ImageJ Freeware Research Articles

A Quantitative Description of Lipid and Extracellular Matrix Proteinaceous Fibers in Hepatic Fibrosis of a Rat Model by ImageJ using Nano-Images

We made a histological and ultrastructural studies using ImageJ freeware and Origin pro statistical analysis of the site of lipid accumulations and ECM pertinacious collagen fibers in a rat model of hepatic fibrosis using CCl4 and ethanol dual effect and using nano GTE as the main factor to diminishes both hepatic lipid and ECM fibers. This imageJ observation indicates and re-enforces the notion previously published by the same author that Nano GTE is a strong therapeutic agent to work against the oxidative effects of chemicals like CCl4 and ethanol.

Histopathology of Chloroplast Ultrastructure and Inclusion Bodies in CMV-Infected Plant Tissues Using Electron Microscopic Examination and Image Analysis

Histopathology of Chloroplast Ultrastructure and Inclusion Bodies in CMV-Infected Plant Tissues Using Electron Microscopic Examination and ImageJ Analysis Cucumber mosaic virus (CMV) is an economically important virus that causes wide-spread diseases damaging crops worldwide. Two strains of the virus were detected using ELISA with specific antibodies, and isolated from Kuwait environment designated CMVKU1 and CMV-KU2. The KU1 is a mild strain and is associated with a benign viral satellite RNA, while KU2 isolate, is a severe strain and lacking viral satellite RNA. Both strains were used in a comparative study to investigate the effects of CMV infection on the ultrastructure of chloroplasts and inclusion bodies in test plants of tomato (Lycopersicon esculentum) and squash (Cucurbita pepo). Transmission electron microscopy (TEM) was used to determine the extent of infection and damage caused by virus isolates on chloroplast structure, and cellular inclusions including plastoglobules, and starch granules. Chloroplasts, the photosynthetic apparatus of plants, in infected tissues were severely affected with KU2 strain that caused a reduction and damage of their main structure, number, size, and inclusion bodies compared to the benign strain of KU1 and healthy control plants. Electron micrographs of ultra-thin sections proved the presence of the viral nanoparticles. The formation of viral crystalline bodies, chloroplast damage and malformation due to the viral infection. As the infection progressed, chloroplasts were destroyed and grana were disorganized and scattered into the cytoplasm that causes reduction in chlorophyll contents in plant tissues. ImageJ freeware analysis were used to determine the extent of infection and host-virus interactions and damage caused by virus isolates...

Lower energy and pulse stacking. A safer alternative for skin tightening using fractional CO2 laser.

To evaluate the effect of different energies and stacking in skin shrinkage. Three decreasing settings of a fractional CO2 laser were applied to the abdomen of Twenty five Wistar rats divided into three groups. Group I (n=5) was histologically evaluated for microthermal zones dimensions. Groups II and III (n=10 each) were macroscopic evaluated with freeware ImageJ for area contraction immediately and after 30 and 60 days. No statistical significance was found within microthermal zone histological dimensions (Group I) in all settings studied. (Ablation depth: 76.90 to 97.18µm; Coagulation depth: 186.01 to 219.84 µm). In Group II, macroscopic evaluation showed that all settings cause significant immediate skin contraction. The highest setting cause significant more intense tightening effect initially, contracting skin area from 258.65 to 179.09 mm2. The same pattern was observed in Group III. At 30 and 60 days, the lowest setting significantly sustained contraction. Lower fractional CO2 laser energies associated to pulse stacking could cause consistent and long lasting tissue contraction in rats.

Opacificación capsular posterior en lentes intraoculares hidrofílicos en comparación con lentes intraoculares hidrofóbicos usando imágenes de Scheimpflug

PurposeTo determine the visual outcomes and posterior capsule opacification in acrylic intraocular lenses (IOL). MethodsProspective analysis of 38 eyes of 19 patients who had uneventful phacoemulsification with implantation of a IOL Akreos MI60 or a IOL Acrysof IQ from February 2013 to January 2014. Patients from a single surgeon, the protocol included the determination logMAR best corrected visual acuity, spherical equivalent, contrast sensitivity assessed by the freiburg visual acuity test and posterior capsule opacification scoring using the evaluation of posterior capsular opacification system (EPCO2000) by digital retroillumination photography and high resolution tomograms from Scheimpflug reconstructions analyzed using ImageJ freeware. ResultsThe mean follow up was 6.2 months±1.4 (SD). Contrast sensitivity and best corrected visual acuity were similar between the lenses. Mean pixel intensity value using EPCO2000 software for Akreos MI60 was 1.08U and 1.08U for Acrysof IQ (P=.785). Using ImageJ mean value in pixels for Akreos MI60 was 1.76U and for Acrysof IQ was 2.56U (P=.027). There were no significant correlationship between EPCO2000 and Scheimpflug images (r=.141, P=.412). ConclusionsThis study found that the results of visual function were comparable between both lenses. The posterior capsular opacification was seen to be lesser extent with the Akreos MI60 IOL when measured by Scheimpflug images, probably due to the very design of the lens which showed no correlation compared to the digital retroillumination photoghaphy.

