expression of genes related to extracellular matrix composition, stability and turnover. Conclusions Inflammatory response and extracellular matrix regulation signalling was strongly represented at the DEV and OG1 times. These results indicate that the LBEC is not only a casualty but also an active participant in lamellar events leading to structural failure of the digital lamellae in equine laminitis. PMID:24750316

  20. Differentiation of Multipotent Vascular Stem Cells Contributes to Vascular Diseases

    PubMed Central

    Tang, Zhenyu; Wang, Aijun; Yuan, Falei; Yan, Zhiqiang; Liu, Bo; Chu, Julia S.; Helms, Jill A.

    2012-01-01

    It is generally accepted that the de-differentiation of smooth muscle cells (SMCs) from contractile to proliferative/synthetic phenotype has an important role during vascular remodeling and diseases. Here we provide evidence that challenges this theory. We identify a new type of multipotent vascular stem cell (MVSC) in blood vessel wall. MVSCs express markers including Sox17, Sox10 and S100β, are cloneable, have telomerase activity, and can differentiate into neural cells and mesenchymal stem cell (MSC)-like cells that subsequently differentiate into SMCs. On the other hand, we use lineage tracing with smooth muscle myosin heavy chain as a marker to show that MVSCs and proliferative or synthetic SMCs do not arise from the de-differentiation of mature SMCs. Upon vascular injuries, MVSCs, instead of SMCs, become proliferative, and MVSCs can differentiate into SMCs and chondrogenic cells, thus contributing to vascular remodeling and neointimal hyperplasia. These findings support a new hypothesis that the differentiation of MVSCs, rather than the de-differentiation of SMCs, contributes to vascular remodeling and diseases. PMID:22673902

  1. Early experience on peripheral vascular application of the vascular plugs

    PubMed Central

    Rohit, Manoj Kumar; Sinha, Alok Kumar; Kamana, Naveen Krishna

    2013-01-01

    Background Transcatheter closure of various congenital and acquired vascular malformations with Amplatzer Vascular plugs I and II has been established. Here we present our experience with device closure. Materials and methods Between October 2006 and August 2012, nine (three males and six females) patients aged between 11 months and 62 years (mean age 19 years) underwent percutaneous device closure with AVP I and II vascular plugs for congenital and acquired arteriovenous malformation and cardiac diverticulum are presented here. Results One case of coronary cameral fistula, four cases of pulmonary arteriovenous fistula, one case of large major aortopulmonary collaterals (in tetralogy of Fallot closed before intracardiac repair), one case of congenital cardiac diverticulum, one case of fistula between external carotid artery and internal jugular vein and one case of iatrogenic carotid jugular fistula were successfully closed with AVP I and II plugs. Overall in nine cases, 16 AVP I and II plugs were deployed to occlude feeding vessels and one cardiac diverticulum. The technical success rate was 100%. No major complications were observed. Conclusion Amplatzer vascular plugs can be used successfully for closure of various congenital and acquired vascular malformations with good result. PMID:24206877

  2. Vascular endothelial growth factor coordinates islet innervation via vascular scaffolding

    PubMed Central

    Reinert, Rachel B.; Cai, Qing; Hong, Ji-Young; Plank, Jennifer L.; Aamodt, Kristie; Prasad, Nripesh; Aramandla, Radhika; Dai, Chunhua; Levy, Shawn E.; Pozzi, Ambra; Labosky, Patricia A.; Wright, Christopher V. E.; Brissova, Marcela; Powers, Alvin C.

    2014-01-01

    Neurovascular alignment is a common anatomical feature of organs, but the mechanisms leading to this arrangement are incompletely understood. Here, we show that vascular endothelial growth factor (VEGF) signaling profoundly affects both vascularization and innervation of the pancreatic islet. In mature islets, nerves are closely associated with capillaries, but the islet vascularization process during embryonic organogenesis significantly precedes islet innervation. Although a simple neuronal meshwork interconnects the developing islet clusters as they begin to form at E14.5, the substantial ingrowth of nerve fibers into islets occurs postnatally, when islet vascularization is already complete. Using genetic mouse models, we demonstrate that VEGF regulates islet innervation indirectly through its effects on intra-islet endothelial cells. Our data indicate that formation of a VEGF-directed, intra-islet vascular plexus is required for development of islet innervation, and that VEGF-induced islet hypervascularization leads to increased nerve fiber ingrowth. Transcriptome analysis of hypervascularized islets revealed an increased expression of extracellular matrix components and axon guidance molecules, with these transcripts being enriched in the islet-derived endothelial cell population. We propose a mechanism for coordinated neurovascular development within pancreatic islets, in which endocrine cell-derived VEGF directs the patterning of intra-islet capillaries during embryogenesis, forming a scaffold for the postnatal ingrowth of essential autonomic nerve fibers. PMID:24574008

  3. Understanding lasers

    SciTech Connect

    Gibilisco, S.

    1989-01-01

    Covering all different types of laser applications-Gibilisco offers an overview of this fascinating phenomenon of light. Here he describes what lasers are and how they work and examines in detail the different kinds of lasers in use today. Topics of particular interest include: the way lasers work; the different kinds of lasers; infrared, ultraviolet and x-ray lasers; use of lasers in industry and manufacturing; use of lasers for long-distance communications; fiberoptic communications; the way laser shows work; the reality of Star Wars; lasers in surgical and medical applications; and holography and the future of laser technology.

  4. Proteomic analysis of human papillomavirus-related oral squamous cell carcinoma: identification of thioredoxin and epidermal-fatty acid binding protein as upregulated protein markers in microdissected tumor tissue.

    PubMed

    Melle, Christian; Ernst, Günther; Winkler, Robert; Schimmel, Bettina; Klussmann, Jens Peter; Wittekindt, Claus; Guntinas-Lichius, Orlando; von Eggeling, Ferdinand

    2009-04-01

    Human papillomavirus (HPV) infection has been identified as an etiologic agent for a subset of oral squamous cell carcinoma (OSCC) with increasing incidence. HPV DNA-positivity may confer better prognosis but the related oncogenic mechanisms are unknown. For the identification of HPV relevant proteins, we analyzed microdissected cells from HPV DNA-positive (n = 17) and HPV DNA-negative (n = 7) OSCC tissue samples. We identified 18 proteins from tumor tissues by peptide fingerprint mapping and SELDI MS that were separated using 2-DE. Among a number of signals that were detected as significantly different in the protein profiling analysis, we identified thioredoxin (TRX) and epidermal-fatty acid binding protein as upregulated in HPV related tumor tissue. This study, investigating for the first time proteomic changes in microdissected HPV infected tumor tissue, provides an indication on the oncogenic potential of viruses. PMID:19337991

  5. Direct cDNA selection with DNA microdissected from mouse chromosome 16: isolation of novel clones and construction of a partial transcription map of the C3-C4 region.

    PubMed

    Wei, J; Hodes, M E; Wang, Y; Feng, Y; Ghetti, B; Dlouhy, S R

    1996-08-01

    A group of cDNA segments was selected by direct hybridization of mouse cerebellar cDNAs against genomic DNA pools generated by microdissection of the mouse chromosome 16 (MMU16) C3-C4 region. After elimination of repetitive sequences and adjustment for redundancy among clones, 34 novel cDNA fragments were isolated. The MMU16 origin of clones was confirmed by genetic linkage mapping. Reverse transcription PCR indicated that approximately 68% of the cDNAs represent transcripts that are expressed in adult mouse cerebellum. Northern blotting showed that some of these are predominantly or solely expressed in brain. This work demonstrates that DNA microdissected from banded MMU16 can be used for direct cDNA selection, thus enabling construction of a new, region-specific partial transcription map. This selected cDNA library should be a useful reagent for further molecular neurobiological studies.

  6. Laser-based technique for controlled damage of mesenchymal cell spheroids: a first step in studying reparation in vitro

    PubMed Central

    Ilina, I. V.; Zurina, I. M.; Roskova, A. E.; Gorkun, A. A.; Ovchinnikov, A. V.; Agranat, M. B.; Saburina, I. N.

    2016-01-01

    ABSTRACT Modern techniques of laser microsurgery of cell spheroids were used to develop a new simple reproducible model for studying repair and regeneration in vitro. Nanosecond laser pulses (wavelength 355 nm, frequency 100 Hz, pulse duration 2 ns) were applied to perform a microdissection of the outer and the inner zones of human bone marrow multipotent mesenchymal stromal cells (BM MMSC) spheroids. To achieve effective dissection and preservation of spheroid viability, the energy of laser pulses was optimized and adjusted in the range 7-9 μJ. After microdissection, the edges of the wound surface opened and the angular opening reached a value of more than 180°. The destruction of the initial spheroid structure was observed in the wound area, with surviving cells changing their shape into a round one. Partial restoration of a spheroid form took place in the first six hours. The complete structure restoration accompanying the reparative processes occurred gradually over seven days due to remodelling of surviving cells. PMID:27334698

  7. Vascular tissue engineering: towards the next generation vascular grafts.

    PubMed

    Naito, Yuji; Shinoka, Toshiharu; Duncan, Daniel; Hibino, Narutoshi; Solomon, Daniel; Cleary, Muriel; Rathore, Animesh; Fein, Corey; Church, Spencer; Breuer, Christopher

    2011-04-30

    The application of tissue engineering technology to cardiovascular surgery holds great promise for improving outcomes in patients with cardiovascular diseases. Currently used synthetic vascular grafts have several limitations including thrombogenicity, increased risk of infection, and lack of growth potential. We have completed the first clinical trial evaluating the feasibility of using tissue engineered vascular grafts (TEVG) created by seeding autologous bone marrow-derived mononuclear cells (BM-MNC) onto biodegradable tubular scaffolds. Despite an excellent safety profile, data from the clinical trial suggest that the primary graft related complication of the TEVG is stenosis, affecting approximately 16% of grafts within the first seven years after implantation. Continued investigation into the cellular and molecular mechanisms underlying vascular neotissue formation will improve our basic understanding and provide insights that will enable the rationale design of second generation TEVG. PMID:21421015

  8. Vascular changes in hard palate sialolipoma: Sialoangiolipoma or vascular malformation?

    PubMed Central

    Handra-Luca, Adriana

    2015-01-01

    Palate sialolipomas are rare. Less than 10 cases located in the hard palate are reported to our knowledge. We report a case of hard palate sialolipoma, peculiar by the intratumor vascular patterns. A 67-year-old man presented with a 1.5 cm lesion of the oral hard palate. The lesion was surgically resected. On microscopy, the lesion, partly encapsulated, consisted of a proliferation of mature adipocytes containing normal minor salivary gland tissue and branching intratumoral vessels of varied size with irregularly thickened wall and papillary projections or tufts. The microscopic features of the tumor we report suggest that vascular malformation-like patterns may occur in sialolipomas of the hard palate. This morphological vascular peculiarity should be acknowledged since it may represent source of hemorrhage. PMID:26604516

  9. Laser applications in neurosurgery

    NASA Astrophysics Data System (ADS)

    Cerullo, Leonard J.

    1985-09-01

    beam makes the laser superior to all conventional destructive instruments. 4)|The coagulative properties of certain chromophoric lasers has allowed a new attack on certain vascular tumors and malformations of the brain and spinal cord which had been operated only with trepidation or not at all. Early reports are sobering but encouraging. 5)|Finally, the use of the laser with tissue photosensitization, albeit it in its infancy, offers great promise. This is particularly true in the case of primary brain cancer, where the infiltration of tumorous tissue among normal pathways precludes the classical oncologic surgery practice of resection of a "safe margin". The ability to track and destroy these cells, without affecting adjacent cells, may be the greatest single contribution of the laser to neurosurgery in the future. The present applications of the laser are relatively crude by comparison with what is expected. Endoscopic laser surgery, both vascular and subarachnoid, will diminish morbidity and improve results. From the exotic treatment of aneurysms and arteriovenous malformations of the brain to the mundane care of herniated disks of the spine, it is anticipated that the laser will play an important role. The use of a laser, coupled with computerized imagining devices, will allow increasing precision in arrival to and treatment of deep seated lesions of the brain, brainstem, and spinal cord. The use of different wavelengths, perhaps in the X-ray and ultraviolet spectra, will allow increasing precision with decreasing invasion. Manipulation of wavelength, time, and treatment area will allow subcellular surgery, perhaps in the treatment of personality disorders and movement disorders as well as epilepsy. Tissue welding will allow heightened regenerative and recuperative powers to be exploited. The possibility of laser biostimulation must also be considered. In short, it appears that the future of the laser in neurosurgery is limited only by the imagination of the

  10. Vascular potassium channels in NVC.

    PubMed

    Yamada, K

    2016-01-01

    It has long been proposed that the external potassium ion ([K(+)]0) works as a potent vasodilator in the dynamic regulation of local cerebral blood flow. Astrocytes may play a central role for producing K(+) outflow possibly through calcium-activated potassium channels on the end feet, responding to a rise in the intracellular Ca(2+) concentration, which might well reflect local neuronal activity. A mild elevation of [K(+)]0 in the end feet/vascular smooth muscle space could activate Na(+)/K(+)-ATPase concomitant with inwardly rectifying potassium (Kir) channels in vascular smooth muscle cells, leading to a hyperpolarization of vascular smooth muscle and relaxation of smooth muscle actin-positive vessels. Also proposed notion is endothelial calcium-activated potassium channels and/or inwardly rectifying potassium channel-mediated hyperpolarization of vascular smooth muscle. A larger elevation of [K(+)]0, which may occur pathophysiologically in such as spreading depression or stroke, can trigger a depolarization of vascular smooth muscle cells and vasoconstriction instead. PMID:27130411

  11. Adverse Outcome Pathways for Embryonic Vascular Disruption and Alternative Methods to Identify Chemical Vascular Disruptor

    EPA Science Inventory

    Chemically induced vascular toxicity during embryonic development can result in a wide range of adverse prenatal outcomes. We used information from genetic mouse models linked to phenotypic outcomes and a vascular toxicity knowledge base to construct an embryonic vascular disrupt...

