Expression of Selected miRNAs in Undifferentiated Carcinoma with Osteoclast-like Giant Cells (UCOGC) of the Pancreas: Comparison with Poorly Differentiated Pancreatic Ductal Adenocarcinoma.

Undifferentiated carcinoma with osteoclast-like giant cells (UCOGC) of the pancreas represents a rare subtype of pancreatic ductal adenocarcinoma (PDAC). Despite a distinct morphology and specific clinical behavior, UCOGCs exhibit unexpected similarities in regard to DNA mutational profiles with conventional PDAC. Treating pancreatic ductal adenocarcinoma is particularly challenging, with limited prospects for cure. As with many other malignant neoplasms, the exploration of microRNAs (miRNAs, miRs) in regulating the biological characteristics of pancreatic cancer is undergoing extensive investigation to enhance tumor diagnostics and unveil the therapeutic possibilities. Herein, we evaluated the expression of miR-21, -96, -148a, -155, -196a, -210, and -217 in UCOGCs and poorly differentiated (grade 3, G3) PDACs. The expression of miR-21, miR-155, and miR-210 in both UCOGCs and G3 PDACs was significantly upregulated compared to the levels in normal tissue, while the levels of miR-148a and miR-217 were downregulated. We did not find any significant differences between cancerous and normal tissues for the expression of miR-96 and miR-196a in G3 PDACs, whereas miR-196a was slightly, but significantly, downregulated in UCOGCs. On the other hand, we have not observed significant differences in the expression of the majority of miRNAs between UCOGC and G3 PDAC, with the exception of miR-155. UCOGC samples demonstrated lower mean levels of miR-155 in comparison with those in G3 PDACs.

Expression of Selected miRNAs in Normal and Cancer-Associated Fibroblasts and in BxPc3 and MIA PaCa-2 Cell Lines of Pancreatic Ductal Adenocarcinoma.

Therapy for pancreatic ductal adenocarcinoma remains challenging, and the chances of a complete cure are very limited. As in other types of cancer, the expression and role of miRNAs in controlling the biological properties of this type of tumor have been extensively studied. A better insight into miRNA biology seems critical to refining diagnostics and improving their therapeutic potential. In this study, we focused on the expression of miR-21, -96, -196a, -210, and -217 in normal fibroblasts, cancer-associated fibroblasts prepared from a ductal adenocarcinoma of the pancreas, and pancreatic carcinoma cell lines. We compared these data with miRNAs in homogenates of paraffin-embedded sections from normal pancreatic tissues. In cancer-associated fibroblasts and cancer cell lines, miRNAs differed significantly from the normal tissue. In detail, miR-21 and -210 were significantly upregulated, while miR-217 was downregulated. Similar transcription profiles were earlier reported in cancer-associated fibroblasts exposed to hypoxia. However, the cells in our study were cultured under normoxic conditions. We also noted a relation to IL-6 production. In conclusion, cultured cancer-associated fibroblasts and carcinoma cells reflect miR-21 and -210 expression similarly to the cancer tissue samples harvested from the patients.

Senescence-Associated miRNAs and Their Role in Pancreatic Cancer.

Replicative senescence is irreversible cell proliferation arrest for somatic cells which can be circumvented in cancers. Cellular senescence is a process, which may play two opposite roles. On the one hand, this is a natural protection of somatic cells against unlimited proliferation and malignant transformation. On the other hand, cellular secretion caused by senescence can stimulate inflammation and proliferation of adjacent cells that may promote malignancy. The main genes controlling the senescence pathways are also well known as tumor suppressors. Almost 140 genes regulate both cellular senescence and cancer pathways. About two thirds of these genes (64%) are regulated by microRNAs. Senescence-associated miRNAs can stimulate cancer progression or act as tumor suppressors. Here we review the role playing by senescence-associated miRNAs in development, diagnostics and treatment of pancreatic cancer.

Hemorheological alterations of red blood cells induced by 450-nm and 520-nm laser radiation.

