Growth Pattern Of Cells Research Articles

Unsupervised class discovery in pancreatic ductal adenocarcinoma reveals cell-intrinsic mesenchymal features and high concordance between existing classification systems

Pancreatic ductal adenocarcinoma (PDAC) has the worst prognosis of all common cancers. However, divergent outcomes exist between patients, suggesting distinct underlying tumor biology. Here, we delineated this heterogeneity, compared interconnectivity between classification systems, and experimentally addressed the tumor biology that drives poor outcome. RNA-sequencing of 90 resected specimens and unsupervised classification revealed four subgroups associated with distinct outcomes. The worst-prognosis subtype was characterized by mesenchymal gene signatures. Comparative (network) analysis showed high interconnectivity with previously identified classification schemes and high robustness of the mesenchymal subtype. From species-specific transcript analysis of matching patient-derived xenografts we constructed dedicated classifiers for experimental models. Detailed assessments of tumor growth in subtyped experimental models revealed that a highly invasive growth pattern of mesenchymal subtype tumor cells is responsible for its poor outcome. Concluding, by developing a classification system tailored to experimental models, we have uncovered subtype-specific biology that should be further explored to improve treatment of a group of PDAC patients that currently has little therapeutic benefit from surgical treatment.

Complex patterns of cell growth in the placenta in normal pregnancy and as adaptations to maternal diet restriction.

The major milestones in mouse placental development are well described, but our understanding is limited to how the placenta can adapt to damage or changes in the environment. By using stereology and expression of cell cycle markers, we found that the placenta grows under normal conditions not just by hyperplasia of trophoblast cells but also through extensive polyploidy and cell hypertrophy. In response to feeding a low protein diet to mothers prior to and during pregnancy, to mimic chronic malnutrition, we found that this normal program was altered and that it was influenced by the sex of the conceptus. Male fetuses showed intrauterine growth restriction (IUGR) by embryonic day (E) 18.5, just before term, whereas female fetuses showed IUGR as early as E16.5. This difference was correlated with differences in the size of the labyrinth layer of the placenta, the site of nutrient and gas exchange. Functional changes were implied based on up-regulation of nutrient transporter genes. The junctional zone was also affected, with a reduction in both glycogen trophoblast and spongiotrophoblast cells. These changes were associated with increased expression of Phlda2 and reduced expression of Egfr. Polyploidy, which results from endoreduplication, is a normal feature of trophoblast giant cells (TGC) but also spongiotrophoblast cells. Ploidy was increased in sinusoidal-TGCs and spongiotrophoblast cells, but not parietal-TGCs, in low protein placentas. These results indicate that the placenta undergoes a range of changes in development and function in response to poor maternal diet, many of which we interpret are aimed at mitigating the impacts on fetal and maternal health.

Dermatan sulfate epimerase 1 expression and mislocalization may interfere with dermatan sulfate synthesis and breast cancer cell growth

Dermatan sulfate (DS) is a glycosaminoglycan (GAG) that is produced through the epimerization of the glucuronic acid on chondroitin sulfate into iduronic acid (IduA) by dermatan sulfate epimerase (DS-epi) 1 and 2. Proteoglycans (PGs) play essential physiological and pathological roles during cellular development, proliferation, differentiation, and cancer metastasis. DS proteoglycans play vital roles during the process of tumorigenesis, due to the increased flexibility of the polysaccharide chain in the presence of IduA residues, which facilitate specific interactions with proteins, such as growth factors, cytokines, and angiogenic factors. Furthermore, DS-epi is highly expressed in many tumors, especially in esophageal squamous cell carcinoma. This study aimed to investigate the expression of DS-epi1 in multiple breast cancer cell lines, including MCF7 (luminal A), MDA-MB-231 (triple-negative) and SKBR3 (human epidermal growth factor receptor 2-positive), and its involvement in cancer progression. A SKBR3 variant, SKBR3m, presented the most erratic cell growth pattern when compared with those for MCF7 and MDA-MB-231. Moreover, SKBR3m cells demonstrated the highest level of DS-epi1 gene expression and higher 35S-DS content. However, at the protein level, MCF7 cells displayed the highest protein level for DS-epi1, whereas MDA-MB-231 cells had the lowest level. DS-epi1 was found in vesicles and in the perinuclear compartment only in SKBR3m cells, suggesting localization in the Golgi apparatus in these cells, in contrast with the cytoplasmic localization observed in MCF7 and MDA-MB-231 cells. The cytoplasm location of DS-epi1 likely compromised the formation of DS chains, but the core protein was detected using a decorin antibody. Golgi-specific labeling confirmed the localization of DS-epi1 in SKBR3m cells at the Golgi apparatus, indicating that the location of the enzyme was a determinant for the synthesis of DS in this cell line, suggesting that DS may play a decisive role in the tumor growth observed in this breast cancer cell line.