  12. Potential candidate genomic biomarkers of drug induced vascular injury in the rat

    SciTech Connect

    Dalmas, Deidre A.; Scicchitano, Marshall S.; Mullins, David; Hughes-Earle, Angela; Tatsuoka, Kay; Magid-Slav, Michal; Frazier, Kendall S.; Thomas, Heath C.

    2011-12-15

    Drug-induced vascular injury is frequently observed in rats but the relevance and translation to humans present a hurdle for drug development. Numerous structurally diverse pharmacologic agents have been shown to induce mesenteric arterial medial necrosis in rats, but no consistent biomarkers have been identified. To address this need, a novel strategy was developed in rats to identify genes associated with the development of drug-induced mesenteric arterial medial necrosis. Separate groups (n = 6/group) of male rats were given 28 different toxicants (30 different treatments) for 1 or 4 days with each toxicant given at 3 different doses (low, mid and high) plus corresponding vehicle (912 total rats). Mesentery was collected, frozen and endothelial and vascular smooth muscle cells were microdissected from each artery. RNA was isolated, amplified and Affymetrix GeneChip Registered-Sign analysis was performed on selectively enriched samples and a novel panel of genes representing those which showed a dose responsive pattern for all treatments in which mesenteric arterial medial necrosis was histologically observed, was developed and verified in individual endothelial cell- and vascular smooth muscle cell-enriched samples. Data were confirmed in samples containing mesentery using quantitative real-time RT-PCR (TaqMan Trade-Mark-Sign ) gene expression profiling. In addition, the performance of the panel was also confirmed using similarly collected samples obtained from a timecourse study in rats given a well established vascular toxicant (Fenoldopam). Although further validation is still required, a novel gene panel has been developed that represents a strategic opportunity that can potentially be used to help predict the occurrence of drug-induced mesenteric arterial medial necrosis in rats at an early stage in drug development. -- Highlights: Black-Right-Pointing-Pointer A gene panel was developed to help predict rat drug-induced mesenteric MAN. Black

  13. Laser treatment of port-wine stains

    PubMed Central

    Brightman, Lori A; Geronemus, Roy G; Reddy, Kavitha K

    2015-01-01

    Port-wine stains are a type of capillary malformation affecting 0.3% to 0.5% of the population. Port-wine stains present at birth as pink to erythematous patches on the skin and/or mucosa. Without treatment, the patches typically darken with age and may eventually develop nodular thickening or associated pyogenic granuloma. Laser and light treatments provide improvement through selective destruction of vasculature. A variety of vascular-selective lasers may be employed, with the pulsed dye laser being the most common and well studied. Early treatment produces more optimal results. Advances in imaging and laser treatment technologies demonstrate potential to further improve clinical outcomes. PMID:25624768

  14. Vascular gene expression: a hypothesis

    PubMed Central

    Martínez-Navarro, Angélica C.; Galván-Gordillo, Santiago V.; Xoconostle-Cázares, Beatriz; Ruiz-Medrano, Roberto

    2013-01-01

    The phloem is the conduit through which photoassimilates are distributed from autotrophic to heterotrophic tissues and is involved in the distribution of signaling molecules that coordinate plant growth and responses to the environment. Phloem function depends on the coordinate expression of a large array of genes. We have previously identified conserved motifs in upstream regions of the Arabidopsis genes, encoding the homologs of pumpkin phloem sap mRNAs, displaying expression in vascular tissues. This tissue-specific expression in Arabidopsis is predicted by the overrepresentation of GA/CT-rich motifs in gene promoters. In this work we have searched for common motifs in upstream regions of the homologous genes from plants considered to possess a “primitive” vascular tissue (a lycophyte), as well as from others that lack a true vascular tissue (a bryophyte), and finally from chlorophytes. Both lycophyte and bryophyte display motifs similar to those found in Arabidopsis with a significantly low E-value, while the chlorophytes showed either a different conserved motif or no conserved motif at all. These results suggest that these same genes are expressed coordinately in non-vascular plants; this coordinate expression may have been one of the prerequisites for the development of conducting tissues in plants. We have also analyzed the phylogeny of conserved proteins that may be involved in phloem function and development. The presence of CmPP16, APL, FT, and YDA in chlorophytes suggests the recruitment of ancient regulatory networks for the development of the vascular tissue during evolution while OPS is a novel protein specific to vascular plants. PMID:23882276

  15. Vascular ultrasound for atherosclerosis imaging

    PubMed Central

    de Korte, Chris L.; Hansen, Hendrik H. G.; van der Steen, Anton F. W.

    2011-01-01

    Cardiovascular disease is a leading cause of death in the Western world. Therefore, detection and quantification of atherosclerotic disease is of paramount importance to monitor treatment and possible prevention of acute events. Vascular ultrasound is an excellent technique to assess the geometry of vessel walls and plaques. The high temporal as well as spatial resolution allows quantification of luminal area and plaque size and volume. While carotid arteries can be imaged non-invasively, scanning of coronary arteries requires invasive intravascular catheters. Both techniques have already demonstrated their clinical applicability. Using linear array technology, detection of disease as well as monitoring of pharmaceutical treatment in carotid arteries are feasible. Data acquired with intravascular ultrasound catheters have proved to be especially beneficial in understanding the development of atherosclerotic disease in coronary arteries. With the introduction of vascular elastography not only the geometry of plaques but also the risk for rupture of plaques might be identified. These so-called vulnerable plaques are frequently not flow-limiting and rupture of these plaques is responsible for the majority of cerebral and cardiac ischaemic events. Intravascular ultrasound elastography studies have demonstrated a high correlation between high strain and vulnerable plaque features, both ex vivo and in vivo. Additionally, pharmaceutical intervention could be monitored using this technique. Non-invasive vascular elastography has recently been developed for carotid applications by using compound scanning. Validation and initial clinical evaluation is currently being performed. Since abundance of vasa vasorum (VV) is correlated with vulnerable plaque development, quantification of VV might be a unique tool to even prevent this from happening. Using ultrasound contrast agents, it has been demonstrated that VV can be identified and quantified. Although far from routine

  16. Treatment of congential vascular disorders: classification, step program, and therapeutic procedures

    NASA Astrophysics Data System (ADS)

    Philipp, Carsten M.; Poetke, Margitta; Engel-Murke, Frank; Waldschmidt, J.; Berlien, Hans-Peter

    1994-02-01

    Because of the different step programs concerning the preoperative diagnostic and the onset of therapy for the various types of congenital vascular disorders (CVD) a clear classification is important. One has to discern the vascular malformations, including the port wine stain, from the real hemangiomas which are vascular tumors. As former classification, mostly based on histological findings, showed little evidence to a clinical step program, we developed a descriptive classification which allows an early differentiation between the two groups of CVD. In most cases this can be done by a precise medical history of the onset and development of the disorder, a close look to the clinical signs and by Duplex-Ultrasound and MRI-diagnostic. With this protocol and the case adapted use of different lasers and laser techniques we have not seen any severe complications as skin necrosis or nerve lesions.

  17. [Vascular Calcification - Pathological Mechanism and Clinical Application - . Role of vascular smooth muscle cells in vascular calcification].

    PubMed

    Kurabayashi, Masahiko

    2015-05-01

    Vascular calcification is commonly seen with aging, chronic kidney disese (CKD), diabetes, and atherosclerosis, and is closely associated with cardiovascular morbidity and mortality. Vascular calcification has long been regarded as the final stage of degeneration and necrosis of arterial wall and a passive, unregulated process. However, it is now known to be an active and tightly regulated process involved with phenotypic transition of vascular smooth muscle cells (VSMC) that resembles bone mineralization. Briefly, calcium deposits of atherosclerotic plaque consist of hydroxyapatite and may appear identical to fully formed lamellar bone. By using a genetic fate mapping strategy, VSMC of the vascular media give rise to the majority of the osteochondrogenic precursor- and chondrocyte-like cells observed in the calcified arterial media of MGP (- / -) mice. Osteogenic differentiation of VSMC is characterized by the expression of bone-related molecules including bone morphogenetic protein (BMP) -2, Msx2 and osteopontin, which are produced by osteoblasts and chondrocytes. Our recent findings are that (i) Runx2 and Notch1 induce osteogenic differentiation, and (ii) advanced glycation end-product (AGE) /receptor for AGE (RAGE) and palmitic acid promote osteogenic differentiation of VSMC. To understand of the molecular mechanisms of vascular calcification is now under intensive research area. PMID:25926569

  18. [Vascular Calcification - Pathological Mechanism and Clinical Application - . Role of vascular smooth muscle cells in vascular calcification].

    PubMed

    Kurabayashi, Masahiko

    2015-05-01

    Vascular calcification is commonly seen with aging, chronic kidney disese (CKD), diabetes, and atherosclerosis, and is closely associated with cardiovascular morbidity and mortality. Vascular calcification has long been regarded as the final stage of degeneration and necrosis of arterial wall and a passive, unregulated process. However, it is now known to be an active and tightly regulated process involved with phenotypic transition of vascular smooth muscle cells (VSMC) that resembles bone mineralization. Briefly, calcium deposits of atherosclerotic plaque consist of hydroxyapatite and may appear identical to fully formed lamellar bone. By using a genetic fate mapping strategy, VSMC of the vascular media give rise to the majority of the osteochondrogenic precursor- and chondrocyte-like cells observed in the calcified arterial media of MGP (- / -) mice. Osteogenic differentiation of VSMC is characterized by the expression of bone-related molecules including bone morphogenetic protein (BMP) -2, Msx2 and osteopontin, which are produced by osteoblasts and chondrocytes. Our recent findings are that (i) Runx2 and Notch1 induce osteogenic differentiation, and (ii) advanced glycation end-product (AGE) /receptor for AGE (RAGE) and palmitic acid promote osteogenic differentiation of VSMC. To understand of the molecular mechanisms of vascular calcification is now under intensive research area.

  19. Vascular contributions to cognitive impairment

    PubMed Central

    Flores, Alan

    2015-01-01

    Summary Unlike many neurodegenerative causes of cognitive impairment and dementia, vascular damage is preventable. Despite the heterogeneity of vascular cognitive impairment (VCI) and the complexity of its clinical presentations, the potential for limiting progression and changing the trajectory of damage makes it all the more important for physicians to be educated about the syndrome and to remain vigilant when taking care of patients. In this review, we outline an approach to patients with possible VCI, summarize current treatment and prevention guidelines, and provide an overview with case examples. PMID:26124978

  20. Genetic basis for vascular anomalies.

    PubMed

    Kirkorian, A Yasmine; Grossberg, Anna L; Püttgen, Katherine B

    2016-03-01

    The fundamental genetics of many isolated vascular anomalies and syndromes associated with vascular anomalies have been elucidated. The rate of discovery continues to increase, expanding our understanding of the underlying interconnected molecular pathways. This review summarizes genetic and clinical information on the following diagnoses: capillary malformation, venous malformation, lymphatic malformation, arteriovenous malformation, PIK3CA-related overgrowth spectrum (PROS), Proteus syndrome, SOLAMEN syndrome, Sturge-Weber syndrome, phakomatosis pigmentovascularis, congenital hemangioma, verrucous venous malformation, cutaneomucosal venous malformation, blue rubber bleb nevus syndrome, capillary malformation-arteriovenous malformation syndrome, Parkes-Weber syndrome, and Maffucci syndrome. PMID:27607321

  1. Three-region specific microdissection libraries for the long arm of human chromosome 2, regions q33-q35, q31-q32, and q23-q24

    SciTech Connect

    Yu, J.; Tong, S.; Whittier, A.

    1995-09-01

    Three region-specific libraries have been constructed from the long arm of human chromosome 2, including regions 2q33-35 (2Q2 library), 2q31-32 (2Q3) and 2q23-24 (2Q4). Chromosome microdissection and the MboI linker-adaptor microcloning techniques were used in constructing these libraries. The libraries comprised hundreds of thousands of microclones in each library. Approximately half of the microclones in the library contained unique or low-copy number sequence inserts. The insert sizes ranged between 50 and 800 bp, with a mean of 130-190 bp. Southern blot analysis of individual unique sequence microclones showed that 70-94% of the microclones were derived from the dissected region. 31 unique sequence microclones from the 2Q2 library, 31 from 2Q3, and 30 from 2Q4, were analyzed for insert sizes, the hybridizing genomic HindIII fragment sizes, and cross-hybridization to rodent species. These libraries and the short insert microclones derived from the libraries should be useful for high resolution physical mapping, sequence-ready reagents for large scale genomic sequencing, and positional cloning of disease-related genes assigned to these regions, e.g. the recessive familial amyotrophic lateral sclerosis assigned to 2q33-q35, and a type I diabetes susceptibility gene to 2q31-q33. 17 refs., 5 figs., 2 tabs.