Proper rheological properties of red blood cells (RBC) including flexibility and aggregability are essential for healthy blood microcirculation. Excessive RBC aggregation has been observed to be associated with many pathological conditions and is crucial in acute circulatory problems. Low-level laser radiation (LLLR) has been found to have positive effects on the rheology of human blood, however, the detailed mechanisms of blood photobiomodulation remains unclear. In this study, utilizing the single-cell technique optical tweezers (OT) and traditional light microscopy, the influence of photobiomodulation of human RBC was examined under different conditions of laser irradiation. The results revealed that high radiant exposure (over 170.5 J/cm2 radiant fluence) caused enhanced RBC aggregation and cell shape transformation while the aggregation force between single RBC remained unchanged. LLLR with radiant fluence below 9.5 J/cm2 by 450 nm wavelength improved the RBC deformability, weakened the strength of cell-cell interaction in the RBC disaggregation process, and showed rejuvenating effects on RBC suspended in a harsh cell environment.

Development of oral cancer tissue-mimicking phantom based on polyvinyl chloride plastisol and graphite for terahertz frequencies.

SIGNIFICANCE: A new concept of a biotissue phantom for terahertz (THz) biomedical applications is needed for reliable and long-term usage. AIM: We aimed to develop a new type of biotissue phantom without water content and with controllable THz optical properties by applying graphite powders into a polyvinyl chloride plastisol (PVCP) matrix and to give a numerical description to the THz optical properties of the phantoms using the Bruggeman model (BM) of the effective medium theory (EMT). APPROACH: The THz optical properties of graphite and the PVCP matrix were measured using THz time-domain spectroscopy, which works in the frequency range from 0.1 to 1 THz. Two phantoms with 10% and 12.5% graphite were fabricated to evaluate the feasibility of describing phantoms using the EMT. The EMT then was used to determine the concentration of graphite required to mimic the THz optical properties of human cancerous and healthy oral tissue. RESULTS: The phantom with 16.7% of graphite has the similar THz optical properties as human cancerous oral tissue in the frequency range of 0.2 to 0.7 THz. The THz optical properties of the phantom with 21.9% of graphite are close to those of human healthy oral tissue in the bandwidth from 0.6 to 0.8 THz. Both the refractive index and absorption coefficient of the samples increase with an increase of graphite concentration. The BM of the EMT was used as the numerical model to describe the THz optical properties of the phantoms. The relative error of the BM for the refractive index estimation and the absorption coefficient is up to 4% and 8%, respectively. CONCLUSIONS: A water-free biotissue phantom that mimics the THz optical properties of human cancerous oral tissue was developed. With 21.9% of graphite, the phantom also mimics human healthy oral tissue in a narrow frequency range. The BM proved to be a suitable numerical model of the phantom.

Evaluating ß-amyloidosis progression in Alzheimer's disease with Mueller polarimetry.

We applied the wide-field Mueller imaging polarimetry for the screening of formalin-fixed paraffin-embedded samples of mouse brain tissue at different stages of brain ß-amyloidosis in Alzheimer's disease (AD). The accumulation of amyloid-beta (Aß) deposits throughout the brain tissue is one of the key pathological hallmarks observed with the AD progression. We demonstrate that the presence of Aß plaques influences the properties of backscattered polarized light, in particular, its degree of depolarization. By means of statistical analysis, we demonstrate that the high-order statistical moments of depolarization distributions, acquired with the multi-spectral Mueller imaging polarimetry, can be used as sensitive markers of the growing presence of Aß plaques. The introduced label-free polarimetric approach has a potential to facilitate the current practice of the histopathology screening in terms of diagnosis accuracy, time and cost efficiency.

Publisher Correction: Tumor-necrosis factor impairs CD4+ T cell-mediated immunological control in chronic viral infection.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Colon cancer detection by using Poincaré sphere and 2D polarimetric mapping of ex vivo colon samples.