DROUGHT STRESS IMPACTS ON WHEAT AND ITS RESISTANCE MECHANISMS

Scarcity of water has been a serious agricultural hindrance to crop productivity since antiquity. Drought-stressed loss in wheat yield likely exceeds losses from all other causes, since both the severity and duration of the stress are censorious. Here, we have reviewed the effects of drought stress on the morphological, physiological, and biochemical attributes along with the growth impacts, water relations, and photosynthesis impacts in wheat. This review also illustrates the mechanism of drought resistance in wheat. Historical drought years in Nepal have been identified and the yield losses were assessed. Wheat encounters a variety of morphological, physiological, biochemical responses at cellular and molecular levels towards prevailing water stress, thus making it a complex phenomenon. Drought stress affects leaf size, stems elongation and root proliferation, imbalance plant-water relations and decline water-use efficiency. Different types of physiological research are ongoing to find out the changes occurs in the wheat plant as a result of drought stress. Morphological changes can be looked through two ways: changes in root system and changes in shoot system such as effects on height, leaf senescence, flowering, and so on. Physiological changes involve changes in cell growth pattern, chlorophyll contents, photosynthetic disturbances, plant-water relations, etc. Biochemical changes occur in different chemical, biomolecules, and enzymes. Plants portray several mechanisms to withstand drought stress which can be classified as Drought escape, Drought avoidance, and Drought tolerance. Selection of wheat genotype that can tolerate water scarcity would be suitable for the breeding program aiming to development of drought tolerant variety under water limited regions.

Free thiol groups on poly(aspartamide) based hydrogels facilitate tooth-derived progenitor cell proliferation and differentiation.

Cell-based tissue reconstruction is an important field of regenerative medicine. Stem and progenitor cells derived from tooth-associated tissues have strong regeneration potential. However, their in vivo application requires the development of novel scaffolds that will provide a suitable three-dimensional (3D) environment allowing not only the survival of the cells but eliciting their proliferation and differentiation. Our aim was to study the viability and differentiation capacity of periodontal ligament cells (PDLCs) cultured on recently developed biocompatible and biodegradable poly(aspartamide) (PASP)-based hydrogels. Viability and behavior of PDLCs were investigated on PASP-based hydrogels possessing different chemical, physical and mechanical properties. Based on our previous results, the effect of thiol group density in the polymer matrix on cell viability, morphology and differentiation ability is in the focus of our article. The chemical composition and 3D structures of the hydrogels were determined by FT Raman spectroscopy and Scanning Electron Microscopy. Morphology of the cells was examined by phase contrast microscopy. To visualize cell growth and migration patterns through the hydrogels, two-photon microscopy were utilized. Cell viability analysis was performed according to a standardized protocol using WST-1 reagent. PDLCs were able to attach and grow on PASP-based hydrogels. An increase in gel stiffness enhanced adhesion and proliferation of the cells. However, the highest population of viable cells was observed on the PASP gels containing free thiol groups. The presence of thiol groups does not only enhance viability but also facilitates the osteogenic direction of the differentiating cells. These cell-gel structures seem to be highly promising for cell-based tissue reconstruction purposes in the field of regenerative medicine.