  2. Thrombospondin-2 Expression During Retinal Vascular Development and Neovascularization

    PubMed Central

    Fei, Ping; Palenski, Tammy L.; Wang, Shoujian; Gurel, Zafer; Hankenson, Kurt D.; Sorenson, Christine M.

    2015-01-01

    Abstract Purpose: To determine thrombospondin-2 (TSP2) expression and its impact on postnatal retinal vascular development and retinal neovascularization. Methods: The TSP2-deficient (TSP2−/−) mice and a line of TSP2 reporter mice were used to assess the expression of TSP2 during postnatal retinal vascular development and neovascularization. The postnatal retinal vascularization was evaluated using immunostaining of wholemount retinas prepared at different postnatal days by collagen IV staining and/or TSP2 promoter driven green fluorescent protein (GFP) expression. The organization of astrocytes was evaluated by glial fibrillary acidic protein (GFAP) staining. Retinal vascular densities were determined using trypsin digestion preparation of wholemount retinas at 3- and 6-weeks of age. Retinal neovascularization was assessed during the oxygen-induced ischemic retinopathy (OIR). Choroidal neovascularization (CNV) was assessed using laser-induced CNV. Results: Using the TSP2-GFP reporter mice, we observed significant expression of TSP2 mRNA in retinas of postnatal day 5 (P5) mice, which increased by P7 and remained high up to P42. Similar results were observed in retinal wholemount preparations, and western blotting for GFP with the highest level of GFP was observed at P21. In contrast to high level of mRNA at P42, the GFP fluorescence or protein level was dramatically downregulated. The primary retinal vasculature developed at a faster rate in TSP2−/− mice compared with TSP2+/+ mice up to P5. However, the developing retinal vasculature in TSP2+/+ mice caught up with that of TSP2−/− mice after P7. No significant differences in retinal vascular density were observed at 3- or 6-weeks of age. TSP2−/− mice also exhibited a similar sensitivity to the hyperoxia-mediated vessel obliteration and similar level of neovascularization during OIR as TSP2+/+ mice. Lack of TSP2 expression minimally affected laser-induced CNV compared with TSP2+/+ mice. Conclusions

  3. Three Hundred Patients Treated with Ultrapulsed 980 nm Diode Laser for Skin Disorders.

    PubMed

    Wollina, Uwe

    2016-01-01

    The use of lasers in skin diseases is quite common. In contrast to other laser types, medical literature about 980 nm ultrapulsed diode laser is sparse in dermatology. Herein, we report the use of ultrapulsed diode 980 nm laser in 300 patients with vascular lesions, cysts and pseudocysts, infectious disease, and malignant tumors. This laser is a versatile tool with excellent safety and efficacy in the hands of the experienced user. PMID:27688445

  4. Three Hundred Patients Treated with Ultrapulsed 980 nm Diode Laser for Skin Disorders

    PubMed Central

    Wollina, Uwe

    2016-01-01

    The use of lasers in skin diseases is quite common. In contrast to other laser types, medical literature about 980 nm ultrapulsed diode laser is sparse in dermatology. Herein, we report the use of ultrapulsed diode 980 nm laser in 300 patients with vascular lesions, cysts and pseudocysts, infectious disease, and malignant tumors. This laser is a versatile tool with excellent safety and efficacy in the hands of the experienced user.

  5. Three Hundred Patients Treated with Ultrapulsed 980 nm Diode Laser for Skin Disorders

    PubMed Central

    Wollina, Uwe

    2016-01-01

    The use of lasers in skin diseases is quite common. In contrast to other laser types, medical literature about 980 nm ultrapulsed diode laser is sparse in dermatology. Herein, we report the use of ultrapulsed diode 980 nm laser in 300 patients with vascular lesions, cysts and pseudocysts, infectious disease, and malignant tumors. This laser is a versatile tool with excellent safety and efficacy in the hands of the experienced user. PMID:27688445

  6. Three Hundred Patients Treated with Ultrapulsed 980 nm Diode Laser for Skin Disorders.

    PubMed

    Wollina, Uwe

    2016-01-01

    The use of lasers in skin diseases is quite common. In contrast to other laser types, medical literature about 980 nm ultrapulsed diode laser is sparse in dermatology. Herein, we report the use of ultrapulsed diode 980 nm laser in 300 patients with vascular lesions, cysts and pseudocysts, infectious disease, and malignant tumors. This laser is a versatile tool with excellent safety and efficacy in the hands of the experienced user.

  7. Treatment of vascular malformation of the gastrointestinal tract

    NASA Astrophysics Data System (ADS)

    Waldschmidt, Juergen; Stroedter, L.; Doede, T.; Kischkel, A.

    2000-06-01

    Vascular malformations of the gastrointestinal tract are rare phenomenon. They are generally manifested by upper or lower GI - bleeding and do not resolve spontaneously. Emergency intervention is necessary. This paper reports on 10 cases, treated in the Dept. of Pediatric surgery of the FU Berlin, recorded from 1981 to 1999. We use the Nd:YAG laser 1064 nm, Fibertom 5100, Dornier, Germany, with a 600nm barefiber. Reduction in size of the hemangiomas and stop of the GI-bleeding was achieved in all cases.

  8. Focused ultrasound surgery-induced vascular occlusion in fetal medicine

    NASA Astrophysics Data System (ADS)

    Rivens, Ian H.; Rowland, Ian; Denbow, Mark; Fisk, Nicholas M.; Leach, Martin O.; ter Haar, Gail R.

    1998-04-01

    Aim: This study investigates whether it is possible to occlude blood flow in vivo using high intensity focused ultrasound surgery (FUS). Such an effect could be used in the non-invasive treatment of fetal dysfunctions. Conclusion: Our ability to curtail blood flow using FUS allows the possibility of non-invasively treating feto-fetal transfusion syndrome by occluding the placental shunt vessels responsible for the vascular imbalance in twins sharing a placenta. This would have advantages over currently available interventional treatments (surgery or intrauterine lasers), which have significant related mortality and morbidity.

  9. Overgrowth syndromes with vascular malformations.

    PubMed

    Hagen, Solveig L; Hook, Kristen P

    2016-03-01

    This review provides a clinically-oriented summary of the most commonly encountered overgrowth syndromes associated with vascular malformations. This manuscript will outline morphologic features, clinical evaluation and management of this complex group of patients. Recent genetic advances have aided in classification and help to explain overlapping clinical features in many cases. PMID:27607325

  10. The relationships of vascular plants.

    PubMed Central

    Kenrick, P

    2000-01-01

    Recent phylogenetic research indicates that vascular plants evolved from bryophyte-like ancestors and that this involved extensive modifications to the life cycle. These conclusions are supported by a range of systematic data, including gene sequences, as well as evidence from comparative morphology and the fossil record. Within vascular plants, there is compelling evidence for two major clades, which have been termed lycophytes (clubmosses) and euphyllophytes (seed plants, ferns, horsetails). The implications of recent phylogenetic work are discussed with reference to life cycle evolution and the interpretation of stratigraphic inconsistencies in the early fossil record of land plants. Life cycles are shown to have passed through an isomorphic phase in the early stages of vascular plant evolution. Thus, the gametophyte generation of all living vascular plants is the product of massive morphological reduction. Phylogenetic research corroborates earlier suggestions of a major representational bias in the early fossil record. Mega-fossils document a sequence of appearance of groups that is at odds with that predicted by cladogram topology. It is argued here that the pattern of appearance and diversification of plant megafossils owes more to changing geological conditions than to rapid biological diversification. PMID:10905613

  11. Vascular effects of intermittent hypoxia.

    PubMed

    Kanagy, Nancy L

    2009-01-01

    Obstructive sleep apnea is characterized by repeated upper airway obstruction during sleep and affects between 5% and 20% of the population. Epidemiological studies reveal that sleep apnea and associated intermittent hypoxemia increase the risk for hypertension and vascular disease but the mechanisms underlying these effects are incompletely understood. This review reports the results of rodent models of intermittent hypoxia (IH) and relates them to the observed hemodynamic and vascular consequences of sleep apnea. These animal studies have demonstrated that IH exposure in the absence of any other comorbidity causes hypertension, endothelial dysfunction, and augmented constrictor sensitivity, all due at least in part to increased vascular oxidative stress. Animal studies have used a variety of exposure paradigms to study intermittent hypoxia and these different exposure protocols can cause hypocapnia or hypercapnia-or maintain eucapnia-with accompanying alterations in plasma pH. It appears that these different profiles of arterial blood gases can lead to divergent results but the impact of these differences is still being investigated. Overall, the studies in rodents have clearly demonstrated that the vascular and hemodynamic impact of intermittent hypoxia provides a strong rationale for treating clinical sleep apnea to prevent the resulting cardiovascular morbidity and mortality.

  12. Nilotinib-associated vascular events.

    PubMed

    Quintás-Cardama, Alfonso; Kantarjian, Hagop; Cortes, Jorge

    2012-10-01

    Anecdotal evidence suggests that nilotinib therapy may be associated with severe peripheral artery occlusive disease (PAOD). The authors describe the experience at M.D. Anderson Cancer Center regarding vascular events associated with nilotinib therapy in patients with chronic myeloid leukemia. Overall, 5 cases of PAOD were identified among 233 patients, for an incidence of 2%. Nilotinib is a highly selective inhibitor of the inactive conformation of ABL1 kinase. An improved topologic fit to the ABL1 protein-binding surface contributes to its increased potency over imatinib. This higher selectivity in vitro translated to an improved tolerability in vivo. In fact, nilotinib therapy in the frontline phase III ENESTnd (Evaluating Nilotinib Efficacy and Safety in Clinical Trials-Newly Diagnosed Patients) study was associated with an improved toxicity profile compared with that of imatinib. Intriguingly, several cases of severe peripheral artery occlusive disease (PAOD) have been reported among patients treated with nilotinib in small series. We have identified 5 patients with chronic myeloid leukemia (CML) in whom vascular events developed that were likely related to nilotinib therapy among 233 (2%) patients treated at our institution: 1 patient had recurrent Raynaud syndrome, a second patient had recurrent cerebrovascular accidents, and 3 other patients had PAOD (2 of them with other vascular events, including coronary artery disease and pulmonary emboli, respectively). Risk factors for vascular disease were present in only 1 patient with a history of diabetes mellitus. Although the incidence of vascular events is low, this potential complication should be taken into account when selecting nilotinib for the treatment of CML.

  13. Lasers and laser-like devices: part one.

    PubMed

    Stewart, Nicholas; Lim, Adrian C; Lowe, Patricia M; Goodman, Greg

    2013-08-01

    Lasers have been used in dermatology for nearly 50 years. Through their selective targeting of skin chromophores they have become the preferred treatment for many skin conditions, including vascular malformations, photorejuvenation and acne scars. The technology and design of lasers continue to evolve, allowing greater control of laser parameters and resulting in increased safety and efficacy for patients. Innovations have allowed the range of conditions and the skin types amenable to treatment, in both general and cosmetic dermatology, to expand over the last decade. Integrated skin cooling and laser beam fractionation, for example, have improved safety, patient tolerance and decreased downtime. Furthermore, the availability and affordability of quality devices continues to increase, allowing clinicians not only to access laser therapies more readily but also to develop their personal experience in this field. As a result, most Australian dermatologists now have access to laser therapies, either in their own practice or within referable proximity, and practical knowledge of these technologies is increasingly required and expected by patients. Non-laser energy devices utilising intense pulsed light, plasma, radiofrequency, ultrasound and cryolipolysis contribute to the modern laser practitioners' armamentarium and will also be discussed.

  14. History of vascular access for haemodialysis.

    PubMed

    Konner, Klaus

    2005-12-01

    The history of vascular access is a history of vascular surgery as well as a history of dialysis therapy. This survey is a personal view on the history of vascular access without the ambition to cover every detail, but with an effort to mention the major steps in a fascinating panorama.

