Changes In Tissue Architecture Research Articles

Biological Characterization of Polymeric Matrix and Graphene Oxide Biocomposites Filaments for Biomedical Implant Applications: A Preliminary Report.

Carbon nanostructures application, such as graphene (Gr) and graphene oxide (GO), provides suitable efforts for new material acquirement in biomedical areas. By aiming to combine the unique physicochemical properties of GO to Poly L-lactic acid (PLLA), PLLA-GO filaments were produced and characterized by X-ray diffraction (XRD). The in vivo biocompatibility of these nanocomposites was performed by subcutaneous and intramuscular implantation in adult Wistar rats. Evaluation of the implantation inflammatory response (21 days) and mesenchymal stem cells (MSCs) with PLLA-GO took place in culture for 7 days. Through XRD, new crystallographic planes were formed by mixing GO with PLLA (PLLA-GO). Using macroscopic analysis, GO implanted in the subcutaneous region showed particles’ organization, forming a structure similar to a ribbon, without tissue invasion. Histologically, no tissue architecture changes were observed, and PLLA-GO cell adhesion was demonstrated by scanning electron microscopy (SEM). Finally, PLLA-GO nanocomposites showed promising results due to the in vivo biocompatibility test, which demonstrated effective integration and absence of inflammation after 21 days of implantation. These results indicate the future use of PLLA-GO nanocomposites as a new effort for tissue engineering (TE) application, although further analysis is required to evaluate their proliferative capacity and viability.

Interplay Between Notch and YAP/TAZ Pathways in the Regulation of Cell Fate During Embryo Development.

Cells in growing tissues receive both biochemical and physical cues from their microenvironment. Growing evidence has shown that mechanical signals are fundamental regulators of cell behavior. However, how physical properties of the microenvironment are transduced into critical cell behaviors, such as proliferation, progenitor maintenance, or differentiation during development, is still poorly understood. The transcriptional co-activators YAP/TAZ shuttle between the cytoplasm and the nucleus in response to multiple inputs and have emerged as important regulators of tissue growth and regeneration. YAP/TAZ sense and transduce physical cues, such as those from the extracellular matrix or the actomyosin cytoskeleton, to regulate gene expression, thus allowing them to function as gatekeepers of progenitor behavior in several developmental contexts. The Notch pathway is a key signaling pathway that controls binary cell fate decisions through cell–cell communication in a context-dependent manner. Recent reports now suggest that the crosstalk between these two pathways is critical for maintaining the balance between progenitor maintenance and cell differentiation in different tissues. How this crosstalk integrates with morphogenesis and changes in tissue architecture during development is still an open question. Here, we discuss how progenitor cell proliferation, specification, and differentiation are coordinated with morphogenesis to construct a functional organ. We will pay special attention to the interplay between YAP/TAZ and Notch signaling pathways in determining cell fate decisions and discuss whether this represents a general mechanism of regulating cell fate during development. We will focus on research carried out in vertebrate embryos that demonstrate the important roles of mechanical cues in stem cell biology and discuss future challenges.

Hesperidin and its aglycone hesperetin in breast cancer therapy: A review of recent developments and future prospects.