Antibacterial activity of various chitosan forms against Xanthomonas axonopodis pv. glycines

In this study, the antibacterial activities of colloidal chitosan, chitosan solution, and chitooligosaccharide solution were evaluated against Xanthomonas axonopodis pv. glycines grown in peptone sucrose broth (PSB) medium. Treatment with colloidal chitosan (0.01, 0.025, and 0.05%) inhibited X. axonopodis pv. glycines growth only until 36 h. Thin-layer chromatography analysis detected some metabolites, consistently with the cell growth pattern. Two chitooligosaccharides (1–3 kDa and 5–10 kDa) dissolved in distilled water and acetic acid did not exhibit antibacterial activity against X. axonopodis pv. glycines at all tested concentrations (0, 0.001, 0.005, 0.01, 0.015, 0.02, 0.025, and 0.05%). Compared to the control, the chitosan solution decreased X. axonopodis pv. glycines cell growth by 58.7% and 99.0% at concentrations of 0.015% and 0.02%, respectively, after 3 d of incubation. The chitosan solution exhibited the highest antibacterial activity at pH 6.5. However, the antibacterial activity of chitosan decreased in the presence of NaCl and MgCl2.

ConvPath: A software tool for lung adenocarcinoma digital pathological image analysis aided by a convolutional neural network.

BackgroundThe spatial distributions of different types of cells could reveal a cancer cell's growth pattern, its relationships with the tumor microenvironment and the immune response of the body, all of which represent key “hallmarks of cancer”. However, the process by which pathologists manually recognize and localize all the cells in pathology slides is extremely labor intensive and error prone.MethodsIn this study, we developed an automated cell type classification pipeline, ConvPath, which includes nuclei segmentation, convolutional neural network-based tumor cell, stromal cell, and lymphocyte classification, and extraction of tumor microenvironment-related features for lung cancer pathology images. To facilitate users in leveraging this pipeline for their research, all source scripts for ConvPath software are available at https://qbrc.swmed.edu/projects/cnn/.FindingsThe overall classification accuracy was 92.9% and 90.1% in training and independent testing datasets, respectively. By identifying cells and classifying cell types, this pipeline can convert a pathology image into a “spatial map” of tumor, stromal and lymphocyte cells. From this spatial map, we can extract features that characterize the tumor micro-environment. Based on these features, we developed an image feature-based prognostic model and validated the model in two independent cohorts. The predicted risk group serves as an independent prognostic factor, after adjusting for clinical variables that include age, gender, smoking status, and stage.InterpretationThe analysis pipeline developed in this study could convert the pathology image into a “spatial map” of tumor cells, stromal cells and lymphocytes. This could greatly facilitate and empower comprehensive analysis of the spatial organization of cells, as well as their roles in tumor progression and metastasis.

PTEN-knockdown disrupts the morphology, growth pattern and function of Nthy-Ori 3-1 cells by downregulating PAX8 expression.

The incidence of thyroid disorders, which are common endocrine diseases, has rapidly increased in recent years. However, the etiology and pathogenesis of these disorders remain unclear. Phosphatase and tension homolog (PTEN) is a dual-specific phosphatase that is associated with multiple thyroid disorders; however, the role of PTEN in thyroid disorders remains unknown. In the present study, the human thyroid follicular epithelial cell line Nthy-Ori 3-1 was used to determine the role of PTEN in thyroid disorders. PTEN expression was knocked down using a PTEN-specific short hairpin RNA. Western blotting was subsequently used to determine protein expression, the Matrigel tube formation assay and iodide uptake assay were applied for evaluating the morphology and function of thyroid cells. The results showed that PTEN knockdown decreased the protein expression of paired box 8 (PAX8). The morphology and tubular-like growth pattern of thyroid cells were therefore disrupted, and restoration of PAX8 expression reversed these effects. Furthermore, PTEN-knockdown decreased the expression of specific thyroid proteins (thyroglobulin, TG; thyroid peroxidase, TPO; and sodium/iodide symporter, NIS) and inhibited the iodide uptake ability of thyroid cells by downregulating PAX8, suggesting that PTEN deficiency may impair the function of thyroid cells. In conclusion, the present study reported an important function of PTEN in normal thyroid cells and identified the involvement of PAX8. These results may improve understanding of the role of PTEN in the pathogenesis of thyroid disorders.

Effects of substrate patterning on cellular spheroid growth and dynamics measured by gradient light interference microscopy (GLIM).