  15. Motoneurons are essential for vascular pathfinding

    PubMed Central

    Lim, Amy H.; Suli, Arminda; Yaniv, Karina; Weinstein, Brant; Li, Dean Y.; Chien, Chi-Bin

    2011-01-01

    The neural and vascular systems share common guidance cues that have direct and independent signaling effects on nerves and endothelial cells. Here, we show that zebrafish Netrin 1a directs Dcc-mediated axon guidance of motoneurons and that this neural guidance function is essential for lymphangiogenesis. Specifically, Netrin 1a secreted by the muscle pioneers at the horizontal myoseptum (HMS) is required for the sprouting of dcc-expressing rostral primary motoneuron (RoP) axons and neighboring axons along the HMS, adjacent to the future trajectory of the parachordal chain (PAC). These axons are required for the formation of the PAC and, subsequently, the thoracic duct. The failure to form the PAC in netrin 1a or dcc morphants is phenocopied by laser ablation of motoneurons and is rescued both by cellular transplants and overexpression of dcc mRNA. These results provide a definitive example of the requirement of axons in endothelial guidance leading to the parallel patterning of nerves and vessels in vivo. PMID:21828101

  16. Intraoperative fluorescence vascular angiography: during tibial bypass.

    PubMed

    Perry, Diana; Bharara, Manish; Armstrong, David G; Mills, Joseph

    2012-01-01

    Preventing amputations in persons with lower extremity complications of diabetes is a complex endeavor, particularly in those with concomitant ischemia and tissue loss. Fluorescence angiography (Novadaq SPY system) may provide a tool for objective evaluations of tissue viability in the diabetic foot, which is an important indicator of the ability of the diabetic ulcer to heal adequately. The SPY system uses a low-power laser coupled with a charge-coupled device camera and indocyanine green (ICG) to sequence perfusion at the surface of the skin. We present an illustrated example of the potential utility of ICG fluorescence angiography (ICGFA) before and after vascular intervention in a high-risk limb. ICGFA appeared to reveal demarcation between viable and nonviable tissue and real-time perfusion, specifically capillary fill. ICGFA clarified the extent of necessary debridement and provided an immediate indication of improvement in regional perfusion status following revascularization. Future studies involving ICGFA may include pre- and postdebridement and closure perfusion, comparison of tissue perfusion pre- and post-endovascular therapy, and lower extremity flap viability. Future works will also address the consistency of results with ICGFA by analyzing a larger cohort of patients being treated by our unit. PMID:22401340

  17. Vascular diseases: aortitis, aortic aneurysms, and vascular calcification.

    PubMed

    Ladich, Elena; Yahagi, Kazuyuki; Romero, Maria E; Virmani, Renu

    2016-01-01

    Inflammatory diseases of the aorta broadly include noninfectious and infectious aortitis, periaortitis, atherosclerosis, and inflammatory atherosclerotic aneurysms. Aortitis is uncommon but is increasingly recognized as an important cause of aortic aneurysms and dissections. Abdominal (AAA) and thoracic aortic aneurysms (TAA) have different pathologies and etiologies. AAAs are the most common type of aortic aneurysm, and the vast majority of these are atherosclerotic. The causes of TAA vary depending on the site of involvement, but medial degeneration is a common pathologic substrate, regardless of etiology, and genetic influences play a prominent role in TAA expression. Standardized classification schemes for inflammatory and degenerative diseases of the aorta have only recently been added to the pathology literature. A brief overview of the new histopathologic classifications for aortic inflammatory and degenerative diseases has recently been published by the Society for Cardiovascular Pathology and the Association for European Cardiovascular Pathology as a consensus document on the surgical pathology of the aorta. Vascular calcification is a highly regulated biologic process, and the mechanisms leading to vascular calcification are under investigation. Calcification may occur in the intima (atherosclerotic) or in the media secondary to metabolic disease. Rarely, vascular calcification may be associated with genetic disorders. PMID:27526100

  18. Akt isoforms in vascular disease.

    PubMed

    Yu, Haixiang; Littlewood, Trevor; Bennett, Martin

    2015-08-01

    The mammalian serine/threonine Akt kinases comprise three closely related isoforms: Akt1, Akt2 and Akt3. Akt activation has been implicated in both normal and disease processes, including in development and metabolism, as well as cancer and cardiovascular disease. Although Akt signalling has been identified as a promising therapeutic target in cancer, its role in cardiovascular disease is less clear. Importantly, accumulating evidence suggests that the three Akt isoforms exhibit distinct tissue expression profiles, localise to different subcellular compartments, and have unique modes of activation. Consistent with in vitro findings, genetic studies in mice show distinct effects of individual Akt isoforms on the pathophysiology of cardiovascular disease. This review summarises recent studies of individual Akt isoforms in atherosclerosis, vascular remodelling and aneurysm formation, to provide a comprehensive overview of Akt function in vascular disease.

  19. Akt isoforms in vascular disease

    PubMed Central

    Yu, Haixiang; Littlewood, Trevor; Bennett, Martin

    2015-01-01

    The mammalian serine/threonine Akt kinases comprise three closely related isoforms: Akt1, Akt2 and Akt3. Akt activation has been implicated in both normal and disease processes, including in development and metabolism, as well as cancer and cardiovascular disease. Although Akt signalling has been identified as a promising therapeutic target in cancer, its role in cardiovascular disease is less clear. Importantly, accumulating evidence suggests that the three Akt isoforms exhibit distinct tissue expression profiles, localise to different subcellular compartments, and have unique modes of activation. Consistent with in vitro findings, genetic studies in mice show distinct effects of individual Akt isoforms on the pathophysiology of cardiovascular disease. This review summarises recent studies of individual Akt isoforms in atherosclerosis, vascular remodelling and aneurysm formation, to provide a comprehensive overview of Akt function in vascular disease. PMID:25929188

  20. Locally vascularized pelvic accessory spleen.

    PubMed

    Iorio, F; Frantellizzi, V; Drudi, Francesco M; Maghella, F; Liberatore, M

    2016-01-01

    Polysplenism and accessory spleen are congenital, usually asymptomatic anomalies. A rare case of polysplenism with ectopic spleen in pelvis of a 67-year-old, Caucasian female is reported here. A transvaginal ultrasound found a soft well-defined homogeneous and vascularized mass in the left pelvis. Patient underwent MRI evaluation and contrast-CT abdominal scan: images with parenchymal aspect, similar to spleen were obtained. Abdominal scintigraphy with 99mTc-albumin nanocolloid was performed and pelvic region was studied with planar scans and SPECT. The results showed the presence of an uptake area of the radiopharmaceutical in the pelvis, while the spleen was normally visualized. These findings confirmed the presence of an accessory spleen with an artery originated from the aorta and a vein that joined with the superior mesenteric vein. To our knowledge, in the literature, there is just only one case of a true ectopic, locally vascularized spleen in the pelvis.

  1. Vascular Pathophysiology in Hearing Disorders

    PubMed Central

    Trune, Dennis R.; Nguyen-Huynh, Anh

    2014-01-01

    The inner ear vasculature is responsible for maintenance of the blood-labyrinth barrier, transport of systemic hormones for ion homeostasis, and supplying nutrients for metabolic functions. Unfortunately, these blood vessels also expose the ear to circulating inflammatory factors resulting from systemic diseases. Thus, while the inner ear blood vessels are critical for normal function, they also are facilitating pathologic mechanisms that result in hearing and vestibular dysfunction. In spite of these numerous critical roles of inner ear vasculature, little is known of its normal homeostatic functions and how these are compromised in disease. The objective of this review is to discuss the current concepts of vascular biology, how blood vessels naturally respond to circulating inflammatory factors, and how such mechanisms of vascular pathophysiology may cause hearing loss. PMID:25346568

  2. Metoclopramide and renal vascular resistance.

    PubMed

    Manara, A R; Bolsin, S; Monk, C R; Hartnell, G; Harris, R A

    1991-01-01

    We have studied the effect of i.v. metoclopramide on renal vascular resistance in nine healthy volunteers. Peak systolic and end-diastolic frequencies were measured using duplex Doppler ultrasound of a renal interlobar artery, before and after the administration of i.v. metoclopramide 10 mg, and the resistance index derived. There was no significant change in mean arterial pressure or resistance index following metoclopramide.

  3. Vascular compression of the duodenum.

    PubMed Central

    Moskovich, R; Cheong-Leen, P

    1986-01-01

    Compression of the third or fourth part of the duodenum by the superior mesenteric artery or one of its branches is the anatomic basis for some cases of duodenal obstruction. Two cases of vascular obstruction of the duodenum after surgical correction of scoliosis are presented. The embryologic and pathoanatomic bases for this condition, and the rationale for treatment, are described. Images Figure 1. Figure 2. Figure 3. PMID:3761291

  4. Metoclopramide and renal vascular resistance.

    PubMed

    Manara, A R; Bolsin, S; Monk, C R; Hartnell, G; Harris, R A

    1991-01-01

    We have studied the effect of i.v. metoclopramide on renal vascular resistance in nine healthy volunteers. Peak systolic and end-diastolic frequencies were measured using duplex Doppler ultrasound of a renal interlobar artery, before and after the administration of i.v. metoclopramide 10 mg, and the resistance index derived. There was no significant change in mean arterial pressure or resistance index following metoclopramide. PMID:1997046

  5. [Jaboulay, vascular surgeon at Lyon].

    PubMed

    Bouchet, Mathieu

    2010-01-01

    Mathieu Jaboulay was an excellent surgeon. He was the first to come up with the principles of vascular surgery as he made a surgical anastomosis between two arteries of a dog by an eversion circular suture. In 1902, he implemented a suture between an artery and a vein in an arteritis by obliteration. Jaboulay also tried the graft of an animal kidney on the crease of a human elbow in 1906. Jaboulay was an initiator for Alexis Carrel. PMID:20527332

  6. The European experience with vascular injuries.

    PubMed

    Fingerhut, Abe; Leppäniemi, Ari K; Androulakis, George A; Archodovassilis, F; Bouillon, Bertil; Cavina, Enrico; Chaloner, Eddie; Chiarugi, Massimo; Davidovic, Lazar; Delgado-Millan, Miguel Angel; Goris, Jan; Gunnlaugsson, Gunnar H; Jover, Jose Maria; Konstandoulakis, Manoussos M; Kurtoglu, Mehmet; Lepäntalo, Mauri; Llort-Pont, Carme; Meneu-Diaz, Juan Carlos; Moreno-Gonzales, Enrique; Navarro-Soto, Salvador; Panoussis, P; Ryan, James M; Salenius, Juha P; Seccia, Massimo; Takolander, Rabbe; Taviloglu, Korhan; Tiesenhausen, Kurt; Torfason, Bjarni; Uranüs, Selman

    2002-02-01

    The rich and diverse heritage of the management of vascular injuries in the 45 independent European countries prevents the authors from revealing a uniform picture of the European experience, but some trends are clearly emerging. In countries with a low incidence of penetrating trauma and increasing use of interventional vascular procedures, the proportion of iatrogenic vascular trauma exceeds 40% of all vascular injuries, whereas on other parts of the continent, armed conflicts are still a major cause of vascular trauma. National vascular registries, mostly in the Scandinavian countries, produce useful, nationwide data about vascular trauma and its management but suffer still from inadequate data collection. Despite a relatively low incidence of vascular trauma in most European countries, the results are satisfactory, probably in most cases because of active and early management by surgeons on call, whether with vascular training or not, treating all kinds of vascular surgical emergencies. In some countries, attempts at developing a trauma and emergency surgical specialty, including expertise in the management of vascular injuries, are on their way. PMID:11905944

  7. The European experience with vascular injuries.

    PubMed

    Fingerhut, Abe; Leppäniemi, Ari K; Androulakis, George A; Archodovassilis, F; Bouillon, Bertil; Cavina, Enrico; Chaloner, Eddie; Chiarugi, Massimo; Davidovic, Lazar; Delgado-Millan, Miguel Angel; Goris, Jan; Gunnlaugsson, Gunnar H; Jover, Jose Maria; Konstandoulakis, Manoussos M; Kurtoglu, Mehmet; Lepäntalo, Mauri; Llort-Pont, Carme; Meneu-Diaz, Juan Carlos; Moreno-Gonzales, Enrique; Navarro-Soto, Salvador; Panoussis, P; Ryan, James M; Salenius, Juha P; Seccia, Massimo; Takolander, Rabbe; Taviloglu, Korhan; Tiesenhausen, Kurt; Torfason, Bjarni; Uranüs, Selman

    2002-02-01

    The rich and diverse heritage of the management of vascular injuries in the 45 independent European countries prevents the authors from revealing a uniform picture of the European experience, but some trends are clearly emerging. In countries with a low incidence of penetrating trauma and increasing use of interventional vascular procedures, the proportion of iatrogenic vascular trauma exceeds 40% of all vascular injuries, whereas on other parts of the continent, armed conflicts are still a major cause of vascular trauma. National vascular registries, mostly in the Scandinavian countries, produce useful, nationwide data about vascular trauma and its management but suffer still from inadequate data collection. Despite a relatively low incidence of vascular trauma in most European countries, the results are satisfactory, probably in most cases because of active and early management by surgeons on call, whether with vascular training or not, treating all kinds of vascular surgical emergencies. In some countries, attempts at developing a trauma and emergency surgical specialty, including expertise in the management of vascular injuries, are on their way.

  8. Mechanosensing at the Vascular Interface

    PubMed Central

    Tarbell, John M.; Simon, Scott I.; Curry, Fitz-Roy E.

    2015-01-01

    Mammals are endowed with a complex set of mechanisms that sense mechanical forces imparted by blood flow to endothelial cells (ECs), smooth muscle cells, and circulating blood cells to elicit biochemical responses through a process referred to as mechanotransduction. These biochemical responses are critical for a host of other responses, including regulation of blood pressure, control of vascular permeability for maintaining adequate perfusion of tissues, and control of leukocyte recruitment during immunosurveillance and inflammation. This review focuses on the role of the endothelial surface proteoglycan/glycoprotein layer—the glycocalyx (GCX)—that lines all blood vessel walls and is an agent in mechanotransduction and the modulation of blood cell interactions with the EC surface. We first discuss the biochemical composition and ultrastructure of the GCX, highlighting recent developments that reveal gaps in our understanding of the relationship between composition and spatial organization. We then consider the roles of the GCX in mechanotransduction and in vascular permeability control and review the prominent interaction of plasma borne sphingosine-1 phosphate (S1P), which has been shown to regulate both the composition of the GCX and the endothelial junctions. Finally, we consider the association of GCX degradation with inflammation and vascular disease and end with a final section on future research directions. PMID:24905872

  9. [Vascular Ehlers-Danlos syndrome].