Breast cancer (BC) has high incidence and mortality rates, making it a major global health issue. BC treatment has been challenging due to the presence of drug resistance and the limited availability of therapeutic options for triple-negative and metastatic BC, thereby urging the exploration of more effective anti-cancer agents. Hesperidin and its aglycone hesperetin, two flavonoids from citrus species, have been extensively evaluated for their anti-cancer potentials. In this review, available literatures on the chemotherapeutic and chemosensitising activities of hesperidin and hesperetin in preclinical BC models are reported. The safety and bioavailability of hesperidin and hesperetin as well as the strategies to enhance their bioavailability are also discussed. Overall, hesperidin and hesperetin can inhibit cell proliferation, migration and BC stem cells as well as induce apoptosis and cell cycle arrest in vitro. They can also inhibit tumour growth, metastasis and neoplastic changes in tissue architecture in vivo. Moreover, the co-administration of hesperidin or hesperetin with doxorubicin, letrozole or tamoxifen can enhance the efficacies of these clinically available agents. These chemotherapeutic and chemosensitising activities of hesperidin and hesperetin have been linked to several mechanisms, including the modulation of signalling pathways, glucose uptake, enzymes, miRNA expression, oxidative status, cell cycle regulatory proteins, tumour suppressor p53, plasma and liver lipid profiles as well as DNA repair mechanisms. However, poor water solubility, extensive phase II metabolism and apical efflux have posed limitations to the bioavailability of hesperidin and hesperetin. Various strategies for bioavailability enhancement have been studied, including the utilisation of nano-based drug delivery systems and the co-administration of hesperetin with other flavonoids. In particular, nanoformulated hesperidin and hesperetin possess greater chemotherapeutic and chemosensitising activities than free compounds. Despite promising preclinical results, further safety and efficacy evaluation of hesperidin and hesperetin as well as their nanoformulations in clinical trials is required to ascertain their potentials to be developed as clinically useful agents for BC treatment.

Impact of switching to raltegravir and/or adding losartan in lymphoid tissue fibrosis and inflammation in people living with HIV. A randomized clinical trial.

Persistent inflammation and immune activation are associated with lymph node fibrosis and end-organ diseases in treatment-suppressed people living with HIV (PLWH). We investigated the effect of switching to raltegravir and/or adding losartan on lymphoid tissue fibrosis and on the inflammatory/immune-activation mediators in treated HIV patients. Chronic HIV-infected patients treated with two nucleoside reverse transcriptase inhibitors (2NRTI) and one non-NRTI (NNRTI) or protease inhibitor (PI) during at least 48weeks were randomized to four groups (n=48): 2NRTI+efavirenz (EFV), 2NRTI+EFV+losartan, 2NRTI+raltegravir and 2NRTI+raltegravir+losartan for 48weeks. Tonsillar biopsy and peripheral blood markers of CD4 and CD8T-lymphocyte activation and senescence, monocyte activation and soluble markers of inflammation were determined at baseline and at week 48 and compared between groups. No changes in lymphoid tissue architecture were observed. Adding losartan had no impact on lymphocyte subsets. Conversely, patients who switched to raltegravir showed a higher decrease in all activated [CD4+CD38+HLA-DR+, -0.3 vs. 0.48 (P=0.033); CD8+CD38+HLA-DR+, -1.6 vs. 1.3 (P=0.02)] and senescent [CD4+CD28-CD57+, -0.3 vs. 0.26 (P=0.04); CD8+CD28-CD57+, -6.1 vs. 3.8 (P=0.002)] T lymphocytes. In addition, the median CD4/CD8 ratio increased by 0.35 in patients in the raltegravir group vs. 0.03 in the other arms (P=0.002). Differences between groups in monocyte subpopulations or soluble inflammation markers were not observed. Losartan had no effect on lymphoid fibrosis or immune activation/inflammation. Conversely, switching to a regimen with raltegravir significantly decreased activated and senescent T-lymphocyte subpopulations and increased CD4/CD8 ratio in successfully treated PLWH.

A Straightforward Method for 3D Visualization of B Cell Clusters and High Endothelial Venules in Lymph Nodes Highlights Differential Roles of TNFRI and -II

Whole mount tissue immunolabeling and imaging of complete organs has tremendous benefits in characterizing organ morphology. Here, we present a straightforward method for immunostaining, clearing and imaging of whole murine peripheral lymph nodes (PLNs) for detailed analysis of their architecture and discuss all procedures in detail in a step-by-step approach. Given the importance of tumor necrosis factor receptor (TNFR) signaling in development of PLNs we used TNFRI-/- and TNFRII-/- mice models as proof-of-concept for this technique by visualizing and analyzing structural changes in PLN B cell clusters and high endothelial venules (HEVs). Samples were subjected to de- and rehydration with methanol, labeled with antibodies for B cells, T cells and high endothelial venules (HEVs) and optically cleared using benzyl alcohol-benzyl benzoate. Imaging was done using LaVision light sheet microscope and analysis with Imaris software. Using these techniques, we confirmed previous findings that TNFRI signaling is essential for formation of individual B cell clusters. In addition, Our data suggest that TNFRII signaling is also to some extent involved in this process as TNFRII-/- PLNs had a B cell cluster morphology reminiscent of TNFRI-/- PLNs. Moreover, visualization and objective quantification of the complete PLN high endothelial vasculature unveiled reduced volume, length and branching points of HEVs in TNFRI-/- PLNs, revealing an earlier unrecognized contribution of TNFRI signaling in HEV morphology. Together, these results underline the potential of whole mount tissue staining and advanced imaging techniques to unravel even subtle changes in lymphoid tissue architecture.