The development of three-dimensional (3D) cellular architectures during development and pathological processes involves intricate migratory patterns that are modulated by genetics and the surrounding microenvironment. The substrate composition of cell cultures has been demonstrated to influence growth, proliferation and migration in 2D. Here, we study the growth and dynamics of mouse embryonic fibroblast cultures patterned in a tissue sheet which then exhibits 3D growth. Using gradient light interference microscopy (GLIM), a label-free quantitative phase imaging approach, we explored the influence of geometry on cell growth patterns and rotational dynamics. We apply, for the first time to our knowledge, dispersion-relation phase spectroscopy (DPS) in polar coordinates to generate the radial and rotational cell mass-transport. Our data show that cells cultured on engineered substrates undergo rotational transport in a radially independent manner and exhibit faster vertical growth than the control, unpatterned cells. The use of GLIM and polar DPS provides a novel quantitative approach to studying the effects of spatially patterned substrates on cell motility and growth.

Offshore Crude Oil Disrupts Retinoid Signaling and Eye Development in Larval Atlantic Haddock

The aim of this study was to examine the impact of dispersed crude oil on retinoid metabolism and signaling in developing haddock larvae (Melanogrammus aeglefinus). Retinoids and its active metabolites are involved in the regulation of several developmental processes as cell differentiation, growth and limb patterning. Thus, disruption of retinoid signaling, especially during early developmental stages, may have detrimental effects on the organism. In the present study, crude oil exposure during embryonic development and early larvae development disrupted vitamin A signaling by affecting retinoid levels and genes involved in retinoid metabolism. The disruption of retinoid signaling was also evident when looking at the gene expression pattern at several time-points during and after exposure, demonstrating the complexity and temporal factor of retinoid signaling disruption. Exposure during the embryonic period (pre hatch) resulted in reduced eye size and increased incidence of abnormal eye morphology. The observed changes in expression of retinoid metabolic genes related to the visual cycle indicates that there is a link between the modulated retinoid levels (all-trans retinoic acid and retinol) and increased incidence of abnormal eye development as a result of crude oil exposure. This is the first study to demonstrate disrupted retinoid signaling in fish following crude oil exposure at environmental relevant levels, giving novel insight in to the mechanism of toxicity.

Ellipse packing in two-dimensional cell tessellation: a theoretical explanation for Lewis's law and Aboav-Weaire's law.

BackgroundLewis’s law and Aboav-Weaire’s law are two fundamental laws used to describe the topology of two-dimensional (2D) structures; however, their theoretical bases remain unclear.MethodsWe used R software with the Conicfit package to fit ellipses based on the geometric parameters of polygonal cells of ten different kinds of natural and artificial 2D structures.ResultsOur results indicated that the cells could be classified as an ellipse’s inscribed polygon (EIP) and that they tended to form the ellipse’s maximal inscribed polygon (EMIP). This phenomenon was named as ellipse packing. On the basis of the number of cell edges, cell area, and semi-axes of fitted ellipses, we derived and verified new relations of Lewis’s law and Aboav-Weaire’s law.ConclusionsEllipse packing is a short-range order that places restrictions on the cell topology and growth pattern. Lewis’s law and Aboav-Weaire’s law mainly reflect the effect of deformation from circle to ellipse on cell area and the edge number of neighboring cells, respectively. The results of this study could be used to simulate the dynamics of cell topology during growth.

Novel xanthine oxidase-based cell model using HK-2 cell for screening antihyperuricemic functional compounds

Hyperuricemia is a metabolic disease caused by disorders of purine metabolism, the prevalence of which has increased worldwide. Here, a cell model for high uric acid production was established in vitro employing cultured human kidney cells (HK-2 cells), and its molecular basis was analyzed using gene expression profile. High performance liquid chromatography (HPLC) was used to monitor the content of metabolites in cell culture media. Adenosine addition was found to induce HK-2 cells to produce uric acid precursors (inosine and hypoxanthine). Furthermore, the cell model was verified by confirming the antihyperuricemic effect of the widely used antihyperuricemic drugs allopurinol, probenecid, and febuxostat, as well as reported bioactive peptides and amino acids, encompassing glutathione, tryptophan and carnosine, which significantly reduced uric acid production in the HK-2 cells (p < 0.05). RNA-Seq technology was used to perform a wide transcriptome analysis of the hyperuricemic cell model, and the results demonstrated that it has the potential to be used as a rapid and valid in vitro model to screen antihyperuricemic compounds that mimics in vivo cell growth patterns.

