Cell-extracellular Matrix Communication Research Articles

Diabetes induces modifications in costameric proteins and increases cardiomyocyte stiffness.

Several studies have demonstrated that Diabetes mellitus can increase the risk of cardiovascular disease and remains the principal cause of death in these patients. Costameres connect the sarcolemma with the cytoskeleton and extracellular matrix, facilitating the transmission of mechanical forces and cell signaling. They are related to cardiac physiology because individual cardiac cells are connected by intercalated discs that synchronize muscle contraction. Diabetes impacts the nano-mechanical properties of cardiomyocytes, resulting in increased cellular and left ventricular stiffness, as evidenced in clinical studies of these patients. The question of whether costameric proteins are affected by diabetes in the heart has not been studied. This work analyzes whether T1DM modifies the costameric proteins and coincidentally changes the cellular mechanics in the same cardiomyocytes. The samples were analyzed by immunotechniques using laser confocal microscopy. Significant statistical differences were found in the spatial arrangement of the costameric proteins. However, these differences are not due to their expression. Atomic force microscopy was used to compare intrinsic cellular stiffness between diabetic and normal cardiomyocytes and obtain the first elasticity map sections of diabetic living cardiomyocytes. Data obtained demonstrated that diabetic cardiomyocytes had higher stiffness than control. The present work shows experimental evidence that intracellular changes related to cell-cell and cell-extracellular matrix communication occur, which could be related to cardiac pathogenic mechanisms. These changes could contribute to alterations in the mechanical and electrical properties of cardiomyocytes and consequently, to diabetic cardiomyopathy.

Comparison of primordial germ cell differences at different developmental time points in chickens.

Recently, the application in the field of germplasm resource conservation has become an important application of primordial germ cells (PGCs). However, due to the lack of deep understanding of the biological characteristics of PGCs at different time points, there is no systematic scheme for the selection of PGCs at which time points in practical application, which affects the practical application effect of PGCs. This study aims to clarify the differences in PGCs during development. Here, migration experiment, EdU proliferation assay and cell apoptosis assay were conducted to compare the differences in the migration ability, the proliferation ability and the recovery efficiency among female and male PGCs at E3.5, E4.5 and E5.5, which were explained by the following transcriptome sequencing analysis. We found that there were larger differences between female and male PGCs at different embryonic ages, while smaller differences between female and male PGCs at the same embryonic age. Further comparison showed that the cell migration ability of female and male PGCs decreased gradually during development, so female and male PGCs at E3.5 are more suitable for in vitro allotransplantation. At the same time, the proliferation ability of PGCs gradually decreased during development, and cell adhesion and extracellular matrix communication were weakened, indicating that female and male PGCs of E3.5 are more suitable for in vitro long-term culture cell line establishment. Interestingly, female and male PGCs at E5.5 showed strong DNA damage repair ability, thus more suitable for in vitro long-term cryopreservation. This study provides a theoretical basis for systematically selecting PGCs at suitable developmental time points as cell materials for efficient utilization by analyzing the characteristics of female and male PGCs at different developmental time points based on transcriptome.

MECHANICALLY INSTRUCTIVE SCAFFOLDS TO STEER TISSUE REGENERATION: MERGING MECHANOBIOLOGY WITH BIOFABRICATION

Regenerative medicine (RM) promises to restore both the mechanical functionality and the biological composition of tissues after damage. Three-dimensional scaffolds are used in RM to host cells and let them produce proteins that are the building blocks of the native tissues. While regenerating tissues evolve over time through dynamic biomechanical and biochemical changes, current scaffolds’ generation are passive causing mechanical mismatch, suboptimal growth, and pain. Furthermore, current scaffolds ignore the complexity of the reciprocal bio-mechanics regulation, hindering the design of the next-gen scaffolds. To regenerate tissues and organs, biofabrication strategies that impart spatiotemporal control over cell-cell and cell-extracellular matrix communication, often through control over cell and material deposition and placement, are being developed. To achieve these targets, the spatiotemporal control over biological signals at the interface between cells and materials is often aimed for. Alternatively, biological activity can be triggered through the control of mechanical cues, harnessing more fundamental know-how in mechanobiology that could be combined with biofabrication strategies. Here, I present some of our most recent advancements in merging mechanobiology with biofabrication that enabled the control of cell activity, moving towards enhanced tissue regeneration as well as the possibility to create more complex 3D in vitro models to study biological processes.