TISSUE NA+ EXCESS UPON EXPERIMENTAL SALT-LOADING IS SYSTEMIC AND REFLECTS ISOTONIC OEDEMA

Objective: Skin was identified as a large depot for Na+ accumulation. Its proposed water-independent (hypertonic) nature has the potential to induce local pathogenic changes, including activation of the osmotic stress sensor TonEBP-mediated lymphangiogenic signalling. Blockade of this response led to salt-sensitive hypertension. Our study tested the hypertonic nature and organ-specific distribution of tissue Na+ accumulation upon salt-loading. Design and method: Twelve weeks-old male and female stroke-prone spontaneously hypertensive (SHRSP) and control Wistar-Kyoto (WKY) rats were treated with 1% NaCl in drinking water (high salt, HS) or normal water (NS) for 3 weeks (n = 8–10/sex/group). Tissue water and Na+/K+ content, as well as TonEBP gene expression in abdominal skin, liver, lung, myocardium and skeletal muscle were measured by gravimetric approach (wet–dry weight), flame photometry and qRT-PCR, respectively. The functional effects of in-vivo and ex-vivo exposure of resistance arteries to salt excess (HS treatment and incubation in +15 mM NaCl, respectively) were assessed and compared by wire myography. Results: Tissue Na+ increased in skin (+20.7%), but also liver (+10.3%), lungs (+14.8%) and skeletal muscle (+23.6%) in HS-SHRSP compared to NS; apart from skeletal muscle, it was consistently paralleled by water (skin: +14.1%; liver: +4.8%; lungs: +9.0%; p < 0.05 for all). Patterns were similar between males and females and absent in WKY. In both strains and all organs, K+ + Na+ concentration was not affected by HS and sat within physiological limits, thus pointing against hypertonicity. Nevertheless, TonEBP gene expression changes matched Na+ and water isotonic accumulation in SHRSP-HS skin, liver and lungs. Hypertonic incubation did not induce hypercontractile responses but exacerbated the sensitivity to NO-mediated relaxation in arteries from both strains (δIC50, p < 0.001). The effect of in-vivo HS, in keeping with classic hypertensive vascular changes, was opposite (p < 0.001, in SHRSP only). Conclusions: If water independent, interstitial Na accumulation is unlikely to underpin the development of experimental hypertension via a direct impact on vascular function. Tissue Na+ excess upon HS rather reflects isotonic oedema accumulation and systemic changes in tissue architecture, not solely confined to skin, and sensed by TonEBP.

Ploidy influences wheat mesophyll cell geometry, packing and leaf function.

Leaf function is influenced by leaf structure, which is itself related not only to the spatial arrangement of constituent mesophyll cells, but also their size and shape. In this study, we used confocal microscopy to image leaves of Triticum genotypes varying in ploidy level to extract 3D information on individual mesophyll cell size and geometry. Combined with X‐ray Computed Tomography and gas exchange analysis, the effect of changes in wheat mesophyll cell geometry upon leaf structure and function were investigated. Mesophyll cell size and shape were found to have changed during the course of wheat evolution. An unexpected linear relationship between mesophyll cell surface area and volume was discovered, suggesting anisotropic scaling of mesophyll cell geometry with increasing ploidy. Altered mesophyll cell size and shape were demonstrated to be associated with changes in mesophyll tissue architecture. Under experimental growth conditions, CO2 assimilation did not vary with ploidy, but stomatal conductance was lower in hexaploid plants, conferring a greater instantaneous water‐use efficiency. We propose that as wheat mesophyll cells have become larger with increased ploidy, this has been accompanied by changes in cell geometry and packing which limit water loss while maintaining carbon assimilation.