This work is dedicated to the diagnosis and grading of colon cancer by a combined use of Poincaré sphere and 2D Stokes vector polarimetry mapping approaches. The major challenge consists in exploring the applicability of polarized light for noninvasive screening of the histological abnormalities within the samples of biological tissues. Experimental studies were conducted in ex vivo colon sample, excised after surgical procedure for colon tumor removal of G2-adenocarcinoma lesion. Polarimetric measurements in linear and circular regime were carried via personally developed polarimetric, optical set-up, using supercontinuous fiber laser with irradiation fixed at 635 nm. We apply the Poincaré sphere and two-dimensional Stokes vector scanning approach for screening the corresponding tissue samples. A comparison between linear and circular polarization states is made both for quantitative and qualitative evaluations. It is shown that circular polarization has better diagnostic capabilities than linear polarization, with higher dynamic ranges of the polarimetric parameters and better values of the diagnostic quantities. In addition to the standard polarimetry parameters, utilized as essential diagnostic markers, we apply statistical analysis to obtain more detailed information in frame of the applied diagnostic approach.

Biodegradable Nanocarriers Resembling Extracellular Vesicles Deliver Genetic Material with the Highest Efficiency to Various Cell Types.

Efficient delivery of genetic material to primary cells remains challenging. Here, efficient transfer of genetic material is presented using synthetic biodegradable nanocarriers, resembling extracellular vesicles in their biomechanical properties. This is based on two main technological achievements: generation of soft biodegradable polyelectrolyte capsules in nanosize and efficient application of the nanocapsules for co-transfer of different RNAs to tumor cell lines and primary cells, including hematopoietic progenitor cells and primary T cells. Near to 100% efficiency is reached using only 2.5 × 10-4 pmol of siRNA, and 1 × 10-3 nmol of mRNA per cell, which is several magnitude orders below the amounts reported for any of methods published so far. The data show that biodegradable nanocapsules represent a universal and highly efficient biomimetic platform for the transfer of genetic material with the utmost potential to revolutionize gene transfer technology in vitro and in vivo.

Double-Stranded DNA Fragments Bearing Unrepairable Lesions and Their Internalization into Mouse Krebs-2 Carcinoma Cells.

Murine Krebs-2 tumor-initiating stem cells are known to natively internalize extracellular double-stranded DNA fragments. Being internalized, these fragments interfere in the repair of chemically induced interstrand cross-links. In the current investigation, 756 bp polymerase chain reaction (PCR) product containing bulky photoreactive dC adduct was used as extracellular DNA. This adduct was shown to inhibit the cellular system of nucleotide excision repair while being resistant to excision by this DNA repair system. The basic parameters for this DNA probe internalization by the murine Krebs-2 tumor cells were characterized. Being incubated under regular conditions (60 min, 24°C, 500 µL of the incubation medium, in the dark), 0.35% ± 0.18% of the Krebs-2 ascites cells were shown to natively internalize modified DNA. The saturating amount of the modified DNA was detected to be 0.37 µg per 106 cells. For the similar unmodified DNA fragments, this ratio is 0.73 µg per 106 cells. Krebs-2 tumor cells were shown to be saturated internalizing either (190 ± 40) × 103 molecules of modified DNA or (1,000 ± 100) × 103 molecules of native DNA. On internalization, the fragments of DNA undergo partial and nonuniform hydrolysis of 3' ends followed by circularization. The degree of hydrolysis, assessed by sequencing of several clones with the insertion of specific PCR product, was 30-60 nucleotides.

Deep Airway Inflammation and Respiratory Disorders in Nanocomposite Workers.