#136 : Osteopontin Facilitates Embryo Growth, Implantation, and Cellular Metabolism via Interacting with αvβ3 and RGS2 Pathway: Evidence from Mouse Embryos and a 3D In-Vitro Implantation Model

Background and Aims: We conducted a study on mice to investigate how osteopontin, a mediator of cell-cell and cell-extracellular matrix (ECM) communication and invasion, influences embryonic development, metabolism, and implantation potential, with confirmation by a 3D in-vitro implantation model. Method: Embryos were collected from female mice and cultured under different conditions: HTF only, HTF supplemented with OPN at different doses, and OPN-supplemented HTF with anti-[Formula: see text]v[Formula: see text]3 Ab or RGS2 inhibitors. Embryo morphology, development, and implantation rates were evaluated. Microarrays were used to evaluate gene expression profiling. Choriocarcinoma JAR and epithelial RL95-2 cell lines were adopted for the in-vitro implantation model. Oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were used to measure cellular metabolism levels. Results: The supplementation of OPN best enhanced the rate of embryo hatching-out and implantation onto the matrigel at 100 nM, but a higher degeneration rate was seen at a higher concentration ([Formula: see text]200 nM). In OPN-treated embryos, microarray analysis showed 51 significantly down-regulated genes and 60 up-regulated genes, including RGS2. Inhibition of [Formula: see text]v[Formula: see text]3 integrin decreased embryo implantation rates and down-regulated RGS2. Validation of the 3D implantation model using JAR spheroids and RL95-2 monolayer showed a similar effect. OPN also increased OCR and ECAR of the JAR cells, and migration assays revealed increased migration ability in OPN-overexpressed JAR cells, which was then inhibited by [Formula: see text]v[Formula: see text]3 and RGS2 inhibitors. Conclusion: Osteopontin, at a reasonable concentration, could enhance embryo development and implantation by interacting with [Formula: see text]v[Formula: see text]3 integrin, subsequently facilitating the RGS2 pathway. OPN has the potential to be a supplement during in vitro embryo culture in ART intervention.

RF05 | PSUN357 Scaffold Protein IQGAP1 Bridges Proliferative Signaling Between Bile Acids and the Cellular Microenvironment to Mediate Hepatic Cell Proliferation

Abstract The cell-cell and cell-extracellular matrix (ECM) communication is central to maintain cellular functions, including proliferation. Scaffold proteins can mediate these communications by binding specific protein effectors of signaling pathways and tethering them into a stable complex. IQ-domain containing GTPase Activating Protein 1 (IQGAP1) is a multi-domain scaffold protein that is involved in cytokinesis, proliferation and apoptosis. Each of the five domains of IQGAP1 bind distinct effector proteins to transduce cellular signals. In the liver, supraphysiological levels of bile acids, a cholesterol metabolite, is linked to the increase in IQGAP1 levels and is positively associated with transcriptional co-activator YAP. However, how bile acids induce expression of IQGAP1 remains unknown. We hypothesize that supraphysiological levels of bile acids act on the cell microenvironment to activate IQGAP1. As bile acids are induced during liver regeneration, we examined and found that Iqgap1 levels are increased. Remarkably this increase in Iqgap1 transcript coincides with that of bile acids after 2/3 partial hepatectomy in mice. In addition, IQGAP1 demonstrated increased localization into puncta at the cell surface in HepG2 cells, seeded on ECM microarrays with higher stiffness indicating mechano-sensing signals can alter IQGAP1 protein levels. We will examine IQGAP1 localization during liver regeneration and investigate if there is a cross talk between ECM stiffness, bile acid and IQGAP1 expression. Using domain deletion constructs, we will also identify the IQGAP1 domain responsible for coordinating the bile acid and cell-ECM communication in the liver. Collectively, these results will help elucidate the IQGAP1-specific cell signaling pathways used in facilitating liver cell proliferation, and test if IQGAP1 scaffold can integrate metabolite cues (bile acids) to communication between liver cells and the cellular microenvironment. Presentation: Saturday, June 11, 2022 1:06 p.m. - 1:11 p.m., Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

Single-cell RNA-sequencing analysis of aortic valve interstitial cells demonstrates the regulation of integrin signaling by nitric oxide.