Chronic exposure of bisphenol S (BPS) affect hypothalamic-pituitary-testicular activities in adult male rats: possible in estrogenic mode of action

BackgroundThe industrial revolution has resulted in increased synthesis and the introduction of a variety of compounds into the environment and their potentially hazardous effects have been observed in the biota. The present study was aimed to evaluate the potential endocrine-disrupting effects of chronic exposure to the low concentrations of bisphenol S (BPS) in male rats.MethodsWeaning male Sprague-Dawley rats (22 days old) were either exposed to water containing 0.1% ethanol for control or different concentrations of BPS (0.5, 5, and 50 μg/L) in drinking water for 48 weeks in the chronic exposure study. After completion of the experimental period, animals were dissected and different parameters (hormone concentrations, histology of testis and epididymis, oxidative stress and level of antioxidant enzymes in the testis, daily sperm production (DSP), and sperm parameters) were determined.ResultsResults of the present study showed a significant alteration in the gonadosomatic index (GSI) and relative reproductive organ weights. Oxidative stress in the testis was significantly elevated while sperm motility, daily sperm production, and the number of sperm in epididymis were reduced. Plasma testosterone, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) concentrations were reduced and estradiol levels were high in the 50 μg/L-exposed group. Histological observations involved a significant reduction in the epithelial height of the testis along with disrupted spermatogenesis, an empty lumen of the seminiferous tubules, and the caput region of the epididymis.ConclusionThese results suggest that exposure to 5 and 50 μg/L of BPS for the chronic duration started from an early age can induce structural changes in testicular tissue architecture and endocrine alterations in the male reproductive system which may lead to infertility in males.

Self‐Assembling Nanofibers Inhibit Inflammation in a Murine Model of Crohn's‐Disease‐Like Ileitis

AbstractInflammatory bowel diseases are characterized by chronic inflammation, leading to the destruction of gastrointestinal tract tissue. Strategies specifically utilized to manage intestinal inflammation in patients afflicted with Crohn's disease (CD) are met with numerous obstacles. These include treatment‐specific physiological side effects and the development of resistance to biological and pharmacological therapies. As an alternative treatment approach, high aspect ratio supramolecular anti‐inflammatory peptide amphiphiles (AIF‐PAs) are engineered to mitigate small intestinal inflammation within the established, preclinical CD‐like ilieitis senescence accelerated mouse (SAMP1)/YitFcsJ murine model. Data derived from this study demonstrate ileitis that the direct injection of AIF‐PAs into inflammatory small intestinal skip lesions minimizes pathologic change in tissue architecture and intestinal physiology. When compared to untreated or saline injected control animals, AIF‐PAs substantially reduce lesion size and tissue‐specific inflammation, inciting cytokines tumor necrosis factor alpha (TNFα), interferon gamma (IFNγ), interleukin (IL)‐1β, and IL‐13 along with a concomitant decrease including in tissue proinflammatory innate and adaptive immune cells. Finally, this is accompanied by increased small bowel intestinal transit that aligns with improved gross intestinal pathology scoring. Taken together, this study provides evidence that AIF‐PAs may serve as a novel stand‐alone or combinatorial therapy to enhance efficacy in order to treat localized intestinal inflammation related to CD.

A modifier screen identifies regulators of cytoskeletal architecture as mediators of Shroom-dependent changes in tissue morphology.