Alternative Approach for Fighting Bacteria and Fungi: Use of Modified Fluorapatite.

The recent studies on fluorapatite and hydroxyapatite, in the context of their medical applications, have shown that the former has relatively higher thermal stability and better mechanical properties than the latter. Moreover, the presence of fluoride ions is relevant for protection of teeth from dental caries since they stimulate processes of mineralization and crystallization. In this report, a silver modified fluorapatite (FAP/Ag), considered as a novel biomedical compound, was tested for its bactericidal, fungicidal and cytotoxicity activity. All these features were compared to the impact of pure FAP and were evaluated against the bacteria and fungi strains, which constitute main pathogenic species among orthopedic clinical isolates of implant-associated infections. Generally, considerable increase in the antimicrobial activity was observed when silver modified fluorapatite was compared to the pure material. This was manifested among others by disturbance of cell growth pattern and various deformations in the final cell shape as revealed using atomic force microscopy (AFM). Regarding toxicity nowadays as a major issue in implantation, we additionally examined whether the Ag+ ions have an impact on human keratinocytes and mouse fibroblasts using a cytotoxicity assay. To conclude, owing to a great bactericidal potential without collateral cytotoxicity effect the fluorapatite doped with silver ions may be considered a promising biocomponent useful in medical and healthcare applications.

Cold stress affects cell wall deposition and growth pattern in tobacco pollen tubes

Cold is an abiotic stress seriously threatening crop productivity by decreasing biomass production. The pollen tube is a target of cold stress, but also a useful model to address questions on cell wall biosynthesis. We here provide (immuno)cytological data relative to the impact of cold on the pollen tube cell wall. We clearly show that the growth pattern is severely affected by the stress, since the typical pulsed-growth mechanism accompanied by the periodic deposition of pectin rings is absent/severely reduced. Additionally, pectins and cellulose accumulate in bulges provoked by the stress, while callose, which colocalizes with pectins in the periodic rings formed during pulsed growth, accumulates randomly in the stressed samples. The altered distribution of the cell wall components is accompanied by differences in the localization of glucan synthases: cellulose synthase shows a more diffuse localization, while callose synthase shows a more frequent cytoplasmic accumulation, thereby denoting a failure in plasma membrane insertion. The cell wall observations are complemented by the analysis of intracellular Ca2+, pH and reactive oxygen species (ROS): while in the case of pH no major differences are observed, a less focused Ca2+ and ROS gradients are present in the stressed samples. The standard oscillatory growth of pollen tubes is recovered by transient changes of turgor pressure induced by hypoosmotic media. Overall our data contribute to the understanding of the impact that cold stress has on the normal development of the pollen tube and unveil the cell wall-related aberrant features accompanying the observed alterations.

A case of a malignant peripheral nerve sheath tumor in a guinea pig

Here, we describe the clinical and histopathological characteristics of a malignant peripheral nerve sheath tumor (MPNST) extending from the dorsal subcutis to the periphery of the spine in a female guinea pig aged 3 years 7 months. The patient presented with pleural and blood-like pericardial effusion and died. The tumor had invaded the spine and the surrounding muscles and had grown in hypercellular and hypocellular arrangements of round, broad-spindle, and elongated-spindle cells. We observed a fascicular growth pattern, nuclear palisading, and perivascular accumulations of cells that responded positively to anti-S100, sox10, and CD56 antibodies. This is the first report of a MPSNT in a guinea pig.

Size distribution of bioaerosols from biomass burning emissions: Characteristics of bacterial and fungal communities in submicron (PM1.0) and fine (PM2.5) particles