Calcific aortic valve disease (CAVD) is an increasingly prevalent condition among the elderly population that is associated with significant morbidity and mortality. Insufficient understanding of the underlying disease mechanisms has hindered the development of pharmacologic therapies for CAVD. Recently, we described nitric oxide (NO) mediated S-nitrosylation as a novel mechanism for preventing the calcific process. We demonstrated that NO donor or an S-nitrosylating agent, S-nitrosoglutathione (GSNO), inhibits spontaneous calcification in porcine aortic valve interstitial cells (pAVICs) and this was supported by single-cell RNA sequencing (scRNAseq) that demonstrated NO donor and GSNO inhibited myofibroblast activation of pAVICs. Here, we investigated novel signaling pathways that are critical for the calcification of pAVICs that are altered by NO and GSNO by performing an in-depth analysis of the scRNA-seq dataset. Transcriptomic analysis revealed 1,247 differentially expressed genes in pAVICs after NO donor or GSNO treatment compared to untreated cells. Pathway-based analysis of the differentially expressed genes revealed an overrepresentation of the integrin signaling pathway, along with the Rho GTPase, Wnt, TGF-β, and p53 signaling pathways. We demonstrate that ITGA8 and VCL, two of the identified genes from the integrin signaling pathway, which are known to regulate cell-extracellular matrix (ECM) communication and focal adhesion, were upregulated in both in vitro and in vivo calcific conditions. Reduced expression of these genes after treatment with NO donor suggests that NO inhibits calcification by targeting myofibroblast adhesion and ECM remodeling. In addition, withdrawal of NO donor after 3 days of exposure revealed that NO-mediated transcriptional and translational regulation is a transient event and requires continuous NO exposure to inhibit calcification. Overall, our data suggest that NO and S-nitrosylation regulate the integrin signaling pathway to maintain healthy cell-ECM interaction and prevent CAVD.

Abstract 265: Inhibiting proteasome in extracellular matrix (ECM) detached cells promotes anoikis

Abstract The underlying mediators for altered cell-extracellular matrix (ECM) communication in tumors remain to be understood and are likely to be multifactorial. Detached cells from the ECM are often cleared by a death process called ‘anoikis’. Anoikis is critical for health and homeostasis maintenance. However, aggressive cancer cells acquire anoikis resistance to promote invasion and metastasis in new sites. We previously identified metabolic reprogramming and upregulation of Vacuolar-ATPase members responsible for anoikis resistance while the manipulation of these pathways induced anoikis. However, the mechanisms underlying anoikis resistance and subsequent metastasis are yet to be fully elucidated, thus limiting anoikis-targeting drug exploration for metastasis control. Herein, our RNA-seq data revealed upregulated genes associated with proteasome activity (such as PSMA, PSMB, PSMD, and POMP) in ECM detached cancer cells compared to adherent cancer cells. Targeting this, using proteasome inhibitors (MG132 or bortezomib) in breast cancer, cervical, and melanoma cell lines sensitized cancer cells to anoikis through increased cellular, mitochondrial stress, and misfolded protein accumulation. Moreover, Bortezomib treatment in vivo in melanoma tumor model repressed metastasis. These data demonstrate proteasome activity is hijacked by ECM detached cancer cells to maintain proteostasis thus promoting anoikis resistance and metastasis. Regardless of the impressive outcome in treating hematological malignancies, proteasome inhibitors show little effect against solid tumors in several clinical trials that utilized selective and more potent inhibitors either as a single or in combinatory therapy. In this sense, our finding suggests proteasome inhibitors may be more attractive for metastasis control rather than local tumor treatment. Altogether, our findings demonstrate enhanced proteasome activity within the tumor cells is critical for anoikis resistance, and targeting this can markedly overcome resistance and control tumor metastasis. Citation Format: Funmilayo Adeshakin, Guizhong Zhang, Adeleye Adeshakin, Xiaochun Wan. Inhibiting proteasome in extracellular matrix (ECM) detached cells promotes anoikis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 265.