    PubMed

    Frank, Michael

    2009-04-20

    Vascular type Ehlers-Danlos syndrome (EDS) is a rare inherited disease with an autosomal dominant trait. The mutation of the COL3A1 gene which encodes type III collagen, is responsible of early vascular (spontaneous arterial rupture or dissection), digestive (perforation) and obstetrical events (uterine and arterial rupture). Diagnosis of the disease is primarily clinical, especially in case of characteristic morphologic features. Diagnostic certainty is obtained by evidencing the mutation of the COL3A1 gene. Some arterial lesions are suggestive of the disease, as dissecting aneurysms of the internal carotid, of the iliac arteries, and of the anterior visceral aortic branches, fusiform aneurisms of the splenic artery, and the occurrence of a non traumatic direct carotid-cavernous fistula. The occurrence of a spontaneous peritonitis or of an extensive perineal tear after delivery should also draw physician's attention. Because of the unpredictability of arterial or organ rupture, any patient diagnosed with vascular type EDS presenting with an acute pain syndrome should be considered as a trauma situation and be investigated straightaway by CT-scan or MRI testing, in order to eliminate a life threatening complication. PMID:19462862

  10. New Approaches for Vascular Anastomoses.

    PubMed

    Bonilla, Luis F.; Sullivan, Daniel J.

    2001-02-01

    Increasing interest in minimally invasive cardiac surgical procedures has generated a renewed interest in facilitated methods to create vascular anastomosis. These devices, in order to be viable, must perform equally or better than sutures. An extensive review of the literature was performed on the different ways to create a mechanical vascular anastomosis. The experience of the authors with the development of a family of connectors (The Symmetry Bypass System; St. Jude Medical, Minneapolis, MN) is presented. The Aortic Connector System (St. Jude Medical) has undergone extensive animal testing with 30-, 90-, and 180-day follow-up. Over 250 human implants have been performed successfully. Preliminary animal studies with the stainless steel coronary connector are also presented. The creation of a facilitated mechanical vascular anastomosis should not compromise quality or patency rates, and produce at least the same results as those obtained with standard suturing techniques. These devices dramatically reduce the time to perform an anastomosis and eliminate the need for aortic clamping and cardiopulmonary bypass. These advantages should result in a significant reduction in surgical complications, and may facilitate a move to true endoscopic and/or percutaneous bypass. PMID:11177719

  11. [Vascular dementia: facts and controversies].

    PubMed

    Pavlović, Aleksandra; Pavlović, Dragan; Aleksić, Vuk; Sternić, Nadezda

    2013-01-01

    Vascular dementia (VaD) is the second most frequent dementia after Alzheimer's disease, and is diagnosed during lifetime in 20% of demented patients. Five-year survival rate in VaD is 39%, while it is estimated to be 75% in healthy persons of the same age. It is therefore important to make correct diagnosis of VaD early in the course of the disease. Risk factors forVaD are identical to stroke risk factors, and there are significant possibilities for the prevention of vascular cognitive decline. Cognitive decline develops acutely or step-by-step within three months after stroke, but more gradual progression of intellectual decline is also possible. Neurological examination can reveal pyramidal and extrapyramidal signs, pseudobulbar palsy, gait disturbance and urinary incontinence. Neuropsychological profile comprises the loss of cognitive set shifting, decline in word fluency, verbal learning difficulties, perseverations, difficulties in complex figure copying, and in patients with cortically located lesions also problems with speech and praxia. The basis of the diagnosis is, besides history, neurological examination and neuropsychological assessment, computed tomography and/ or magnetic resonance brain imaging. Vascular risk factors control is the most important measure in VaD prevention. Modern guidelines for the treatment of cognitive decline in VaD emphasize that donepezil can be useful in the improvement of cognitive status at the level of Class Ila recommendation at the level of evidence A, while memantine may be useful in patients with mixed VaD and Alzheimer's disease dementia. PMID:23745353

  12. Pregnancy and Vascular Liver Disease

    PubMed Central

    Bissonnette, Julien; Durand, François; de Raucourt, Emmanuelle; Ceccaldi, Pierre-François; Plessier, Aurélie; Valla, Dominique; Rautou, Pierre-Emmanuel

    2015-01-01

    Vascular disorders of the liver frequently affect women of childbearing age. Pregnancy and the postpartum are prothrombotic states. Pregnancy seems to be a trigger for Budd–Chiari syndrome in patients with an underlying prothrombotic disorder. Whether pregnancy is a risk factor for other vascular liver disorders is unknown. In women with a known vascular liver disorder and a desire for pregnancy, stabilisation of the liver disease, including the use of a portal decompressive procedure when indicated, should be reached prior to conception. The presence of esophageal varices should be screened and adequate prophylaxis of bleeding applied in a manner similar to what is recommended for patients with cirrhosis. Most women likely benefit from anticoagulation during pregnancy and the postpartum. Labor and delivery are best managed by a multidisciplinary team with experience in this situation. Assisted vaginal delivery is the preferred mode of delivery. Although the risk of miscarriage and premature birth is heightened, current management of these diseases makes it very likely to see the birth of a live baby when pregnancy reaches 20 weeks of gestation. PMID:25941432

  13. CLINICAL AND MICRODISSECTION GENOTYPING ANALYSES OF THE EFFECT OF INTRA-ARTERIAL CYTOREDUCTIVE CHEMOTHERAPY IN THE TREATMENT OF LACRIMAL GLAND ADENOID CYSTIC CARCINOMA

    PubMed Central

    Tse, David T

    2005-01-01

    Purpose To determine the effect of intra-arterial cytoreductive chemotherapy (IACC) as an adjunct of a multimodality protocol for the treatment of lacrimal gland adenoid cystic carcinoma (ACC). Methods This was a retrospective, comparative, consecutive case series. Nine consecutive patients with lacrimal gland ACC were treated with IACC, followed by orbital exenteration and chemoradiotherapy. This case series was compared with a series of seven patients treated by conventional local therapies. Clinical records, imaging studies, histologic sections, and archival specimens from all 16 patients were reviewed. Information analyzed included site of disease, histologic characteristics, extent of disease, local-regional recurrence or distant metastases, and disease-free survival time. Gene analysis was performed on microdissected tissue samples. Mutational allelotyping targeting nine genomic loci using 15 polymorphic microsatellite markers situated in proximity to known tumor suppressor genes serve as markers for the presence of gene deletion. The effect of IACC was assessed by the radiographic response and survival outcome in comparison to a historical cohort of patients managed by conventional local therapies. A fractional mutation index was used to compare the acquired mutational load between different tumors having nonidentical patterns of microsatellite informativeness. Results The carcinoma cause-specific death rates between the two treatment groups was significant (P = .029, log-rank test). The cumulative 5-year carcinoma cause-specific death rate was 16.7% in the IACC-treated group compared with 57.1% in the conventional treatment group. 1p36 was the single most common site affected by allelic loss for microsatellite markers in this series. Conclusions The preliminary data suggest that IACC as an integral component of a multimodal treatment strategy is potentially effective in improving local disease control and overall disease-free survival in lacrimal gland ACC

  14. PCR walking from microdissection clone M54 identifies three exons from the human gene for the neural cell adhesion molecule L1 (CAM-L1).

    PubMed Central

    Rosenthal, A; MacKinnon, R N; Jones, D S

    1991-01-01

    Microdissection has proved to be a powerful tool in the construction of libraries from specific chromosome segments (11) which are poorly covered by existing RFLP markers. Microclones also represent starting points for finding genes of interest. However, their length (100 to 200 bp) can make their use as probes problematic and identifying them as coding sequence is difficult. We report here that microclones can be extended in vitro by a modified version of our original PCR walking method (10) which utilises oligo-cassettes and the solid phase biotin/streptavidin separation system. We have extended the microclone M54, derived by dissection from Xq27.2 to proximal Xq28 (12), in both directions for approximately 700 bp. Direct sequencing of these products revealed that M54 was located within an intron of the human gene encoding the neural cell adhesion molecule L1 (CAM-L1) which has been recently mapped to Xq28 (13). The extension of M54 also identified three exons of this gene. This information allowed subsequent amplification of a 2.4 kb cDNA molecule from fetal human brain mRNA which encodes most of human CAM-L1. Sequencing of this cDNA revealed a high degree of sequence conservation with the mouse homologue (14). This is the first description of extension of a human derived microclone by PCR mediated walking within total human genomic DNA. These results show that anonymous DNA sequences may be extended into coding or any sequence. Images PMID:1923824

  15. New Medical Applications Of Metal Vapor Lasers

    NASA Astrophysics Data System (ADS)

    Anderson, Robert S.; McIntosh, Alexander I.

    1989-06-01

    The first medical application for metal vapor lasers has been granted marketing approval by the FDA. This represents a major milestone for this technology. Metalaser Technologies recently received this approval for its Vasculase unit in the treatment of vascular lesions such as port wine stains, facial telangiectasia and strawberry hemangiomas.

  16. Krypton red laser photocoagulation of the ocular fundus. 1982.

    PubMed

    Yannuzzi, Lawrence A; Shakin, Jeffrey L

    2012-02-01

    The theoretical rationale, the histopathologic evidence, and the preliminary clinical studies related to krypton red laser (KRL) photocoagulation of the ocular fundus are reviewed. The authors report on their experience with currently available laser systems using this wavelength (647.1 nm) for photocoagulation of retinal vascular proliferative diseases and chorioretinal diseases associated with exudative manifestations. A histopathologic and clinical comparison of argon blue-green laser (ABGL), the pure argon green laser (AGL), and the krypton yellow laser (KYL), with reference to photocoagulation treatment of the ocular fundus is also discussed.

  17. Breaking dogmas: the plant vascular pathogen Xanthomonas albilineans is able to invade non-vascular tissues despite its reduced genome.

    PubMed

    Mensi, Imène; Vernerey, Marie-Stéphanie; Gargani, Daniel; Nicole, Michel; Rott, Philippe

    2014-02-12

    Xanthomonas albilineans, the causal agent of sugarcane leaf scald, is missing the Hrp type III secretion system that is used by many Gram-negative bacteria to colonize their host. Until now, this pathogen was considered as strictly limited to the xylem of sugarcane. We used confocal laser scanning microscopy, immunocytochemistry and transmission electron microscopy (TEM) to investigate the localization of X. albilineans in diseased sugarcane. Sugarcane plants were inoculated with strains of the pathogen labelled with a green fluorescent protein. Confocal microscopy observations of symptomatic leaves confirmed the presence of the pathogen in the protoxylem and metaxylem; however, X. albilineans was also observed in phloem, parenchyma and bulliform cells of the infected leaves. Similarly, vascular bundles of infected sugarcane stalks were invaded by X. albilineans. Surprisingly, the pathogen was also observed in apparently intact storage cells of the stalk and in intercellular spaces between these cells. Most of these observations made by confocal microscopy were confirmed by TEM. The pathogen exits the xylem following cell wall and middle lamellae degradation, thus creating openings to reach parenchyma cells. This is the first description of a plant pathogenic vascular bacterium invading apparently intact non-vascular plant tissues and multiplying in parenchyma cells.

  18. Breaking dogmas: the plant vascular pathogen Xanthomonas albilineans is able to invade non-vascular tissues despite its reduced genome

    PubMed Central

    Mensi, Imène; Vernerey, Marie-Stéphanie; Gargani, Daniel; Nicole, Michel; Rott, Philippe

    2014-01-01

    Xanthomonas albilineans, the causal agent of sugarcane leaf scald, is missing the Hrp type III secretion system that is used by many Gram-negative bacteria to colonize their host. Until now, this pathogen was considered as strictly limited to the xylem of sugarcane. We used confocal laser scanning microscopy, immunocytochemistry and transmission electron microscopy (TEM) to investigate the localization of X. albilineans in diseased sugarcane. Sugarcane plants were inoculated with strains of the pathogen labelled with a green fluorescent protein. Confocal microscopy observations of symptomatic leaves confirmed the presence of the pathogen in the protoxylem and metaxylem; however, X. albilineans was also observed in phloem, parenchyma and bulliform cells of the infected leaves. Similarly, vascular bundles of infected sugarcane stalks were invaded by X. albilineans. Surprisingly, the pathogen was also observed in apparently intact storage cells of the stalk and in intercellular spaces between these cells. Most of these observations made by confocal microscopy were confirmed by TEM. The pathogen exits the xylem following cell wall and middle lamellae degradation, thus creating openings to reach parenchyma cells. This is the first description of a plant pathogenic vascular bacterium invading apparently intact non-vascular plant tissues and multiplying in parenchyma cells. PMID:24522883

  19. Experimental Branch Retinal Vein Occlusion Induces Upstream Pericyte Loss and Vascular Destabilization

    PubMed Central

    Dominguez, Elisa; Raoul, William; Calippe, Bertrand; Sahel, José-Alain; Guillonneau, Xavier; Paques, Michel; Sennlaub, Florian

    2015-01-01

    Aims Branch retinal vein occlusion (BRVO) leads to extensive vascular remodeling and is important cause of visual impairment. Although the vascular morphological changes following experimental vein occlusion have been described in a variety of models using angiography, the underlying cellular events are ill defined. Methods and Results We here show that laser-induced experimental BRVO in mice leads to a wave of TUNEL-positive endothelial cell (EC) apoptosis in the upstream vascular network associated with a transient edema and hemorrhages. Subsequently, we observe an induction of EC proliferation within the dilated vein and capillaries, detected by EdU incorporation, and the edema resolves. However, the pericytes of the upstream capillaries are severely reduced, which was associated with continuing EC apoptosis and proliferation. The vascular remodeling was associated with increased expression of TGFβ, TSP-1, but also FGF2 expression. Exposure of the experimental animals to hypoxia, when pericyte (PC) dropout had occurred, led to a dramatic increase in endothelial cell proliferation, confirming the vascular instability induced by the experimental BRVO. Conclusion Experimental BRVO leads to acute endothelial cells apoptosis and increased permeability. Subsequently the upstream vascular network remains destabilized, characterized by pericyte dropout, un-physiologically high endothelial cells turnover and sensitivity to hypoxia. These early changes might pave the way for capillary loss and subsequent chronic ischemia and edema that characterize the late stage disease. PMID:26208283