A Method For Tree-Ring Analysis UsingDiva-GisFreeware On Scanned Core Images

Tree-ring analysis is a basic technique of paramount importance in forest management, yet it may prove difficult and time-consuming for many slow-growth hardwood tree species. Moreover, it requires the use of specialized tools and proprietary software, which may hinder researchers working with limited budgets. We describe an innovative and inexpensive method using DIVA-GIS freeware software to analyze true color high-resolution scanned images of cores previously enhanced with ImageJ freeware (GIS-SDI), and test its accuracy against the widely-used LINTAB-TSAPWin™ and WinDENDRO™ methods. For this purpose, Abies pinsapo and Pyrus bourgaeana increment cores were processed independently using each of the three methods and the results were statistically compared. Dating results were consistent across all three methods, although identifying rings was easier and quicker to perform on the digital images. Using a modern but affordable flatbed scanner to digitize tree cores and the free DIVA-GIS software to analyze the scanned digital images proved to be an inexpensive but highly accurate and efficient approach to tree-ring analysis. Furthermore, this method greatly facilitates tree-ring analysis in species with inconspicuous rings, and enables a complete digital record of every core analyzed to be stored.

Non-invasive and non-destructive measurements of confluence in cultured adherent cell lines

Many protocols used for measuring the growth of adherent monolayer cells in vitro are invasive, destructive and do not allow for the continued, undisturbed growth of cells within flasks. Protocols often use indirect methods for measuring proliferation. Microscopy techniques can analyse cell proliferation in a non-invasive or non-destructive manner but often use expensive equipment and software algorithms. In this method images of cells within flasks are captured by photographing under a standard inverted phase contract light microscope using a digital camera with a camera lens adaptor. Images are analysed for confluence using ImageJ freeware resulting in a measure of confluence known as an Area Fraction (AF) output. An example of the AF method in use on OVCAR8 and UPN251 cell lines is included.•Measurements of confluence from growing adherent cell lines in cell culture flasks is obtained in a non-invasive, non-destructive, label-free manner.•The technique is quick, affordable and eliminates sample manipulation.•The technique provides an objective, consistent measure of when cells reach confluence and is highly correlated to manual counting with a haemocytometer. The average correlation co-efficient from a Spearman correlation (n=3) was 0.99±0.008 for OVCAR8 (p=0.01) and 0.99±0.01 for UPN251 (p=0.01) cell lines.

Reliability of Proliferation Assessment by Ki-67 Expression in Neuroendocrine Neoplasms: Eyeballing or Image Analysis?

Background: The latest WHO classification for neuroendocrine neoplasms (NEN) of the gastrointestinal tract defines grade according to Ki-67 and mitotic indices. Some have questioned the reproducibility and thus the reliability of Ki-67 assessment. We therefore investigated the accuracy of this proliferation marker in NEN. Methods: The Ki-67 index of tumor specimens of NEN (n = 73) was assessed by two pathologists as in routine practice with eyeballing and twice by image analysis using ImageJ freeware at different magnifications. Results were correlated with overall survival. Results: The intraclass correlation coefficient (ICC) between pathologists was 0.88. The ICC for the measurements using image analysis was 0.85. The ICC between all four measurements (pathologists and ImageJ) was 0.80. If the Ki-67 index was translated to grade as prescribed by the current WHO classification (<3% = grade 1, 3-20% = grade 2, >20% = grade 3), kappa was between 0.61 and 0.75. Grades based on pathologist scoring were often (16-29%) higher than grades assigned by image analysis (p < 0.001). Grade was significantly correlated with survival (p < 0.0001) irrespective of the way Ki-67 was assessed. Conclusion: Assessment of the Ki-67 index by eyeballing correlates remarkably well with the Ki-67 index as calculated by image analysis and is therefore an accurate parameter. Moreover, it is significantly related to survival irrespective of the method used. Yet if the Ki-67 index is translated to grade, the grade should be interpreted with caution due to values around threshold levels.