ABSTRACTRegulation of cell architecture is critical in the formation of tissues during animal development. The mechanisms that control cell shape must be both dynamic and stable in order to establish and maintain the correct cellular organization. Previous work has identified Shroom family proteins as essential regulators of cell morphology during vertebrate development. Shroom proteins regulate cell architecture by directing the subcellular distribution and activation of Rho-kinase, which results in the localized activation of non-muscle myosin II. Because the Shroom-Rock-myosin II module is conserved in most animal model systems, we have utilized Drosophila melanogaster to further investigate the pathways and components that are required for Shroom to define cell shape and tissue architecture. Using a phenotype-based heterozygous F1 genetic screen for modifiers of Shroom activity, we identified several cytoskeletal and signaling protein that may cooperate with Shroom. We show that two of these proteins, Enabled and Short stop, are required for ShroomA-induced changes in tissue morphology and are apically enriched in response to Shroom expression. While the recruitment of Ena is necessary, it is not sufficient to redefine cell morphology. Additionally, this requirement for Ena appears to be context dependent, as a variant of Shroom that is apically localized, binds to Rock, but lacks the Ena binding site, is still capable of inducing changes in tissue architecture. These data point to important cellular pathways that may regulate contractility or facilitate Shroom-mediated changes in cell and tissue morphology.

Pulmonary arteries of Williams syndrome patients exhibit altered serotonin metabolism genes and degenerated medial layer architecture.

Williams-Beuren syndrome (WS) is characterized by cardiovascular abnormalities associated with a multigene deletion on 7q11.23, in particular elastin (ELN). Peripheral pulmonary artery stenosis (PPAS) frequently affects pediatric patients with WS. Molecular investigation of WS pulmonary arterial (PA) tissue is limited by tissue scarcity. We compared transcriptomes, tissue architecture, and localized changes in protein expression in PA tissue from patients with WS (n = 8) and donors (n = 5). Over 100 genes were differentially expressed at the ≥4-fold level, including genes related to the serotonin signaling pathway: >60-fold downregulation of serotonin transporter SLC6A4 and >3-fold upregulation of serotonin receptor HTR2A. Histologic examination revealed abnormal elastin distribution and smooth muscle cell morphology in WS PA, with markedly shorter, disorganized elastin fibers, and expanded proteoglycan-rich extracellular matrix between muscle layers. There were significant abnormalities in the PA expression of genes regulating serotonin signaling, metabolism, and receptors in WS. Those changes were associated with distinct changes in the arterial structure and may play a role in the stenosis-promoting effects of elevated shear stress at PA bifurcations in WS. Serotonin pathway signaling is significantly altered in the pulmonary arteries of patients with Williams syndrome and severe peripheral arterial stenosis. The present study compares the histological and biochemical characteristics of pulmonary arteries from patients with Williams syndrome to those of controls, something that has not, to our knowledge, been done previously. It demonstrates marked abnormalities in the pulmonary arteries of patients with Williams syndrome, especially significant pathologic alterations in the signaling of the serotonin pathway. The findings of this study provide direction for the development of potential therapies to treat pulmonary artery stenosis in patients with Williams syndrome.

Engineering the Extracellular Matrix to Model the Evolving Tumor Microenvironment.

SummaryClinical evidence supports a role for the extracellular matrix (ECM) in cancer risk and prognosis across multiple tumor types, and numerous studies have demonstrated that individual ECM components impact key hallmarks of tumor progression (e.g., proliferation, migration, angiogenesis). However, the ECM is a complex network of fibrillar proteins, glycoproteins, and proteoglycans that undergoes dramatic changes in composition and organization during tumor development. In this review, we will highlight how engineering approaches can be used to examine the impact of changes in tissue architecture, ECM composition (i.e., identity and levels of individual ECM components), and cellular- and tissue-level mechanics on tumor progression. In addition, we will discuss recently developed methods to model the ECM that have not yet been applied to the study of cancer.

Subacute Hepatotoxicity of Extracts of Senna occidentalis Seeds in Swiss Albino Mice.