  20. Purinergic control of vascular tone in the retina.

    PubMed

    Kur, Joanna; Newman, Eric A

    2014-02-01

    Purinergic control of vascular tone in the CNS has been largely unexplored. This study examines the contribution of endogenous extracellular ATP, acting on vascular smooth muscle cells, in controlling vascular tone in the in vivo rat retina. Retinal vessels were labelled by i.v. injection of a fluorescent dye and imaged with scanning laser confocal microscopy. The diameters of primary arterioles were monitored under control conditions and following intravitreal injection of pharmacological agents. Apyrase (500 units ml(-1)), an ATP hydrolysing enzyme, dilated retinal arterioles by 40.4 ± 2.8%, while AOPCP (12.5 mm), an ecto-5'-nucleotidase inhibitor that increases extracellular ATP levels, constricted arterioles by 58.0 ± 3.8% (P < 0.001 for both), demonstrating the importance of ATP in the control of basal vascular tone. Suramin (500 μm), a broad-spectrum P2 receptor antagonist, dilated retinal arterioles by 50.9 ± 3.7% (P < 0.001). IsoPPADS (300 μm) and TNP-ATP (50 μm), more selective P2X antagonists, dilated arterioles by 41.0 ± 5.3% and 55.2 ± 6.1% respectively (P < 0.001 for both). NF023 (50 μm), a potent antagonist of P2X1 receptors, dilated retinal arterioles by 32.1 ± 2.6% (P < 0.001). A438079 (500 μm) and AZ10606120 (50 μm), P2X7 antagonists, had no effect on basal vascular tone (P = 0.99 and P = 1.00 respectively). In the ex vivo retina, the P2X1 receptor agonist α,β-methylene ATP (300 nm) evoked sustained vasoconstrictions of 18.7 ± 3.2% (P < 0.05). In vivo vitreal injection of the gliotoxin fluorocitrate (150 μm) dilated retinal vessels by 52.3 ± 1.1% (P < 0.001) and inhibited the vasodilatory response to NF023 (50 μm, 7.9 ± 2.0%; P < 0.01). These findings suggest that vascular tone in rat retinal arterioles is maintained by tonic release of ATP from the retina. ATP acts on P2X1 receptors, although contributions from other P2X and P2Y receptors cannot be ruled out. Retinal glial cells are a possible source of the vasoconstricting ATP.

  1. Cell-based strategies for vascular regeneration.

    PubMed

    Zou, Tongqiang; Fan, Jiabing; Fartash, Armita; Liu, Haifeng; Fan, Yubo

    2016-05-01

    Vascular regeneration is known to play an essential role in the repair of injured tissues mainly through accelerating the repair of vascular injury caused by vascular diseases, as well as the recovery of ischemic tissues. However, the clinical vascular regeneration is still challenging. Cell-based therapy is thought to be a promising strategy for vascular regeneration, since various cells have been identified to exert important influences on the process of vascular regeneration such as the enhanced endothelium formation on the surface of vascular grafts, and the induction of vessel-like network formation in the ischemic tissues. Here are a vast number of diverse cell-based strategies that have been extensively studied in vascular regeneration. These strategies can be further classified into three main categories, including cell transplantation, construction of tissue-engineered grafts, and surface modification of scaffolds. Cells used in these strategies mainly refer to terminally differentiated vascular cells, pluripotent stem cells, multipotent stem cells, and unipotent stem cells. The aim of this review is to summarize the reported research advances on the application of various cells for vascular regeneration, yielding insights into future clinical treatment for injured tissue/organ. PMID:26864677

  2. Vascular thalamic amnesia: a reappraisal.

    PubMed

    Carlesimo, Giovanni Augusto; Lombardi, Maria Giovanna; Caltagirone, Carlo

    2011-04-01

    In humans lacunar infarcts in the mesial and anterior regions of the thalami are frequently associated with amnesic syndromes. In this review paper, we scrutinized 41 papers published between 1983 and 2009 that provided data on a total of 83 patients with the critical ischemic lesions (i.e. 17 patients with right-sided lesions, 25 with left-sided lesions and 41 with bilateral lesions). We aimed to find answers to the following questions concerning the vascular thalamic amnesia syndrome: (i) Which qualitative pattern of memory impairment (and associated cognitive and behavioral deficits) do these patients present? (ii) Which lesioned intrathalamic structures are primarily responsible for the amnesic syndrome? (iii) Are the recollection and familiarity components of declarative memory underlain by the same or by different thalamic structures? Results of the review indicate that, similar to patients with amnesic syndromes due to mesio-temporal lobe damage, patients with vascular thalamic amnesia display a prevalent deficit of declarative anterograde long-term memory, a less consistent deficit of declarative retrograde long-term memory and substantially spared short-term and implicit memory. Unlike mesio-temporal lobe patients, however, vascular thalamic amnesics often present dysexecutive and behavioral deficits similar to those observed in patients with frontal damage. The presence of an amnesic syndrome in patients with thalamic lacunar infarcts is strongly predicted by involvement of the mammillo-thalamic tract, which connects the anterior nuclei complex to the hippocampus proper via the fornix and the mammillary bodies. Finally, data reported in a few single cases provide support for the hypothesis that thalamic regions connected to distinct areas of the mesio-temporal lobe play differential roles in recollection and familiarity processes. The mammillo-thalamic tract/anterior nuclei axis seems primarily implicated in recollective processes, whereas the

  3. Major vascular injuries complicating knee arthroscopy

    PubMed Central

    Bancu, Serban; Muresan, Mircea; Sala, Daniela

    2015-01-01

    Starting with a case report, we made a detailed review of the literature, with the purpose of identifying and analyzing the type of iatrogenic vascular lesion following knee arthroscopy and the method of vascular repair. A PubMed literature search was undertaken to locate all reported cases of major vascular iatrogenic injuries during arthroscopic knee procedures. We identified 39 papers which report a total of 62 cases of major iatrogenic popliteal lesions after knee arthroscopy, between 1985 and 2014. The type of arthroscopic intervention performed, the type of iatrogenic vascular lesion encountered, the time passed until its discovery and treatment, the method of vascular reconstruction, and the postoperative course are presented. Postarthroscopy vascular complications are infrequent but potentially disastrous for the condition of the affected inferior limb. An early diagnosis and reintervention are mandatory for a good postoperative outcome. PMID:26240627

  4. DNA Damage and Repair in Vascular Disease.

    PubMed

    Uryga, Anna; Gray, Kelly; Bennett, Martin

    2016-01-01

    DNA damage affecting both genomic and mitochondrial DNA is present in a variety of both inherited and acquired vascular diseases. Multiple cell types show persistent DNA damage and a range of lesions. In turn, DNA damage activates a variety of DNA repair mechanisms, many of which are activated in vascular disease. Such DNA repair mechanisms either stall the cell cycle to allow repair to occur or trigger apoptosis or cell senescence to prevent propagation of damaged DNA. Recent evidence has indicated that DNA damage occurs early, is progressive, and is sufficient to impair function of cells composing the vascular wall. The consequences of persistent genomic and mitochondrial DNA damage, including inflammation, cell senescence, and apoptosis, are present in vascular disease. DNA damage can thus directly cause vascular disease, opening up new possibilities for both prevention and treatment. We review the evidence for and the causes, types, and consequences of DNA damage in vascular disease.

  5. The decline and resurgence of vascular dementia.

    PubMed Central

    Hachinski, V C

    1990-01-01

    Arteriosclerotic narrowing of cerebral arteries was once viewed as the key to mental decline. As Alzheimer's disease gained recognition and the concept of multi-infarct dementia achieved acceptance, vascular dementia came to be regarded as uncommon. The changing nature of cerebral vascular disease, the aging of the population and the widespread use of brain imaging techniques have brought new prominence to vascular dementia, chiefly in the form of an epidemic of "Binswanger's disease". Growing evidence suggests that not only grey matter lesions but also white matter lesions contribute to dementia, that vascular factors commonly coexist and interact with Alzheimer changes and that Alzheimer's disease has a vascular and potentially treatable component. Vascular dementia needs to be redefined, reappraised and reinvestigated. PMID:2403832

  6. MicroRNAs Regulate Vascular Medial Calcification.

    PubMed

    Leopold, Jane A

    2014-01-01

    Vascular calcification is highly prevalent in patients with coronary artery disease and, when present, is associated with major adverse cardiovascular events, including an increased risk of cardiovascular mortality. The pathogenesis of vascular calcification is complex and is now recognized to recapitulate skeletal bone formation. Vascular smooth muscle cells (SMC) play an integral role in this process by undergoing transdifferentiation to osteoblast-like cells, elaborating calcifying matrix vesicles and secreting factors that diminish the activity of osteoclast-like cells with mineral resorbing capacity. Recent advances have identified microRNAs (miRs) as key regulators of this process by directing the complex genetic reprogramming of SMCs and the functional responses of other relevant cell types relevant for vascular calcification. This review will detail SMC and bone biology as it relates to vascular calcification and relate what is known to date regarding the regulatory role of miRs in SMC-mediated vascular calcification.

  7. [Pharmacological treatment of vascular cognitive impairment].

    PubMed

    Bidzan, Leszek

    2006-01-01

    Vascular dementia is second most common cause of dementia. The paper highlights the most important trends in pharmacological treatment of vascular dementia. Result of a clinical trial of some agents appears to be promising. Pentoxifyline appears to be useful in multi-infarct vascular dementia. Nimodipine produced improvement in subcortical dementia. Some other agents like ginkgo biloba, acetylocholinesterase inhibitors, memantine and other also have shown mild benefit or at least were associated with some stabilization of dementia. PMID:16969897

  8. Cryptic vascular malformations involving the brainstem

    SciTech Connect

    Yeates, A.; Enzmann, D.

    1983-01-01

    Six patients with angiographically cryptic vascular malformations involving the brainstem were examined with computed tomography (CT). The clinical and CT findings of cryptic vascular malformations of the brainstem are described and distinguished from those of brainstem glioma and multiple sclerosis. Calcification within a brainstem lesion that displays relatively little mass effect and shows little contrast enhancement, particularly when associated with a long history of waxing and waning brainstem symptoms, should suggest a vascular malformation.

  9. Direct intratumoral embolization of intranasal vascular tumors.

    PubMed

    Jang, Hyun-Uk; Kim, Tae-Hoon; Park, Chang-Mook; Kim, Jung-Soo

    2013-02-01

    Embolization is a well established technique that facilitates the subsequent surgical removal of vascularized tumors such as juvenile angiofibroma. Preoperative transarterial embolization has proven beneficial for decreasing intraoperative blood loss. However, the procedure is often incomplete owing to extensive vascular structure. Direct intratumoral embolization may help overcome this limitation. We report our experience with embolization of nasal vascular tumors by means of direct intratumoral injection of n-butyl cyanoacrylate (NBCA).

  10. Supermarket model for vascular disease care.

    PubMed

    Shah, Dhiraj M; Bruni, Karen; Darling, R Clement

    2002-09-01

    A supermarket model for vascular patient care proposes an interdisciplinary group of health care teams such as vascular nurses, interventional radiologists, vascular surgeons, angiologists, internists, cardiologists, and neurologists and facilities such as diagnostic testing laboratories, subcenters such as wound care and foot care centers, atherosclerotic risk prevention centers, rehabilitation centers, vein centers, and socioeconomic follow-up centers that would provide health care of vascular disease in a comprehensive manner in terms of quality care, convenience for patients, 1-stop shopping, education and training, and research and development.