The North China Plain is the agricultural heartland in China. High PM2.5 levels and elevated chemical pollutants have been observed during crop harvest seasons due to open biomass burning. Biomass burning in the wheat-harvest season may significantly deteriorate the regional air quality. The harmful ingredients in smoke particles also have severe implications for toxicity and health effects. Previous studies have illustrated the potential role of bioaerosols as ice-nuclei and cloud condensation nuclei and highlighted their influence on biochemical cycles and human health effects. In a monthly field observation campaign of biomass burning conducted at the summit of Mount Tai in July 2015, we reported the composition, potential role, size distribution of microorganisms in particulate matters PM1.0, PM2.5, and estimated their contribution to particles. The wide-range particle spectrometer suggested that the predominant particles were distributed in submicron particles (PM1.0), which resulted in a similar community structure for bacteria and fungi in PM1.0 and PM2.5. Among bacteria, the predominant Pseudomonas accounted for 18.06% and 21.29% in PM1.0 and PM2.5, respectively. Alternaria covered up to 69.01% and 72.76% of the fungal community in PM1.0 and PM2.5, respectively. A disparity between bacterial communities was identified by the abundance of rare species, such as Bacilli being higher in PM1.0 (2.4%) than in PM2.5 (1.8%), and Defluviicoccus being higher in PM2.5 (2.5%) than in PM1.0 (0.5%), which may be related to cell size and cell growth patterns. Quantitative PCR revealed that microbial cell numbers in PM2.5 were higher than in PM1.0, and that the bacterial cell number was about an order of magnitude greater than the fungal cell number. However, the mass concentration and contribution of fungi to particulate matter was much higher than that of bacteria, suggesting the underestimated role of fungi in atmospheric aerosols. Airborne microorganisms in alpine areas remained less characterized. The findings presented here illustrated the potentially important impacts on air quality and bioaerosol pollution by biomass burning, which provides an essential reference for understanding the transmission and health effects of bioaerosols.

Synthesis, anticancer activity, toxicity evaluation and molecular docking studies of novel phenylaminopyrimidine—(thio)urea hybrids as potential kinase inhibitors

Thirty-two novel urea/thiourea compounds as potential kinase inhibitor were designed, synthesized and evaluated for their cytotoxic activity on breast (MCF7), colon (HCT116) and liver (Huh7) cancer cell lines. Compounds 10, 19 and 30 possessing anticancer activity with IC50 values of 0.9, 0.8 and 1.6 μM respectively on Huh7 cells were selected for further studies. These hit compounds were tested against liver carcinoma panel. Real time cell electronic sensing assay was used to evaluate the effects of the compounds 10, 19 and 30 on the growth pattern of liver cancer cells. Apoptotic cell death and cell cycle analysis upon treatment of liver carcinoma cells with hit compounds were determined. A significant apoptotic cell death was detected upon treatment of Huh7 and Mahlavu cells with compound 30 after 48 h of treatment. Additionally, compound 10 caused cell cycle arrest at G0/G1 phase. Mutagenicity of hit compounds was evaluated. Assertively, these compounds were not found to be mutagenic on Salmonella typhimurium strains TA98 and TA100. To understand the binding modes of the synthesized compounds, molecular docking studies were performed using the crystal data of VEGFR and Src-kinase enzymes in correlation with anticancer activities.

Effects of acrylate/acrylamide polymers on the adhesion, growth and differentiation of Muse cells

Background. Acrylate/acrylamide copolymers have excellent optical properties and biocompatibility and are ideal biomaterials that have been widely used in tissue engineering. Multilineage-differentiating stress-enduring cells (Muse cells) are a specific subset of mesenchymal stem cells that have an excellent potential for the regenerative medicine. Objective. This study was designed to investigate the effects of acrylate/acrylamide copolymers on the adhesion, proliferation and pluripotent-like properties of Muse cells, which were derived from normal human dermal fibroblasts by long-term trypsin incubation. Methods. In an initial experiment, Muse cells were seeded on primary microarrays containing micro-spots of 275 different mixtures of acrylate/acrylamide. Each mixture was composed of two of 11 different monomers in various proportions, and was replicated in four micro-spots each. According to the adhesion and growth characteristics of Muse cells on those substrates, specific polymer candidates for Muse cells were selected and secondary microarrays were prepared. We then observed the effects of those specific polymer candidates on the adherence, proliferation and differentiation of Muse cells and suitable candidates for their optimal culture were identified. Results. According to the adhesion and growth patterns of Muse cells on the primary microarrays, ten suitable mixtures of acrylate/acrylamide copolymers were identified. Muse cells grew well on six of those combinations and around the four other combinations of those polymer mixtures. Muse cells cultured on three of those combinations proliferated and differentiated into long spindle-shaped cells that looked like fibroblasts, while Muse cells cultured on one combination formed clusters that were ring-shaped. Muse cells cultured on some of those combinations of acrylate/acrylamide proliferated and formed clusters that appeared to be very healthy, whereas Muse cells cultured on other combinations formed clusters that expanded outwards. Conclusions. These results identified a polymer combination that was optimum for the adhesion, proliferation and maintenance of Muse cells in an undifferentiated state.