Senna occidentalis is potentially toxic to humans and animals. Its seeds are crop contaminant weeds in some localities where liver disease is prevalent. This study assessed the subacute hepatotoxicity of S. occidentalis seeds in mice model. Three groups of female Swiss Albino mice (25–28 g, aged 8–10 weeks) received distilled water (control), 400, and 1000 mg/kg extract of S. occidentalis seed, respectively. At the end of the study, body weight and liver organ weight were recorded, and tissue and blood samples were collected and analyzed. The results indicated that the extract treated groups, at both doses, showed significant (p ≤ 0.001) decrease in mean body weight gain in the fourth week of the experiment. Besides, the extract treated groups showed significant (p ≤ 0.001) elevation of liver enzyme markers: alanine aminotransferase and aspartate aminotransferase. Also, histopathological examinations of liver tissue showed moderate microvesicular steatosis of hepatocytes and mild inflammation in the 400 mg/kg treated group as well as marked micro- and macrovesicular steatosis, focal area necrosis, and periportal inflammation with mononuclear cell infiltration in the 1000 mg/kg treated group. Thus, these findings show that S. occidentalis seeds exhibit hepatotoxicity in mice, characterized by changes in liver tissue architecture and liver enzyme levels.

Knee Osteoarthritis: A Review of Pathogenesis and State-Of-The-Art Non-Operative Therapeutic Considerations.

Being the most common musculoskeletal progressive condition, osteoarthritis is an interesting target for research. It is estimated that the prevalence of knee osteoarthritis (OA) among adults 60 years of age or older is approximately 10% in men and 13% in women, making knee OA one of the leading causes of disability in elderly population. Today, we know that osteoarthritis is not a disease characterized by loss of cartilage due to mechanical loading only, but a condition that affects all of the tissues in the joint, causing detectable changes in tissue architecture, its metabolism and function. All of these changes are mediated by a complex and not yet fully researched interplay of proinflammatory and anti-inflammatory cytokines, chemokines, growth factors and adipokines, all of which can be measured in the serum, synovium and histological samples, potentially serving as biomarkers of disease stage and progression. Another key aspect of disease progression is the epigenome that regulates all the genetic expression through DNA methylation, histone modifications, and mRNA interference. A lot of work has been put into developing non-surgical treatment options to slow down the natural course of osteoarthritis to postpone, or maybe even replace extensive surgeries such as total knee arthroplasty. At the moment, biological treatments such as platelet-rich plasma, bone marrow mesenchymal stem cells and autologous microfragmented adipose tissue containing stromal vascular fraction are ordinarily used. Furthermore, the latter two mentioned cell-based treatment options seem to be the only methods so far that increase the quality of cartilage in osteoarthritis patients. Yet, in the future, gene therapy could potentially become an option for orthopedic patients. In the following review, we summarized all of the latest and most important research in basic sciences, pathogenesis, and non-operative treatment.

Quantitative analysis of astrocyte and axonal density relationships: Glia to neuron ratio in the optic nerve laminar regions

Astrocytes are critical for the maintenance of retinal ganglion cell (RGC) axonal function and viability, and form a key component of the functional neurovascular unit. Recently, we described the quantitative properties of astrocytes in relation to the capillary distributions in optic nerve laminar regions. Here, we provide a quantitative analysis of astrocytes and RGC axons in longitudinal sections of optic nerve tissue. Histological and immunocytochemical techniques are used to demonstrate the density of astrocytes, RGC axons and glia-neuron ratios across the pre laminar, lamina cribrosa and post laminar compartments of the optic nerve head (ONH). A study of human, pig, horse and rat optic nerves was performed and comparisons are made between species. This study demonstrates that the distribution of astrocytes correlates closely with the density of axonal processes, in accordance with the functional requirement of different regions of the ganglion cell axon. There was a consistency of glia-neuron ratios in the majority of laminar compartments, except for the human and rat prelaminar regions, which demonstrated lower ratios of astrocyte to axonal processes. The distribution of astrocytes may reflect a functional susceptibility to development of disease in the prelaminar region of the optic nerve. Interspecies comparison at the lamina cribrosa showed strikingly consistent glia-neuron ratios. Collectively, our findings suggest there may be a critical ratio of glia to neuron needed to maintain healthy cellular physiology across different laminar compartments of the optic nerve, with particular importance for the health of the lamina cribrosa region. It is possible that, in disease processes, the glia-neuron relationships across the different laminar compartments may be perturbed and this may be relevant for the development of glaucoma. Emerging technologies may further aid our understanding in how the physiology of optic nerve tissue cellular structure may be affected by changes to ONH characteristics and elevated intraocular pressure induced damage. Such findings may also permit the early identification of RGC axonal injury by identifying quantifiable changes in structural tissue architecture when pathophysiological pathways predominate.