Topical treatment with a new matrix therapy agent (RGTA, CACICOL‐20) improves epithelial wound healing after penetrating keratoplasty

AbstractPurpose Complete epithelial wound healing is a milestone in the early post‐operative care after penetrating keratoplasty (PKP). It reduces the infectious risk, allows safe instillation of steroids, and conditions discharge from hospitalization. We assessed a new matrix therapy agent, mimicking heparan sulfates, for the management of post PKP epithelial defectsMethods Prospective, open‐label, uncontrolled, single‐centre study on 33 consecutive patients (33 eyes) who benefited from a 8.25 mm PKP for all‐comers indications. Patients were treated with RGTA eye drops (THEA): one drop in the operating theater immediately after graft, then on alternate days. Follow‐up was performed by slit‐lamp examination and photography with fluorescein‐dye testing repeated every day at a fixed time. Dye area was measured using the freeware ImageJ. Main outcome: time of corneal surface healingResults All corneas displayed complete healing after a mean 2.7±1.1 days (median 3, range; 1 to 6). Complete wound healing was obtained at D1 for 15%, D2 for 33%, D3 for 88%, D4 for 94%, and D6 for 100%. Area of epithelial defect decreased by a mean 75±22% between D1 and D2 (n=28) corresponding to a wound healing speed of 11.5±6.5 mm2/D. There were no systemic or local side effects. For comparison, in the series of all‐comers indications of 1003 PKP without RGTA of Borderie, mean wound healing was 4.6+/‐13.2 days (median 2.5, range 1‐210).Conclusion RGTA seems a potentially useful, noninvasive therapeutic approach in PKP management. A controlled double blind clinical trial vs placebo will be soon conducted to answer this question conclusively.

Using enhanced green fluorescent protein (EGFP) promoter fusions to study gene regulation at single cell and population levels.

Reporter gene fusions based on the enhanced green fluorescent protein (EGFP) are powerful experimental tools that allow real-time changes in gene expression to be monitored both in single cells and in populations. Here we describe the development of a chromosomally integrated transcriptional reporter fusion in Listeria monocytogenes that allows real-time measurements of gene expression. To construct a single copy of an EGFP-based fluorescent reporter fused to a promoter of interest (Px) in L. monocytogenes, a suicide shuttle vector carrying the Px::egfp gene fusion is first constructed in Escherichia coli (as an intermediate host). Then, the vector is transformed into L. monocytogenes and integrated into its chromosome by homologous recombination within the selected promoter region. Subsequently, analysis of fluorescence exhibited by cells carrying a single copy reporter can be performed under selected experimental conditions by stringent sample preparation, optimized image acquisition, and processing of the digital data with the image analysis freeware ImageJ. Thus, the methodology described here can be adapted to investigate the activity and regulation of any promoter in L. monocytogenes both at the cell and population levels.

Non-destructive analyses on a meteorite fragment that fell in the Madrid city centre in 1896

The historical Madrid meteorite chondrite fell in 1896 showing thin melt veins with a 65% of brecciated forsterite fragments surrounded by a fine grained matrix formed by troilite, chromite and Fe–Ni blebs. It exhibits a delicate iron infill, neo-formation of troilite in pockets and shock veins and neo-formation of Na-feldspar formed at high temperature and fast quenching. The semi-quantitative mineral determinations were performed with IMAGEJ freeware and chemical mappings resulting in the following approximated compositions: olivine (∼55%); augite (∼10%); enstatite (∼10%); plagioclase (∼10%); chromite (∼2%); troilite (∼4%), kamacite–taenite α–γ-(Fe, Ni) (∼7%) and merrillite (∼7%). The specimen was also studied by computer tomography, micro-Raman spectroscopy and spectral cathodoluminescence. X-ray diffraction patterns were also recorded in non-destructive way on a polished surface because of the small size of the specimen. This combination of non-destructive techniques provides an improved knowledge on the Madrid-1896 meteorite compared to the previous study performed on the same specimen carried out twenty years ago by electron probe microanalysis and optical microscopy in destructive way. Limits of these techniques are the specimen's size in the analytical chambers and the threshold resolution of the microscopes analyzing shock veins micro-crystals.