αV Integrin Expression and Localization in Male Germ Cells.

Integrins are transmembrane receptors that facilitate cell adhesion and cell–extracellular matrix communication. They are involved in the sperm maturation including capacitation and gamete interaction, resulting in successful fertilization. αV integrin belongs to the integrin glycoprotein superfamily, and it is indispensable for physiological spermiogenesis and testosterone production. We targeted the gene and protein expression of the αV integrin subunit and described its membrane localization in sperm. Firstly, in mouse, we traced αV integrin gene expression during spermatogenesis in testicular fraction separated by elutriation, and we detected gene activity in spermatogonia, spermatocytes, and round spermatids. Secondly, we specified αV integrin membrane localization in acrosome-intact and acrosome-reacted sperm and compared its pattern between mouse, pig, and human. Using immunodetection and structured illumination microscopy (SIM), the αV integrin localization was confined to the plasma membrane covering the acrosomal cap area and also to the inner acrosomal membrane of acrosome-intact sperm of all selected species. During the acrosome reaction, which was induced on capacitated sperm, the αV integrin relocated and was detected over the whole sperm head. Knowledge of the integrin pattern in mature sperm prepares the ground for further investigation into the pathologies and related fertility issues in human medicine and veterinary science.

Epithelial–stromal cell interactions and extracellular matrix mechanics drive the formation of airway-mimetic tubular morphology in lung organoids

SummaryComplex human airway cellular organization where extracellular matrix (ECM) and epithelial and stromal lineages interact present challenges for organ study in vitro. Current in vitro lung models that focus on the lung epithelium do not represent complex airway morphology and cell-ECM interactions seen in vivo. Models including stromal populations often separate them via a semipermeable barrier precluding cell–cell interaction or the effect of ECM mechanics. We investigated the effect of stromal cells on basal epithelial cell-derived bronchosphere structure and function through a triple culture of human bronchial epithelial, lung fibroblast, and airway smooth muscle cells. Epithelial–stromal cross-talk resulted in epithelial cell-driven branching tubules with stromal cells surrounding epithelial cells termed bronchotubules. Agarose– Matrigel scaffold (Agrigel) formed a mechanically tuneable ECM, with adjustable viscoelasticity and stiffness enabling long-term tubule survival. Bronchotubule models may enable research into how epithelial–stromal cell and cell–ECM communication drive tissue patterning, repair, and development of disease.

SPP1 expression in the mouse uterus and placenta: implications for implantation†.

Secreted phosphoprotein 1 (SPP1, also known as osteopontin) binds integrins to mediate cell-cell and cell-extracellular matrix communication to promote cell adhesion, migration, and differentiation. Considerable evidence links SPP1 to pregnancy in several species. Current evidence suggests that SPP1 is involved in implantation and placentation in mice, but in vivo localization of SPP1 and in vivo mechanistic studies to substantiate these roles are incomplete and contradictory. We localized Spp1 mRNA and protein in the endometrium and placenta of mice throughout gestation, and utilized delayed implantation of mouse blastocysts to link SPP1 expression to the implantation chamber. Spp1 mRNA and protein localized to the endometrial luminal (LE), but not glandular epithelia (GE) in interimplantation regions of the uterus throughout gestation. Spp1 mRNA and protein also localized to uterine naturel killer (uNK) cells of the decidua. Within the implantation chamber, Spp1 mRNA localized only to intermittent LE cells, and to the inner cell mass. SPP1 protein localized to intermittent trophoblast cells, and to the parietal endoderm. These results suggest that SPP1: (1) is secreted by the LE at interimplantation sites for closure of the uterine lumen to form the implantation chamber; (2) is secreted by LE adjacent to the attaching trophoblast cells for attachment and invasion of the blastocyst; and (3) is not a component of histotroph secreted from the GE, but is secreted from uNK cells in the decidua to increase angiogenesis within the decidua to augment hemotrophic support of embryonic/fetal development of the conceptus.