Abstract C029: Consumption of dietary AGEs during puberty and increased breast cancer risk: A link between lifestyle and cancer disparity

Abstract Introduction. The focus of this study is on early lifestyle factors and their effect on mammary development during puberty and how they relate to increased breast cancer risk and disparities. At this time we do not understand what biological changes occur during pubertal mammary development which leads to a greater risk of developing cancer in later life. Identifying the molecular mechanisms that cause aberrant pubertal mammary development may lead to defined strategies to reduce breast cancer burden in later life. As our bodies use the sugars that we consume for energy they generate waste chemicals known as advanced glycation end products or AGEs for short. Significantly, low income, obesity and a sedentary lifestyle are established factors driving health disparity that also contribute to increased AGE accumulation levels in our bodies. In particular, AGE content in the Western Diet has consistently increased over the last 50 years due to increased consumption of sugar-laden and cheap processed/manufactured foods which are high in reactive AGE metabolites and can promote obesity. Methods. We use a dietary mouse model to assess impact of AGE on normal mammary development. Wild type FVB/n and RAGE null (RAGE-/-) mice are fed the respective diets from weaning until 7 (pubertal) or 12 (adult) weeks of age. Mammary glands are extracted for whole mounting and paraffin embedded for histology. Fibroblasts were isolated from mammary glands and cultured ex vivo. Transwell migration assays were performed with isolated fibroblasts and HC11 mouse mammary epithelial cells. qPCR was performed on the isolated fibroblasts to assess their activation status. Results. Early life exposures during mammary development influence the breast microenvironment to increase breast cancer risk. We show that due to an innate ability to influence the cellular matrix, dietary AGEs disrupt developmental programs during puberty and promote breast tumor growth. Through receptor for AGE (RAGE) dependent and independent mechanisms, chronic AGE consumption delayed ductal extension, increased ductal branching and caused aberrant terminal end bud (TEB) morphology. Dietary AGE activation of RAGE mediated a program of activated stroma leading to hyperplastic growth and the formation of pre-neoplastic lesions which persisted into adulthood. Importantly, AGE mediated effects remained even after diet intervention after puberty. In dietary-AGE breast tumor models, AGE mediated changes in tissue architecture and cell function were recapitulated and resulted in 3-fold increase in neoplastic growth. Through the perpetual activation of a reactive stroma, AGEs derived from diet represent a common early life exposure which can influence tumor behavior. Conclusions. A greater mechanistic understanding of the link between AGE intake during puberty and increased breast cancer risk may define novel potential strategies for lifestyle and pharmacological intervention aimed at reducing breast cancer risk and cancer disparities. Citation Format: Callan C Frye, Bradley A Krisanits, Reid Schuster, Jaime Randise, Lourdes M Nogueira, Kristi Helke, Amanda C LaRue, David P Turner, Victoria J Findlay. Consumption of dietary AGEs during puberty and increased breast cancer risk: A link between lifestyle and cancer disparity [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr C029.

Tissue Mechanics Regulate Mitotic Nuclear Dynamics during Epithelial Development

SummaryCell divisions are essential for tissue growth. In pseudostratified epithelia, where nuclei are staggered across the tissue, each nucleus migrates apically before undergoing mitosis. Successful apical nuclear migration is critical for planar-orientated cell divisions in densely packed epithelia. Most previous investigations have focused on the local cellular mechanisms controlling nuclear migration. Inter-species and inter-organ comparisons of different pseudostratified epithelia suggest global tissue architecture may influence nuclear dynamics, but the underlying mechanisms remain elusive. Here, we use the developing Drosophila wing disc to systematically investigate, in a single epithelial type, how changes in tissue architecture during growth influence mitotic nuclear migration. We observe distinct nuclear dynamics at discrete developmental stages, as epithelial morphology changes. We use genetic and physical perturbations to show a direct effect of cell density on mitotic nuclear positioning. We find Rho kinase and Diaphanous, which facilitate mitotic cell rounding in confined cell conditions, are essential for efficient apical nuclear movement. Perturbation of Diaphanous causes increasing defects in apical nuclear migration as the tissue grows and cell density increases, and these defects can be reversed by acute physical reduction of cell density. Our findings reveal how the mechanical environment imposed on cells within a tissue alters the molecular and cellular mechanisms adopted by single cells for mitosis.