Thousands of researchers and workers worldwide are employed in nanocomposites manufacturing, yet little is known about their respiratory health. Aerosol exposures were characterized using real time and integrated instruments. Aerosol mass concentration ranged from 0.120 mg/m³ to 1.840 mg/m³ during nanocomposite machining processes; median particle number concentration ranged from 4.8 × 104 to 5.4 × 105 particles/cm³. The proportion of nanoparticles varied by process from 40 to 95%. Twenty employees, working in nanocomposite materials research were examined pre-shift and post-shift using spirometry and fractional exhaled nitric oxide (FeNO) in parallel with 21 controls. Pro-inflammatory leukotrienes (LT) type B4, C4, D4, and E4; tumor necrosis factor (TNF); interleukins; and anti-inflammatory lipoxins (LXA4 and LXB4) were analyzed in their exhaled breath condensate (EBC). Chronic bronchitis was present in 20% of researchers, but not in controls. A significant decrease in forced expiratory volume in 1 s (FEV1) and FEV1/forced vital capacity (FVC) was found in researchers post-shift (p ˂ 0.05). Post-shift EBC samples were higher for TNF (p ˂ 0.001), LTB4 (p ˂ 0.001), and LTE4 (p ˂ 0.01) compared with controls. Nanocomposites production was associated with LTB4 (p ˂ 0.001), LTE4 (p ˂ 0.05), and TNF (p ˂ 0.001), in addition to pre-shift LTD4 and LXB4 (both p ˂ 0.05). Spirometry documented minor, but significant, post-shift lung impairment. TNF and LTB4 were the most robust markers of biological effects. Proper ventilation and respiratory protection are required during nanocomposites processing.

Tumor-necrosis factor impairs CD4(+) T cell-mediated immunological control in chronic viral infection.

Persistent viral infections are characterized by the simultaneous presence of chronic inflammation and T cell dysfunction. In prototypic models of chronicity--infection with human immunodeficiency virus (HIV) or lymphocytic choriomeningitis virus (LCMV)--we used transcriptome-based modeling to reveal that CD4(+) T cells were co-exposed not only to multiple inhibitory signals but also to tumor-necrosis factor (TNF). Blockade of TNF during chronic infection with LCMV abrogated the inhibitory gene-expression signature in CD4(+) T cells, including reduced expression of the inhibitory receptor PD-1, and reconstituted virus-specific immunity, which led to control of infection. Preventing signaling via the TNF receptor selectively in T cells sufficed to induce these effects. Targeted immunological interventions to disrupt the TNF-mediated link between chronic inflammation and T cell dysfunction might therefore lead to therapies to overcome persistent viral infection.

Self-assembly of endohedral metallofullerenes: a decisive role of cooling gas and metal-carbon bonding.

The endohedral metallofullerene (EMF) self-assembly process in Sc/carbon vapor in the presence and absence of an inert cooling gas (helium) is systematically investigated using quantum chemical molecular dynamics simulations. It is revealed that the presence of He atoms accelerates the formation of pentagons and hexagons and reduces the size of the self-assembled carbon cages in comparison with analogous He-free simulations. As a result, the Sc/C/He system simulations produce a larger number of successful trajectories (i.e. leading to Sc-EMFs) with more realistic cage-size distribution than simulations of the Sc/C system. The main Sc encapsulation mechanism involves nucleation of several hexagons and pentagons with Sc atoms already at the early stages of carbon vapor condensation. In such proto-cages, both Sc-C σ-bonds and coordination bonds between Sc atoms and the π-system of the carbon network are present. Sc atoms are thus rather labile and can move along the carbon network, but the overall bonding is sufficiently strong to prevent dissociation even at temperatures around 2000 kelvin. Further growth of the fullerene cage results in the encapsulation of one or two Sc atoms within the fullerene. In agreement with experimental studies, an extension of the simulations to Fe and Ti as the metal component showed that Fe-EMFs are not formed at all, whereas Ti is prone to form Ti-EMFs with small cage sizes, including Ti@C28-Td and Ti@C30-C2v(3).

Small nucleolar RNA U91 is a new internal control for accurate microRNAs quantification in pancreatic cancer.