  11. Vascular anomalies: differential diagnosis and mimickers.

    PubMed

    Garzon, Maria C; Weitz, Nicole; Powell, Julie

    2016-03-01

    Vascular anomalies are very common in children and encompass a wide spectrum of diseases. Many vascular anomalies can be mistaken for infantile hemangioma (IH). In addition, there is a variety of rare disorders including benign and malignant tumors that may mimic IH and other types of vascular anomalies. Understanding the clinical features, natural history, and typical clinical course of different types of vascular anomalies is essential in order to make the correct diagnosis and guide management. Radiologic imaging plays an important role in establishing the diagnosis; and when the diagnosis remains in doubt, a biopsy performed by a surgical specialist with expertise may prove to be lifesaving. PMID:27607326

  12. Vascular nursing in Greece: luxury or necessity?

    PubMed

    Georgakarakos, Efstratios; Bitza, Christina; Papanas, Nikolaos; Matsagkas, Miltiadis; Lazarides, Miltos K

    2013-09-01

    Although peripheral arterial disease is prevalent in the primary care setting, insufficient vascular education among nurses and physicians coupled with certain economic constraints undermines treatment efficacy. Moreover, the burden of advanced venous pathology such as posthrombotic syndrome, venous ulcers, and lymphedema remains suboptimally treated. This article advocates the development of a vascular nursing specialty as a means to improving vascular care especially nowadays, when health care providers dictate comprehensive and cost-effective nursing practice and patient management. It also presents the first attempt to organize a Vascular Nursing Educational Session in Greece. PMID:24043676

  13. Scaffolds in vascular regeneration: current status

    PubMed Central

    Thottappillil, Neelima; Nair, Prabha D

    2015-01-01

    An ideal vascular substitute, especially in <6 mm diameter applications, is a major clinical essentiality in blood vessel replacement surgery. Blood vessels are structurally complex and functionally dynamic tissue, with minimal regeneration potential. These have composite extracellular matrix (ECM) and arrangement. The interplay between ECM components and tissue specific cells gives blood vessels their specialized functional attributes. The core of vascular tissue engineering and regeneration relies on the challenges in creating vascular conduits that match native vessels and adequately regenerate in vivo. Out of numerous vascular regeneration concerns, the relevance of ECM emphasizes much attention toward appropriate choice of scaffold material and further scaffold development strategies. The review is intended to be focused on the various approaches of scaffold materials currently in use in vascular regeneration and current state of the art. Scaffold of choice in vascular tissue engineering ranges from natural to synthetic, decellularized, and even scaffold free approach. The applicability of tubular scaffold for in vivo vascular regeneration is under active investigation. A patent conduit with an ample endothelial luminal layer that can regenerate in vivo remains an unanswered query in the field of small diameter vascular tissue engineering. Besides, scaffolds developed for vascular regeneration, should aim at providing functional substitutes for use in a regenerative approach from the laboratory bench to patient bedside. PMID:25632236

  14. Vascular grafting strategies in coronary intervention

    NASA Astrophysics Data System (ADS)

    Knight, Darryl; Gillies, Elizabeth; Mequanint, Kibret

    2014-06-01

    With the growing need for coronary revascularizations globally, several strategies to restore blood flow to the heart have been explored. Bypassing the atherosclerotic coronary arteries with autologous grafts, synthetic prostheses and tissue-engineered vascular grafts continue to be evaluated in search of a readily available vascular graft with clinically acceptable outcomes. The development of such a vascular graft including tissue engineering approaches both in situ and in vitro is herein reviewed, facilitating a detailed comparison on the role of seeded cells in vascular graft patency.

  15. Vascular tumors and malformations in children, Introduction.

    PubMed

    Maguiness, Sheilagh M

    2016-03-01

    Over the past decade, I have been amazed at the growth in the field of vascular anomalies. The recognition of vascular birthmarks as a defined area of medicine is a relatively recent event. The International Society for the Study of Vascular Anomalies (ISSVA) was founded by Drs John Mulliken and Anthony Young in the late 1970s. Mulliken and Glowacki's sentinel 1982 paper on the biologic classification of vascular anomalies further established the field, by providing clarity of nomenclature and unifying concepts that had previously been lacking. PMID:27607317

  16. Tumor vascular disruption using various radiation types

    PubMed Central

    2014-01-01

    The feasibility of disrupting a tumor’s vascular structure with various radiation types and radionuclides is investigated. Calculated absorbed dose profiles for photons and 4He ions suggest that low-energy beta-gamma and alpha emitting radionuclides can deposit sufficient absorbed dose to disrupt a tumor’s vascular structure while minimizing the dose outside the blood vessel. Candidate radionuclides uniformly distributed in microspheres are theoretically investigated with respect to their vascular disruption potential and to offer an alternative to 90Y microsphere therapy. Requisite activities of candidate low-energy beta-gamma and alpha emitting radionuclides to facilitate vascular disruption are calculated. PMID:24749005

  17. Vascular surgery--is it different?

    PubMed

    DeWeese, J A

    1978-12-01

    In 1972 the Society for Vascular Surgery and the North American Chapter of the International Cardiovascular Society recommended that the American Board of Surgery establish a method for the certification of special competence in vascular surgery. The American Board of Surgery in 1974 judged that, for the present, vascular surgery training best could be upgraded by certifying training programs and not individuals. The Residency Review Committee for Surgery now has approved guidelines which define acceptable vascular surgery training programs. These guidelines require approval of the American Medical Association and the governing bodies of the Liaison Committee for Graduate Medical Education before accreditation procedures can be implemented. It is proposed that vascular surgery is different enough from general surgery and cardiothoracic surgery so that special training programs are needed. Vascular surgeons require a fund of knowledge and surgical skills beyond that learned in most surgical programs as well as a special experience with vascular operations if they are to provide optimal patient care. Acceptance of the principle of the accreditation of surgeons caring for vascular surgical problems is an important step in the upgrading of vascular surgery.

  18. Vascular emergencies in liver trauma.

    PubMed

    Taourel, P; Vernhet, H; Suau, A; Granier, C; Lopez, F M; Aufort, S

    2007-10-01

    The use of CT in the diagnosis and management of liver trauma is responsible for the shift from routine surgical versus non-surgical treatment in the management of traumatic liver injuries, even when they are of high grade. The main cause of complication and of death in liver trauma is related to vascular injury. The goal of this review focussed on the vascular complications of liver trauma is to describe the elementary lesions shown by CT in liver trauma including laceration, parenchymal hematoma and contusions, partial devascularisation, subcapsular hematomas, hemoperitoneum, active bleeding, pseudoaneurysm of the hepatic artery, bile leak, and periportal oedema, to illustrate the possible pitfalls in CT diagnosis of liver trauma and to underline the key-points which may absolutely be present in a CT report of liver trauma. Then we will remind the grading system based on the CT features and we will analyze the interest and limitations of such grading systems. Last we will discuss the diagnostic strategy at the early phase in patients with suspected liver trauma according to their clinical conditions and underline the conditions of arterial embolization, and then we will discuss the diagnosis strategy at the delayed phase according to the suspected complications. PMID:17851012

  19. Leaf hydraulics II: vascularized tissues.

    PubMed

    Rockwell, Fulton E; Holbrook, N Michele; Stroock, Abraham D

    2014-01-01

    Current models of leaf hydration employ an Ohm's law analogy of the leaf as an ideal capacitor, neglecting the resistance to flow between cells, or treat the leaf as a plane sheet with a source of water at fixed potential filling the mid-plane, neglecting the discrete placement of veins as well as their resistance. We develop a model of leaf hydration that considers the average conductance of the vascular network to a representative areole (region bounded by the vascular network), and represent the volume of tissue within the areole as a poroelastic composite of cells and air spaces. Solutions to the 3D flow problem are found by numerical simulation, and these results are then compared to 1D models with exact solutions for a range of leaf geometries, based on a survey of temperate woody plants. We then show that the hydration times given by these solutions are well approximated by a sum of the ideal capacitor and plane sheet times, representing the time for transport through the vasculature and tissue respectively. We then develop scaling factors relating this approximate solution to the 3D model, and examine the dependence of these scaling factors on leaf geometry. Finally, we apply a similar strategy to reduce the dimensions of the steady state problem, in the context of peristomatal transpiration, and consider the relation of transpirational gradients to equilibrium leaf water potential measurements.

  20. Computational modeling of vascular anastomoses.

    PubMed

    Migliavacca, Francesco; Dubini, Gabriele

    2005-06-01

    Recent development of computational technology allows a level of knowledge of biomechanical factors in the healthy or pathological cardiovascular system that was unthinkable a few years ago. In particular, computational fluid dynamics (CFD) and computational structural (CS) analyses have been used to evaluate specific quantities, such as fluid and wall stresses and strains, which are very difficult to measure in vivo. Indeed, CFD and CS offer much more variability and resolution than in vitro and in vivo methods, yet computations must be validated by careful comparison with experimental and clinical data. The enormous parallel development of clinical imaging such as magnetic resonance or computed tomography opens a new way toward a detailed patient-specific description of the actual hemodynamics and structural behavior of living tissues. Coupling of CFD/CS and clinical images is becoming a standard evaluation that is expected to become part of the clinical practice in the diagnosis and in the surgical planning in advanced medical centers. This review focuses on computational studies of fluid and structural dynamics of a number of vascular anastomoses: the coronary bypass graft anastomoses, the arterial peripheral anastomoses, the arterio-venous graft anastomoses and the vascular anastomoses performed in the correction of congenital heart diseases. PMID:15772842

  1. Vascular Remodeling in Pulmonary Hypertension

    PubMed Central

    Shimoda, Larissa A; Laurie, Steven S.

    2013-01-01

    Pulmonary hypertension is a complex, progressive condition arising from a variety of genetic and pathogenic causes. Patients present with a spectrum of histologic and pathophysiological features, likely reflecting the diversity in underlying pathogenesis. It is widely recognized that structural alterations in the vascular wall contribute to all forms of pulmonary hypertension. Features characteristic of the remodeled vasculature in patients with pulmonary hypertension include increased stiffening of the elastic proximal pulmonary arteries, thickening of the intimal and/or medial layer of muscular arteries, development of vaso-occlusive lesions and the appearance of cells expressing smooth muscle specific markers in normally non-muscular small diameter vessels, resulting from proliferation and migration of pulmonary arterial smooth muscle cells and cellular trans-differentiation. The development of several animal models of pulmonary hypertension has provided the means to explore the mechanistic underpinnings of pulmonary vascular remodeling, although none of the experimental models currently used entirely replicates the pulmonary arterial hypertension observed in patients. Herein, we provide an overview of the histological abnormalities observed in humans with pulmonary hypertension and in preclinical models and discuss insights gained regarding several key signaling pathways contributing to the remodeling process. In particular, we will focus on the roles of ion homeostasis, endothelin-1, serotonin, bone morphogenetic proteins, Rho kinase and hypoxia-inducible factor 1 in pulmonary arterial smooth muscle and endothelial cells, highlighting areas of cross-talk between these pathways and potentials for therapeutic targeting. PMID:23334338

  2. Reoperation after vascular ring repair.

    PubMed

    Backer, Carl L; Mongé, Michael C; Russell, Hyde M; Popescu, Andrada R; Rastatter, Jeffrey C; Costello, John M

    2014-01-01

    The majority of patients having surgical intervention for a vascular ring have resolution of their symptoms. However, 5% to 10% of these patients develop recurrent symptoms related either to airway or esophageal compression and may require reoperation. In our series of 300 patients with vascular rings, we performed a reoperation on 26 patients, not all of whom were originally operated on at our institution. The four primary indications for reoperation were Kommerell diverticulum (n = 18), circumflex aorta (n = 2), residual scarring (n = 2), and tracheobronchomalacia requiring aortopexy (n = 4). All patients undergoing reoperation have had preoperative evaluation with bronchoscopy and computed tomographic scanning (CT) with 3-dimensional reconstruction. Patients with dysphagia have had a barium esophagram and esophagoscopy. Patients with a Kommerell diverticulum have undergone resection of the diverticulum and transfer of the left subclavian artery to the left carotid artery. The aortic uncrossing procedure has been used in patients with a circumflex aorta. Aortopexy has been used to treat anterior compression of the trachea by the aorta. Results of these reinterventions have been successful in nearly all cases. Lessons learned from these reoperations can be applied to prevent the need for reoperation by properly selecting the correct initial operation. A dedicated team caring for these children consisting of medical imaging, otolaryngology, cardiovascular-thoracic surgery, and critical care is imperative.

  3. CIRSE Vascular Closure Device Registry

    SciTech Connect

    Reekers, Jim A.; Mueller-Huelsbeck, Stefan; Libicher, Martin; Atar, Eli; Trentmann, Jens; Goffette, Pierre; Borggrefe, Jan; Zelenak, Kamil; Hooijboer, Pieter; Belli, Anna-Maria

    2011-02-15

    Purpose: Vascular closure devices are routinely used after many vascular interventional radiology procedures. However, there have been no major multicenter studies to assess the safety and effectiveness of the routine use of closure devices in interventional radiology. Methods: The CIRSE registry of closure devices with an anchor and a plug started in January 2009 and ended in August 2009. A total of 1,107 patients were included in the registry. Results: Deployment success was 97.2%. Deployment failure specified to access type was 8.8% [95% confidence interval (95% CI) 5.0-14.5] for antegrade access and 1.8% (95% CI 1.1-2.9) for retrograde access (P = 0.001). There was no difference in deployment failure related to local PVD at the access site. Calcification was a reason for deployment failure in only <0.5% of patients. Postdeployment bleeding occurred in 6.4%, and most these (51.5%) could be managed with light manual compression. During follow-up, other device-related complications were reported in 1.3%: seven false aneurysms, three hematoma >5.9 cm, and two vessel occlusions. Conclusion: The conclusion of this registry of closure devices with an anchor and a plug is that the use of this device in interventional radiology procedures is safe, with a low incidence of serious access site complications. There seems to be no difference in complications between antegrade and retrograde access and other parameters.

  4. Optimized laser application in dermatology using infrared thermography

    NASA Astrophysics Data System (ADS)

    Thomas, Roderick A.; Donne, Kelvin E.; Clement, Marc; Kiernan, Michael N.

    2002-03-01

    Infrared thermography can be used to optimize the application of lasers in dermatology with particular reference to the treatment of certain skin disorders such as vascular lesions and depilation. The efficacy of treatment is dependent upon a number of factors including: Optimization and correct selection of laser parameters such as wavelength and spot size. Human factors, such as laser operator skill, patient's skin type and anatomical location. By observing the thermal effects of laser irradiation on the skins surface during treatment results in improved efficacy and minimizes the possible threshold to skin damage, reducing the possibility of burning and scarring. This is of particular significance for example, in the control of purpura for the treatment of vascular lesions. The optimization is validated with reference to a computer model that predicts various skin temperatures based on two different laser spot sizes.

  5. Early Clinical Experience With Argon Ion Laser Endarterectomy

    NASA Astrophysics Data System (ADS)

    Eugene, John; Baribeau, Yvon; Ott, Richard A.; McColgan, Stephen J.; Berns, Michael W.