Implant abutment microgrooves affect soft tissue cells response via connexin 43 pathway

Background The down-regulation of Connexin 43 (CX43) hemichannels (HCs) function in soft tissue cells promotes the up-regulation of numerous pro-healing genes, as the down-regulation of pro-fibrotic ones, leading to its identification as a potential target for the modulation of wound healing. On the other hand, titanium is commonly used as a transmucosal material, both for dental implants and abutments, and its surface micro-topography is known to affect cell responses. Aim/Hypothesis To investigate whether threaded titanium surfaces with different groove characteristics modulate CX43 expression and its organization in HCs rather than in gap junction (GJs) in an immortalized mouse myoblast cell line. In addition, to verify if the growth pattern and arrangement of cells on the grooves. Material and Methods Murine myoblasts C2C12 were seeded on discs with grooved titanium surfaces with different patterns- UTM (Ultrathin Threaded Microsurface) and XA microgrooved surface. A machined surface was used as control (MA). Titanium surface specimens were kindly provided by Sweden & Martina (Padova, Italy). Cells were cultured up to 7 days and daily monitored for viability through chemiluminescence (cell proliferation). Gene expression analysis for CX43, alkaline phosphatase (ALP) and collagen 1 (COL1a) was performed+ after 3 days of culture total RNA was purified, retro-transcribed to cDNA and amplified through quantitative RT-PCR. To localize the expression of CX43 and thus to identify its organization as HCs or as GJs, cells were observed under fluorescence after actin, CX43 and nuclei labeling. Differences among the groups were evaluated through Two-Way ANOVA statistical test. Results Very weak signal for CX43 was observed on UTM surface (mRNA 6.12-fold less than XA and 2.58-fold less than MA). CX43 expression was extremely up-regulated on XA surface after 3 days, when compared to UTM and MA surface (P < 0.0001). Conversely, cell proliferation was significantly reduced (P < 0.0001). On the XA surface, CX43 expression was labelled in the peri-nuclear area, indicating its organization in HCs. MA showed a slightly enhanced level of CX43 (2.37-fold less than XA) and, where cells were confluent, CX43 signal was marked around cell nuclei (HCs) and at the edges of cell profile, indicating GJs with other cells. Noteworthy, the disposition of cells on XA and UTM surfaces followed the grid of the grooved surface topography, with cells laying on the grid hollows and elongated to colonize the surface. Furthermore, gene expression analysis revealed that MA control surface induced a commitment of C2C12 towards an osteoblastic phenotype, showing greater levels of ALP and COL1a. Conclusions and Clinical Implications The microgrooved titanium surfaces showed a modulatory role on myoblasts, promoting an evident cell arrangement along the surface grooves on UTM and XA surfaces. This may lead in vivo to the deposition of an oriented extracellular matrix. Additionally, UTM surface caused CX43 down-regulation, leading to the hypothesis of a more favorable in vivo soft tissues wound healing.

Enhancer function regulated by combinations of transcription factors and cofactors.

Regulation of the expression of diverse genes is essential for making possible the complexity of higher organisms, and the temporal and spatial regulation of gene expression allows for the alteration of cell types and growth patterns. A critical component of this regulation is the DNA sequence-specific binding of transcription factors (TFs). However, most TFs do not independently participate in gene transcriptional regulation, because they lack an effector function. Instead, TFs are thought to work by recruiting cofactors, including Mediator complex (Mediator), chromatin-remodeling complexes (CRCs), and histone-modifying complexes (HMCs). Mediator associates with the majority of transcribed genes and acts as an integrator of multiple signals. On the other hand, CRCs and HMCs are selectively recruited by TFs. Although all the pairings between TFs and CRCs or HMCs are not fully known, there are a growing number of established TF-CRC and TF-HMC combinations. In this review, we focused on the most important of these pairings and discuss how they control gene expression.