BACKGROUND: RT-qPCR quantification of miRNAs expression may play an essential role in pancreatic ductal adenocarcinoma (PDAC) diagnostics. RT-qPCR-based experiments require endogenous controls for the result normalization and reliability. However, expression instability of reference genes in tumors may introduce bias when determining miRNA levels. METHODS: We investigated expression of 6 miRNAs, isolated from FFPE samples of pancreatic adenocarcinomas. Four internal controls were utilized for RT-qPCR result normalization: artificial miR-39 from C. elegans, U6 snRNA, miR-16 and snoRNA U91. RESULTS: We found miR-21, miR-155 or miR-217 expression values in tumors may differ up to several times, depending on selected internal controls. Moreover, different internal controls can produce controversial results for miR-96, miR-148a or miR-196a quantification. Also, expression of our endogenous controls varied significantly in tumors. U6 demonstrated variation from -1.03 to 8.12-fold, miR-16 from -2.94 up to 7.38-fold and the U91 from -3.05 to 4.36-fold respectively. On the other hand, the most stable gene, determined by NormFinder algorithm, was U91. Each miRNA normalized relatively to the spike or U91, demonstrated similar expression values. Thus, statistically significant and insignificant differences between tumors and normal tissues for miRNAs were equal for the spike and the U91. Also, the differences between the spike and U91 were statistically insignificant for all of miRs except miR-217. Among three endogenous controls, U91 had the lowest average expression values and standard deviation in cancer tissues. CONCLUSIONS: We recommend U91 as a new normalizer for miRNA quantification in PDACs.

Use of optical skin phantoms for preclinical evaluation of laser efficiency for skin lesion therapy.

Skin lesions are commonly treated using laser heating. However, the introduction of new devices into clinical practice requires evaluation of their performance. This study presents the application of optical phantoms for assessment of a newly developed 975-nm pulsed diode laser system for dermatological purposes. Such phantoms closely mimic the absorption and scattering of real human skin (although not precisely in relation to thermal conductivity and capacitance); thus, they can be used as substitutes for human skin for approximate evaluation of laser heating efficiency in an almost real environment. Thermographic imaging was applied to measure the spatial and temporal temperature distributions on the surface of laser-irradiated phantoms. The study yielded results of heating with regard to phantom thickness and absorption, as well as laser settings. The methodology developed can be used in practice for preclinical evaluations of laser treatment for dermatology.

Single-step gas-phase polyperfluoroalkylation of naphthalene leads to thermodynamic products.

High-temperature gas-phase, solvent- and catalyst-free reaction of naphthalene with an excess of RF I reagent (RF CF3 , C2 F5 , n-C3 F7 , and n-C4 F9 ) was used for the first time to produce a series of highly perfluoroalkylated naphthalene products NAPH(RF )n with n=2-5. Four 95+ % pure 1,3,5,7-NAPH(RF )4 with RF CF3 , C2 F5 , n-C3 F7 , and n-C4 F9 were isolated using a simple chromatography-free procedure. These new compounds were fully characterized by (19) F and (1) H NMR spectroscopy, X-ray crystallography (for RF CF3 and C2 F5 ), atmospheric-pressure chemical ionization mass spectrometry, and cyclic and square-wave voltammetry. DFT calculations confirm that the proposed synthesis yields the most stable isomers that have not been accessed by alternative preparation techniques.

Free-standing single-atom-thick iron membranes suspended in graphene pores.

The excess of surface dangling bonds makes the formation of free-standing two-dimensional (2D) metals unstable and hence difficult to achieve. To date, only a few reports have demonstrated 2D metal formation over substrates. Here, we show a free-standing crystalline single-atom-thick layer of iron (Fe) using in situ low-voltage aberration-corrected transmission electron microscopy and supporting image simulations. First-principles calculations confirm enhanced magnetic properties for single-atom-thick 2D Fe membranes. This work could pave the way for new 2D structures to be formed in graphene membranes.

Feasibility study of the optical imaging of a breast cancer lesion labeled with upconversion nanoparticle biocomplexes.