    1989-09-01

    This report describes our progress in the development of argon ion laser endarterectomy for arteriosclerotic cardiovascular disease. Nine patients underwent 10 vascular reconstructions for claudication (6), rest pain (1), and gangrene (2). There was 1 aortoiliac endarterectomy, 6 superficial femoral artery endarterectomies, 1 profunda femoris endarterectomy and 2 popliteal endarterectomies. The reconstructions were 6 cm to 60 cm in length. The operations were performed using low power argon ion laser radiation, 1.0 W. All patients experienced symptomatic relief and had palpable pulses postoperatively. There were no perforations and there were no injuries to surrounding tissues from laser radiation. Surgical complications occurred and these were technical problems that should be eliminated from the operation with further developments. The early clinical results show that laser endarterectomy can be performed for peripheral vascular reconstruction using low power argon ion laser radiation.

  6. Laser Analyzer

    NASA Technical Reports Server (NTRS)

    1993-01-01

    Dopant level analysis is important to the laser system designer because it allows him to model the laser's performance. It also allows the end user to determine what went wrong when a laser fails to perform as expected. Under a Small Business Innovation Research (SBIR) contract, Scientific Materials Corporation has developed a process for producing uniform laser rods in which the amount of water trapped in the crystal during growth is reduced. This research led to the formation of a subsidiary company, Montana Analytical Services, which conducts analysis of laser rods for dopant ion concentrations. This is a significant advance in laser technology.

  7. AB127. The value of diagnostic micro-dissection testicular sperm extraction before start of IVF cycles for couples with non-obstructive azoospermia male factor

    PubMed Central

    Huang, William J.; Huang, Shen; Wei, Tzu-Chun

    2015-01-01

    Objective Micro-dissection testicular sperm extraction (mTESE) nowadays has been the major sperm retrieval method for patients with non-obstructive azoospermia (NOA) in assisted reproduction technology (ART). However, there are still 40% to 50% chances that no sperm can be found after the procedure, and the ICSI cycles are then aborted. Therefore the couples need to take the significant physical, psychological and financial risks, including ovulation induction, eggs retrieval and costs for procedures. We introduce the concept of diagnostic mTESE for men with NOA to determine the decision to initiate ovulation cycles. Methods From 2012 to 2014, 152 men received diagnostic mTESE procedures. This study had excluded patients with obstructive azoospermia and needle biopsy-confirmed hypospermatogenesis cases. Patients with undescended testis, Klinefelter’s syndrome, or Y microdeletion were included. The procedure was performed by Schlegel’s method, the testicular tissues were examined under operative microscope up to 24 X. The procedures were started from right testicle, if no sperm found from right side, then left side was opened consequently. The tubules larger than 300 μm were sampled for immediate table side touch print smear examination and the tissues were transferred to Bouin’s fixatives for further pathological examination. The whole procedure was video-taped for future review. The location of sperm-positive areas was recorded in operation note. The testis wound at tunica albuginea was closed using intra-tunical zipper suture with 6-0 Prolene. Results Among the 152 patients, the mean sperm retrieval rate was 45.3%. If we included the patients with biopsy-confirmed hypospermatogenesis receiving only therapeutic mTESE, the overall sperm retrieval rate was about 61%. For the patients with positive results at diagnostic mTESE, the sperm retrieval rate at later therapeutic mTESE for ICSI cycle was 100%. For patients who had no sperm found at diagnostic m

  8. Estimation of vessel diameter and blood flow dynamics from laser speckle images

    PubMed Central

    Postnov, Dmitry D.; Tuchin, Valery V.; Sosnovtseva, Olga

    2016-01-01

    Laser speckle imaging is a rapidly developing method to study changes of blood velocity in the vascular networks. However, to assess blood flow and vascular responses it is crucial to measure vessel diameter in addition to blood velocity dynamics. We suggest an algorithm that allows for dynamical masking of a vessel position and measurements of it’s diameter from laser speckle images. This approach demonstrates high reliability and stability. PMID:27446704

  9. Lasers of All Sizes

    NASA Astrophysics Data System (ADS)

    Balcou, Philippe; Forget, Sébastien Robert-Philip, Isabelle

    2015-10-01

    * Introduction * The Laser in All Its Forms * Gas lasers * Dye lasers * Solid-state lasers * Lasers for Every Taste * The rise of lasers * Lasers of all sizes * The colors of the rainbow... and beyond * Shorter and shorter lasers * Increasingly powerful lasers * Lasers: A Universal Tool? * Cutting, welding, and cleaning * Communicating * Treating illnesses * Measuring * Supplying energy? * Entertaining * Understanding * Conclusion

  10. The epidemiological profile of the Vascular Birthmark Clinic at the Alberta Children’s Hospital

    PubMed Central

    Fraulin, Frankie OG; Flannigan, Ryan K; Sharma, Vishal K; McPhalen, Donald F; Harrop, Robertson A

    2012-01-01

    BACKGROUND: The Vascular Birthmark (VBM) Clinic at the Alberta Children’s Hospital (Calgary, Alberta) is a multidisciplinary clinic dedicated to the evaluation of children with vascular anomalies. OBJECTIVE: To review the characteristics of patients seen at the VBM Clinic. METHOD: A retrospective data analysis of all pediatric patients presenting to the VBM Clinic between 1998 and 2009 was performed. Data including demographic, referring, diagnostic and treatment information were obtained from the clinic’s database. RESULTS: Of 932 patients, 621 with hemangiomas and 311 patients with vascular malformations were found in the database. Hemangiomas were more commonly found in girls (68.5%), and most commonly located on the head and neck (54%), with most patients (72.6%) having only one lesion. Of the patients with hemangiomas, 14.7% underwent diagnostic imaging investigation and 23.7% received treatment including medications, surgery, pulsed-dye laser or dressings. The sex distribution among the 311 patients with vascular malformations was almost equal. Venous malformations accounted for 38.9% of patients, isolated capillary malformations for 31.5%, lymphatic malformations for 11.6%, mixed low-flow malformations for 14.8% and arteriovenous malformations for 2.9%. Overall, 37.9% of patients underwent diagnostic imaging investigation and 42.4% received treatment – either pulsed-dye laser, surgical excision, sclerotherapy or other treatment. CONCLUSIONS: Hemangiomas and vascular malformations can occur at any anatomical site. There is a large variation in clinical presentation necessitating expertise in a variety of diagnostic approaches and treatment modalities. Vascular anomalies are best managed in a multidisciplinary setting. PMID:23730152

  11. Laser microphone

    DOEpatents

    Veligdan, James T.

    2000-11-14

    A microphone for detecting sound pressure waves includes a laser resonator having a laser gain material aligned coaxially between a pair of first and second mirrors for producing a laser beam. A reference cell is disposed between the laser material and one of the mirrors for transmitting a reference portion of the laser beam between the mirrors. A sensing cell is disposed between the laser material and one of the mirrors, and is laterally displaced from the reference cell for transmitting a signal portion of the laser beam, with the sensing cell being open for receiving the sound waves. A photodetector is disposed in optical communication with the first mirror for receiving the laser beam, and produces an acoustic signal therefrom for the sound waves.

  12. Enhanced vascularization of cultured skin substitutes genetically modified to overexpress vascular endothelial growth factor.

    PubMed

    Supp, D M; Supp, A P; Bell, S M; Boyce, S T

    2000-01-01

    Cultured skin substitutes have been used as adjunctive therapies in the treatment of burns and chronic wounds, but they are limited by lack of a vascular plexus. This deficiency leads to greater time for vascularization compared with native skin autografts and contributes to graft failure. Genetic modification of cultured skin substitutes to enhance vascularization could hypothetically lead to improved wound healing. To address this hypothesis, human keratinocytes were genetically modified by transduction with a replication incompetent retrovirus to overexpress vascular endothelial growth factor, a specific and potent mitogen for endothelial cells. Cultured skin substitutes consisting of collagen-glycosaminoglycan substrates inoculated with human fibroblasts and either vascular endothelial growth factor-modified or control keratinocytes were prepared, and were cultured in vitro for 21 d. Northern blot analysis demonstrated enhanced expression of vascular endothelial growth factor mRNA in genetically modified keratinocytes and in cultured skin substitutes prepared with modified cells. Furthermore, the vascular endothelial growth factor-modified cultured skin substitutes secreted greatly elevated levels of vascular endothelial growth factor protein throughout the entire culture period. The bioactivity of vascular endothelial growth factor protein secreted by the genetically modified cultured skin substitutes was demonstrated using a microvascular endothelial cell growth assay. Vascular endothelial growth factor-modified and control cultured skin substitutes were grafted to full-thickness wounds on athymic mice, and elevated vascular endothelial growth factor mRNA expression was detected in the modified grafts for at least 2 wk after surgery. Vascular endothelial growth factor-modified grafts exhibited increased numbers of dermal blood vessels and decreased time to vascularization compared with controls. These results indicate that genetic modification of

  13. Vascular tumors of the choroid and retina

    PubMed Central

    Shanmugam, P Mahesh; Ramanjulu, Rajesh

    2015-01-01

    Vascular tumors of the retina and choroid can be seen occasionally. In the following article, the key clinical and diagnostic features of the major retinal and choroidal vascular tumors, their systemic associations, and the literature pertaining to the most currently available treatment strategies are reviewed. PMID:25827544

  14. 21 CFR 870.3250 - Vascular clip.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Vascular clip. 870.3250 Section 870.3250 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CARDIOVASCULAR DEVICES Cardiovascular Prosthetic Devices § 870.3250 Vascular clip. (a) Identification. A...

  15. 21 CFR 870.3250 - Vascular clip.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Vascular clip. 870.3250 Section 870.3250 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CARDIOVASCULAR DEVICES Cardiovascular Prosthetic Devices § 870.3250 Vascular clip. (a) Identification. A...

  16. A history of vascular and microvascular surgery.

    PubMed

    Rickard, Rory F; Hudson, Donald A

    2014-10-01

    The history of microvascular surgery is intimately linked to that of vascular surgery. Microvascular techniques, developed mainly in China, Japan, Australia, and the United States of America, built on the principles of vascular anastomosis established by pioneers in France, Germany, Italy, and the United States of America. We present a history of the technique here.

  17. Functional preservation of vascular smooth muscle tissue

    NASA Technical Reports Server (NTRS)

    Alexander, W. C.; Hutchins, P. M.; Kimzey, S. L.

    1973-01-01

    The ionic and cellular feedback relationships operating to effect the vascular decompensatory modifications were examined to reveal procedures for implementing protective measures guarding against vascular collapse when returning from a weightless environment to that of the earth's gravity. The surgical procedures for preparing the rat cremaster, and the fixation methods are described. Abstracts of publications resulting from this research are included.

  18. Covariance of lichen and vascular plant floras

    USGS Publications Warehouse

    Bennett, J.P.; Wetmore, C.M.

    1999-01-01

    The geographic relationships among taxonomic groups are important to study to determine patterns of biodiversity and whether or not associations occur between large groups, e.g., birds and vascular plants. This study was undertaken to determine relationships between higher plants and lower plants, specifically vascular plant and lichen floras in nine national parks of the Great Lakes region. No significant relationship was found between vascular plant floras and lichen floras in this area, which spans 1200 km longitudinally, or between an additional 19 areas from North America that were less than 1000 km(2) in area. For areas larger than 1000 km(2), however, a significant positive relationship existed for 33 areas that span one to approximately 150 million km(2). The ratio of numbers of vascular plants to lichens appeared to average just over 6 across the 33 areas. In the Great Lakes parks, between 28-30% of either the vascular plant or lichen species were singletons (occurring in only one park), but the parks that contained the most singletons were not congruent: Isle Royale had the most singleton lichens, while Indiana Dunes had the most vascular plant singletons. Fewer lichen species (2%) than vascular plants (4%) occurred in all nine parks. Latitude appeared to explain some of the variation between the two groups: vascular plants decreased with increasing latitude, while lichens increased.

  19. A large vascular leiomyoma of the leg.

    PubMed

    Cigna, E; Maruccia, M; Malzone, G; Malpassini, F; Soda, G; Drudi, F M

    2012-06-01

    A 69-year-old woman with a subcutaneous, large vascular leiomyoma of the leg is presented. The patient had a painful, slow-growing, right medial malleolus mass. Clinical symptoms, US images and histopathologic features are reported. Vascular leiomyoma should be included in the differential diagnosis of painful, lower extremity subcutaneous masses also in lesions of larger dimensions.

  20. Abdominal Distension and Vascular Collapse.

    PubMed

    Cosentino, Gina; Uwaifo, Gabriel I

    2016-04-01

    We present the case of a 43-year-old gentleman who presented to the emergency room with acute abdominal distension, confusion and vascular collapse. The emergent radiologic imaging obtained showed massive bilateral adrenal enlargement, but despite the initial clinical suspicion of possible overwhelming sepsis and/or massive abdominal/intralesional hemorrhage, lab tests based obtained rapidly confirmed the diagnosis of acute Addisonian crisis which responded dramatically to adrenocorticoid hormone replacement therapy and aggressive fluid resuscitation. The patient's established history of metastatic lung cancer confirmed this as a case of metastatic massive bilateral adrenal metastases with an initial presentation of acute adrenal insufficiency which is uncommon in the setting of metastatic carcinomatosis but more typically associated with lymphomas. Recognition of this clinical possibility is vital to enable rapid diagnosis and consequent life saving therapy. PMID:27328473