【Short Report】A Quick Method to Estimate Root Length in Each Diameter Class Using Freeware ImageJ

Digital images for determining root traits have been analyzed using WinRHIZO, commercial software. A freeware ImageJ has been developed for the analysis but whether root lengths assigned to root diameter classes obtained by this method correspond to those estimated by WinRHIZO has not been confirmed. In this study, root lengths estimated using ImageJ were compared with those estimated using WinRHIZO. In ImageJ, the root lengths were estimated using a triangle thresholding algorithm that was valid for total root length in our previous study. For the small-diameter classes (<0.5 mm), the estimates obtained using ImageJ and the conversion factor 2/3, corresponded closely to the estimates obtained using WinRHIZO. For large-diameter classes (>0.5 mm), root lengths estimated using ImageJ were similar to those obtained using WinRHIZO without using the conversion factor. These results were valid for rice roots. In this report, we propose completely automated estimation of root lengths for roots in each diameter class by using ImageJ.

Quantitative evaluation of efficacy of photodynamic therapy for port-wine stains using erythema index image analysis

Methods for quantitatively evaluate the efficacy of photodynamic therapy (PDT) for port-wine stains (PWS) are still needed for clinical practice and studies. 50 pairs of pictures before and after PDT of 40 PWS patients were selected. Each PWS lesion was labeled with a marker of red color graded 0-9. These pictures were taken without assistance of instruments to keep same position or distance and were referred to as 'experimental' images. 70 labels were photographed at a fixed position and distance with the assistance of a bracket and the images obtained were referred to as 'standardized' ones. An independent group of three experts viewed the photos and assessed the efficacy. The images were processed and measured for erythema index (EI) with the ImageJ freeware. The EI difference (ΔEI) and the percent change of ΔEI ((ratio)ΔEI(%)) of the labels and the PWS lesions was computed separately. Significant differences of EI, ΔEI and (ratio)ΔEI(%) were found between each two grades of the color markers in both standardized and experimental images. ΔEI of lesions achieved 'almost cured' and 'great improvement' after PDT was significantly reduced than before PDT. Significantly greater percentages of lesions were assessed as 'response' and 'significant response' in those of (before)ΔEI≥35 compared to those of (before)ΔEI<35. The (ratio)ΔEI(%) decrease of lesions assessed as 'almost cured', 'great improvement' and 'some improvement' was significantly reduced sequentially. The EI image analysis is a valid method for quantitatively evaluating efficacy of PDT for PWS.

FRET Microscopy for Real-time Monitoring of Signaling Events in Live Cells Using Unimolecular Biosensors

Förster resonance energy transfer (FRET) microscopy continues to gain increasing interest as a technique for real-time monitoring of biochemical and signaling events in live cells and tissues. Compared to classical biochemical methods, this novel technology is characterized by high temporal and spatial resolution. FRET experiments use various genetically-encoded biosensors which can be expressed and imaged over time in situ or in vivo. Typical biosensors can either report protein-protein interactions by measuring FRET between a fluorophore-tagged pair of proteins or conformational changes in a single protein which harbors donor and acceptor fluorophores interconnected with a binding moiety for a molecule of interest. Bimolecular biosensors for protein-protein interactions include, for example, constructs designed to monitor G-protein activation in cells, while the unimolecular sensors measuring conformational changes are widely used to image second messengers such as calcium, cAMP, inositol phosphates and cGMP. Here we describe how to build a customized epifluorescence FRET imaging system from single commercially available components and how to control the whole setup using the Micro-Manager freeware. This simple but powerful instrument is designed for routine or more sophisticated FRET measurements in live cells. Acquired images are processed using self-written plug-ins to visualize changes in FRET ratio in real-time during any experiments before being stored in a graphics format compatible with the build-in ImageJ freeware used for subsequent data analysis. This low-cost system is characterized by high flexibility and can be successfully used to monitor various biochemical events and signaling molecules by a plethora of available FRET biosensors in live cells and tissues. As an example, we demonstrate how to use this imaging system to perform real-time monitoring of cAMP in live 293A cells upon stimulation with a β-adrenergic receptor agonist and blocker.