Innovative luminescent nanomaterials, termed upconversion nanoparticles (UCNPs), have demonstrated considerable promise as molecular probes for high-contrast optical imaging in cells and small animals. The feasibility study of optical diagnostics in humans is reported here based on experimental and theoretical modeling of optical imaging of an UCNP-labeled breast cancer lesion. UCNPs synthesized in-house were surface-capped with an amphiphilic polymer to achieve good colloidal stability in aqueous buffer solutions. The scFv4D5 mini-antibodies were grafted onto the UCNPs via a high-affinity molecular linker barstar:barnase (Bs:Bn) to allow their specific binding to the human epidermal growth factor receptor HER2/neu, which is overexpressed in human breast adenocarcinoma cells SK-BR-3. UCNP-Bs:Bn-scFv4D5 biocomplexes exhibited high-specific immobilization on the SK-BR-3 cells with the optical contrast as high as 10:1 benchmarked against a negative control cell line. Breast cancer optical diagnostics was experimentally modeled by means of epi-luminescence imaging of a monolayer of the UCNP-labeled SK-BR-3 cells buried under a breast tissue mimicking optical phantom. The experimental results were analyzed theoretically and projected to in vivo detection of early-stage breast cancer. The model predicts that the UCNP-assisted cancer detection is feasible up to 4 mm in tissue depth, showing considerable potential for diagnostic and image-guided surgery applications.

The properties of human CD40-activated B cells as antigen-presenting cells are not affected by PGE2.

Tumor vaccination represents a promising immuno-therapeutic strategy in cancer. However, the inherent ability of many tumors to evade immune responses by suppression of immune cell function represents a major barrier. Prostaglandin E2 (PGE2) has been shown to be a critical tumor-derived immunosuppressive factor. It affects a broad range of immune cells including T cells, macrophages and dendritic cells (DCs). CD40-activated B cells are being studied as a potential alternative to DCs as antigen-presenting cells for immunotherapy. So far, it is not known whether PGE2 affects their antigen presenting capacity. We, therefore, investigated the influence of PGE2 on the phenotype, migratory potential and antigen-presenting function of CD40-activated human B cells. Here, we demonstrate that the immunostimulatory properties of CD40-activated B cells are not affected by PGE2. These results support the use of CD40-activated B cells as cellular adjuvants, especially in settings where PGE2 is present in the tumor microenvironment.

Indoleamine 2,3-dioxygenase-expressing myeloid dendritic cells and macrophages in infectious and noninfectious cutaneous granulomas.

BACKGROUND: The enzyme indoleamine 2,3-dioxygenase (IDO) degrades the essential amino acid tryptophan, and this degradation is an immunosuppressive mechanism that is mainly used by antigen-presenting cells. IDO-expressing dendritic cells and macrophages have previously been identified as components of lymph node granulomas after Listeria monocytogenes infection. In this study we undertook an analysis of IDO expression in granulomas of infectious and noninfectious origin in the human skin. METHODS: Lesional skin biopsy specimens (n = 22) from different granulomatous skin disorders (lupus vulgaris, sarcoidosis, granuloma annulare, leprosy) were analyzed. Immunohistochemistry was performed to identify and locate the enzyme IDO within the inflammatory granulomatous infiltrate (IDO, CD11c, CD68, S100, CD3, Foxp3). Two-color immunofluorescence of IDO in combination with multiple markers was applied to characterize the IDO-expressing cells. RESULTS: Cutaneous granulomas of different origin strongly express IDO, mainly in the center and in the ring wall of the granulomas. We demonstrate that in infectious, but also in noninfectious human cutaneous granulomas the large myeloid CD11c(+)S100(+)CD68(-) dendritic cells and the CD68(+) macrophages express IDO. LIMITATIONS: This study was limited by the lack of details about the exact stage or maturity of granuloma formation in the specimens investigated. CONCLUSION: These findings reveal that IDO expression in myeloid dendritic cells and macrophages is part of an integrated response of granuloma formation, which may be a unifying feature of granulomatous reactions in the skin.