Nanoparticles as Versatile Tools for Mechanotransduction in Tissues and Organoids.

Organoids are 3D multicellular constructs that rely on self-organized cell differentiation, patterning and morphogenesis to recapitulate key features of the form and function of tissues and organs of interest. Dynamic changes in these systems are orchestrated by biochemical and mechanical microenvironments, which can be engineered and manipulated to probe their role in developmental and disease mechanisms. In particular, the in vitro investigation of mechanical cues has been the focus of recent research, where mechanical manipulations imparting local as well as large-scale mechanical stresses aim to mimic in vivo tissue deformations which occur through proliferation, folding, invagination, and elongation. However, current in vitro approaches largely impose homogeneous mechanical changes via a host matrix and lack the required positional and directional specificity to mimic the diversity of in vivo scenarios. Thus, while organoids exhibit limited aspects of in vivo morphogenetic events, how local forces are coordinated to enable large-scale changes in tissue architecture remains a difficult question to address using current techniques. Nanoparticles, through their efficient internalization by cells and dispersion through extracellular matrices, have the ability to provide local or global, as well as passive or active modulation of mechanical stresses on organoids and tissues. In this review, we explore how nanoparticles can be used to manipulate matrix and tissue mechanics, and highlight their potential as tools for fate regulation through mechanotransduction in multicellular model systems.

Multidimensional Coculture System to Model Lung Squamous Carcinoma Progression.

Tumor-stroma interactions play a critical role in the development of lung squamous carcinoma (LUSC). However, understanding how these dynamic interactions contribute to tissue architectural changes observed during tumorigenesis remains challenging due to the lack of appropriate models. In this protocol, we describe the generation of a 3D coculture model using a LUSC primary cell culture known as TUM622. TUM622 cells were established from a LUSC patient-derived xenograft (PDX) and have the unique property to form acinar-like structures when seeded in a basement membrane matrix. We demonstrate that TUM622 acini in 3D coculture recapitulate key features of tissue architecture during LUSC progression as well as the dynamic interactions between LUSC cells and components of the tumor microenvironment (TME), including the extracellular matrix (ECM) and cancer-associated fibroblasts (CAFs). We further adapt our principal 3D culturing protocol to demonstrate how this system could be utilized for various downstream analyses. Overall, this organoid model creates a biologically rich and adaptable platform that enables one to gain insight into the cell-intrinsic and extrinsic mechanisms that promote the disruption of epithelial architectures during carcinoma progression and will aid the search for new therapeutic targets and diagnostic markers.

Phasor-FLIM Quantification of Changes in Keratinocyte Metabolism and Tissue Architecture in a Longitudinal Study of UV-induced Skin Cancer

Two of the recognized hallmarks of cancer include fundamental shifts in energy metabolism toward a glycolytic phenotype and a reorganization of tissue architecture to enable tumor cell invasion and metastasis. Phasor-FLIM imaging of NAD(P)H and flavoproteins enables a rapid assessment of changes in metabolism, while fluorescence and scattering from structural proteins reveal changes in tissue architecture. Here, we demonstrate the practicality of conducting a long-term longitudinal study, monitoring the development and progression of UV-induced skin cancer in SKH1 mice. Linear Mixed Model Analysis of the entire dataset of more than 2000 repeated observations reveals statistically significant changes in bound NADH fraction as well as the degree of collagen fluorescence. Recent developments, including modifications to the instrument design to include a three-color, macro-fluorescence imaging mode to guide the selection of suspect regions will be discussed. This publication was made possible by grants from the National Institute for General Medical Science (NIGMS) (5P20GM103427), a component of the National Institutes of Health (NIH), and its contents are the sole responsibility of the authors and do not necessarily represent the official views of NIGMS or NIH.