A simple reproducible and time saving method of semi-automatic dendrite spine density estimation compared to manual spine counting

Estimation of spine number and spine density by manual counting under the assumption that all dendrite protrusions equal spines are often used in studies on neuroplasticity occurring during health, brain diseases, and different experimental paradigms. Manual spine counting is, however, time consuming and biased by inter-observer variation. We present accordingly a quick, reproducible and simple non-stereological semi-automatic spine density estimation method based on the irregularity of the dendrite surface.Using the freeware ImageJ program, microphotographs of Golgi impregnated hippocampal dendrites derived from a previously performed study on the impact of chronic restrained stress were binarized, skeletonized, and the skeleton endings assumed to represent spine positions were counted and the spine densities calculated. The results based on 754 dendrite fragments were compared to manual spine counting of the same dendrite fragments using the Bland–Altman method.The results from both methods were correlated (r=0.79, p<0.0001), The semi-automatic counting method gave a statistically higher (approx. 4%) spine density number, but both counting methods showed similar significant differences between the groups in the CA1 area, and no differences between the groups in the CA3 area.In conclusion, the presented semi-automatic spine density estimation method yields consistently a higher spine density number than manual counting resulting in similar significance between groups. The proposed method may therefore be a reproducible time saving and useful non-stereological approach to spine counting in neuroplasticity studies requiring analysis of hundreds of dendrites.

Comparison of threshold algorithms for automatic image processing of rice roots using freeware ImageJ

High-throughput phenotyping of crop root system architecture using digital image analysis requires costly special software (WinRHIZO). Novel freeware, ImageJ, has also been developed for the similar purpose, but the presetting of the threshold to create root binary images makes this method time-consuming. This study compares the 16 algorithms available in the ImageJ for processing of rice root images. Among the algorithms, the Triangle algorithm proved to be the best binary method, where the coefficient of correlation between ImageJ-estimated and WinRHIZO-estimated root lengths is extremely high ( r = 0.986). However, using the Triangle algorithm, ImageJ overestimated the rice root length compared with WinRHIZO. By multiplying the values obtained using ImageJ by 2/3, the estimates closely corresponded to those estimated by WinRHIZO. The correspondence of root lengths estimated using WinRHIZO and ImageJ with Triangle algorithm was valid for roots of various morphologies, and for rice plants grown in uplands, rainfed lowlands, and irrigated lowlands. This report proposes the completely automated estimation of rice root length using freeware ImageJ with the appropriate threshold algorithm for image processing.

Gas7 Functions with N-WASP to Regulate the Neurite Outgrowth of Hippocampal Neurons

Neuritogenesis, or neurite outgrowth, is a critical process for neuronal differentiation and maturation in which growth cones are formed from highly dynamic actin structures. Gas7 (growth arrest-specific gene 7), a new member of the PCH (Pombe Cdc15 homology) protein family, is predominantly expressed in neurons and is required for the maturation of primary cultured Purkinje neurons as well as the neuron-like differentiation of PC12 cells upon nerve growth factor stimulation. We report that Gas7 co-localizes and physically interacts with N-WASP, a key regulator of Arp2/3 complex-mediated actin polymerization, in the cortical region of Gas7-transfected Neuro-2a cells and growth cones of hippocampal neurons. The interaction between Gas7 and N-WASP is mediated by WW-Pro domains, which is unique in the PCH protein family, where most interactions are of the SH3-Pro kind. The interaction contributes to the formation of membrane protrusions and processes by recruiting the Arp2/3 complex in a Cdc42-independent manner. Importantly, specific interaction between Gas7 and N-WASP is required for regular neurite outgrowth of hippocampal neurons. The data demonstrate an essential role of Gas7 through its interaction with N-WASP during neuronal maturation/differentiation.

Confocal laser‐scanning capillaroscopy: a novel approach to the analysis of skin capillaries in vivo

New techniques for diagnostics and therapy in dermatology are becoming increasingly non-invasive, among which confocal laser-scanning microscopy (CLSM) is the most prevalent. It allows visualization of cellular structures of the skin up to a depth of 300 microm in vivo. Until now, most studies have been conducted on pathologically altered skin, mostly oncologic lesions. We now present a detailed analysis of capillaries located in the upper dermal papillae. Multiple measurements were performed on the dorsal and ventral surface of the right forearm of 30 healthy volunteers (22-88 years) under standard conditions (room temperature, body position, time of day). Images were obtained with the Vivascope 1500 (Lucid) under standard settings and analyzed using the freeware ImageJ with a customwritten macro plugin. The following parameters of the capillaries in vivo were measured: area, perimeter, circularity and maximum diameter. Statistical analysis showed that all four parameters were constant within a narrow range, regardless of the body site, sex and age. In this physiological study, we can clearly demonstrate that by confocal laser-scanning capillaroscopy (CLSC), it is possible to visualize and measure skin capillaries at the extremities in a reproducible manner. This new approach offers a considerable advantage compared with nailfold capillaroscopy, which can only be performed at the proximal nail segment, and over histological analysis, which can be hampered by fixation artifacts resulting in altered size and shape of the vessels to be analyzed. CLSC could allow for precise analysis of in vivo skin vasculature in systemic and proliferative diseases of the skin.