Tissue Regeneration through Cyber‐Physical Systems and Microbots

AbstractTissue engineering is a systematic approach of assembling cells onto a 3D scaffold to form a functional tissue in the presence of critical growth factors. The scaffolding system guides stem cells through topological, physiochemical, and mechanical cues to differentiate and integrate to form a functional tissue. However, cellular communication during tissue formation taking place in a reactor needs to be understood properly to enable appropriate positioning of the cells in a 3D environment. Hence, sensors and actuators integrated with cyber‐physical system (CPS) may be able to sense the tissue microenvironment and direct cells/cellular aggregates to an appropriate position, respectively. This can facilitate better cell‐to‐cell communication and cell–extracellular matrix communication for proper tissue morphogenesis. Advancements are made in the field of smart scaffolds that can morph cells/cellular aggregates after sensing the cellular microenvironment in a desired 3D architecture by providing appropriate cues. Recent scientific developments in the additive manufacturing technology have enabled the fabrication of smart scaffolds to create structural and functional tissue constructs. Sensors/actuators, cyber‐systems, smart materials, and additive manufacturing put together is expected to lead to improved tissue‐engineered medical products. The present review aims to highlight the possibilities of advancement of BioCPS for tissue engineering and regenerative medicine.

ADAM and ADAMTS Proteins, New Players in the Regulation of Hepatocellular Carcinoma Microenvironment

Simple SummaryMembers of the adamalysin family are multi-domain proteins involved in many cancer-related functions. In this review, we will examine the literature on the involvement of adamalysins in hepatocellular carcinoma progression and their importance in the tumor microenvironment where they regulate the inflammatory response and the epithelial–mesenchymal transition. We complete this review with an analysis of adamalysin expression in a large cohort of patients with hepatocellular carcinoma from The Cancer Genome Atlas (TCGA) database. These original results give a new insight into the involvement of all adamalysins in the primary liver cancer.The tumor microenvironment plays a major role in tumor growth, invasion and resistance to chemotherapy, however understanding how all actors from microenvironment interact together remains a complex issue. The tumor microenvironment is classically represented as three closely connected components including the stromal cells such as immune cells, fibroblasts, adipocytes and endothelial cells, the extracellular matrix (ECM) and the cytokine/growth factors. Within this space, proteins of the adamalysin family (ADAM for a disintegrin and metalloproteinase; ADAMTS for ADAM with thrombospondin motifs; ADAMTSL for ADAMTS-like) play critical roles by modulating cell–cell and cell–ECM communication. During last decade, the implication of adamalysins in the development of hepatocellular carcinoma (HCC) has been supported by numerous studies however the functional characterization of most of them remain unsettled. In the present review we propose both an overview of the literature and a meta-analysis of adamalysins expression in HCC using data generated by The Cancer Genome Atlas (TCGA) Research Network.

Transcriptomic profile of Pea3 family members reveal regulatory codes for axon outgrowth and neuronal connection specificity

PEA3 transcription factor subfamily is present in a variety of tissues with branching morphogenesis, and play a particularly significant role in neural circuit formation and specificity. Many target genes in axon guidance and cell–cell adhesion pathways have been identified for Pea3 transcription factor (but not for Erm or Er81); however it was not so far clear whether all Pea3 subfamily members regulate same target genes, or whether there are unique targets for each subfamily member that help explain the exclusivity and specificity of these proteins in neuronal circuit formation. In this study, using transcriptomics and qPCR analyses in SH-SY5Y neuroblastoma cells, hypothalamic and hippocampal cell line, we have identified cell type-specific and subfamily member-specific targets for PEA3 transcription factor subfamily. While Pea3 upregulates transcription of Sema3D and represses Sema5B, for example, Erm and Er81 upregulate Sema5A and Er81 regulates Unc5C and Sema4G while repressing EFNB3 in SH-SY5Y neuroblastoma cells. We furthermore present a molecular model of how unique sites within the ETS domain of each family member can help recognize specific target motifs. Such cell-context and member-specific combinatorial expression profiles help identify cell–cell and cell-extracellular matrix communication networks and how they establish specific connections.