Pentacam Tomograms: A Novel Method for Quantification of Posterior Capsule Opacification

To develop and validate a method to quantify posterior capsule opacification (PCO) in eyes after cataract surgery and intraocular lens implantation using Scheimpflug Pentacam tomograms and compare its validity with slit lamp retroillumination image analysis. One hundred twenty-four pseudophakic eyes of 124 patients were divided into two groups. Group 1 consisted of 40 eyes with visually significant PCO, and group 2 consisted of 84 eyes without visually significant PCO. Pentacam Imaging was performed after full mydriasis using the 50-scan acquisition protocol, and high-resolution tomograms were reconstructed and analyzed using ImageJ freeware. Retroillumination photographs were captured for group 1 eyes using the Topcon digital slit lamp, and these were analyzed using POCOman software to calculate an aggregate severity grade and percentage PCO value. Correlation coefficients were calculated for PCO values obtained using POCOman and ImageJ. Mean PCO percentage value obtained using POCOman software was 23.34 +/- 6.25 U, mean aggregate PCO severity grade was 0.46 +/- 0.28 U, and mean pixel-intensity value using ImageJ was 31.071 +/- 8.26 U. There was a significant positive correlation between the percentage PCO (P = 0.000; r = 0.864) and PCO severity grade (P = 0.001; r = 0.490) obtained for group 1 eyes using slit lamp retroillumination images and PCO pixel intensity obtained using Pentacam tomograms. Retroillumination photographs are the standard for quantifying PCO. Pentacam tomograms are easier to obtain and are free of flash reflections, and they allow for a more objective analysis. The correlation between the two methods demonstrates that ImageJ analysis of Pentacam tomograms is a viable tool for PCO analysis.

Testing Mounting Media to Eliminate Background Noise in Confocal Microscope 3‐D Images of Insect Genitalia

Recently, the confocal laser scanning microscope (CLSM) has been used to image and generate three-dimensional reconstructions of miniscule insect tissues and cuticular structures. These three-dimensional reconstructions provide the investigator with key information concerning the spatial relationship among structures and substructures. Unfortunately, there can be high levels of background "noise" which can obscure the specimen in these three-dimensional reconstructions. This background "noise" might be a result of the mounting media either autofluorescing or reflecting and scattering the imaged specimen's fluorescence. The standard nonpermanent mounting medium is glycerine jelly (a 1:17:17 ratio of porcine gelatin to glycerine to water). In this study, the organic molecule (lipid, protein, or carbohydrate) added to the glycerine water mixture was varied. The relative background to specimen signal (the mean voxel brightness reading in ImageJ freeware) was compared across mountants. The mounting media tested are ranked from best (least background noise) to worst (most background noise) as follows: agarose, agar, pectin, gelatin (the standard), petroleum jelly. A 1% agarose mountant (1:50:50 ratio of agarose to glycerine to water) is recommended because it causes little to no background noise, provides consistent high quality contrast between specimen and background with increasing depth, and is easy to handle.

H-REV107-1 Stimulates Growth in Non-Small Cell Lung Carcinomas via the Activation of Mitogenic Signaling

H-REV107-1, a known member of the class II tumor suppressor gene family, is involved in the regulation of differentiation and survival. We analyzed H-REV107-1 in non-small cell lung carcinomas, in normal lung, and in immortalized and tumor-derived cell lines. Sixty-eight percent of lung tumors revealed positive H-REV107-1-specific staining. Furthermore, survival analysis demonstrated a significant association of cytoplasmic H-REV107-1 with decreased patient survival. This suggested that H-REV107-1, known as a tumor suppressor, plays a different role in non-small cell lung carcinomas. Knock-down of H-REV107-1 expression in lung carcinoma cells inhibited anchorage-dependent and anchorage-independent growth whereas overexpression of H-REV107-1 induced tumor cell proliferation. Consistent with results of the survival analysis, cytoplasmic localization of the protein was essential for this growth-inducing function. Analysis of signaling pathways potentially involved in this process demonstrated that overexpression of H-REV107-1 stimulated RAS-GTPase activity, ERK1,2 phosphorylation, and caveolin-1 expression in the cell lines analyzed. These results indicate that H-REV107-1 is deficient in its function as a tumor suppressor in non-small cell lung carcinomas and is required for proliferation and anchorage-independent growth in cells expressing high levels of the protein, thus contributing to tumor progression in a subset of non-small cell lung carcinomas.