Biofabrication strategies for 3D in vitro models and regenerative medicine.

Organs are complex systems composed of different cells, proteins and signalling molecules that are arranged in a highly ordered structure to orchestrate a myriad of functions in our body. Biofabrication strategies can be applied to engineer 3D tissue models in vitro by mimicking the structure and function of native tissue through the precise deposition and assembly of materials and cells. This approach allows the spatiotemporal control over cell-cell and cell-extracellular matrix communication and thus the recreation of tissue-like structures. In this Review, we examine biofabrication strategies for the construction of functional tissue replacements and organ models, focusing on the development of biomaterials, such as supramolecular and photosensitive materials, that can be processed using biofabrication techniques. We highlight bioprinted and bioassembled tissue models and survey biofabrication techniques for their potential to recreate complex tissue properties, such as shape, vasculature and specific functionalities. Finally, we discuss challenges, such as scalability and the foreign body response, and opportunities in the field and provide an outlook to the future of biofabrication in regenerative medicine.

Combined treatment with Metformin and 2-deoxy glucose induces detachment of viable MDA-MB-231 breast cancer cells in vitro

Triple naegative breast cancer has an increased rate of distant metastasis and consequently poor prognosis. To metastasize, breast cancer cells must detach from the main tumour mass and resist anoikis, a programmed cell death induced by lack of cell-extracellular matrix communication. Although cancer cells must detach to metastasize in vivo, the viability of floating cancer cells in vitro is rarely investigated. Here we show that co-treatment of anoikis-resistant MDA-MB-231 cells with metformin and 2-deoxy-D-glucose (2-DG) increased the percentage of floating cells, of which about 95% were viable. Floating cells resumed their proliferation once they were reseeded in the pharmacological compound-free medium. Similar effects on detachment were observed on anoikis-prone MCF-7 cells. Co-treatment of MDA-MB-231 cells with metformin and 2-DG induced a strong activation of AMP-activated protein kinase (AMPK), which was reduced by AMPK inhibitor compound C that prevented detachment of MDA-MB-231 cells. However, direct AMPK activators A-769662 and AICAR did not have any major effect on the percentage of floating MDA-MB-231 cells, indicating that AMPK activation is necessary but not sufficient for triggering detachment of cancer cells. Our results demonstrate that separate analysis of floating and attached cancer cells might be important for evaluation of anti-cancer agents.

Induced Physical Distension of Rat Mammary Glands Accelerates the Onset of Apoptosis and Involution

Objective: Weaning is a process that results in a reduction in milk secretion, an increase in mammary epithelial cell (MEC) apoptosis, and involution of the mammary gland. The local mechanisms initiating MEC apoptosis and involution are unclear, although the physical morphology of the MEC may influence cell-cell and cell-extracellular matrix communication and thus may alter function. This study examined the effect of physical distension of alveoli on the early molecular events that occur at the onset of involution of rat mammary glands. Methods: Mammary tissue was collected post-mortem from lactating Sprague-Dawley rats at 0, 1, 3, and 6 h (n=6 per time point) following acute physical distension of an inguinal gland with isosmotic sucrose solution (0.8 ml; equivalent to ~6 h worth of milk accumulation) via the teat canal followed by sealing with adhesive (infused). The remaining teats on each rat were either left unsealed for pups to suckle (control), or were sealed to induce milk accumulation and mammary engorgement (engorged). Results: There was a low number of positive in situ-end labeled (ISEL) nuclei in suckled control glands indicating they had a low number of cells with fragmented DNA or were apoptotic. However, there was a greater number of ISEL nuclei in sucrose-infused and milk-engorged teat-sealed glands by 1 and 6 h, respectively, such that these changes were accelerated by sucrose infusion, compared with milk accumulation alone. The timing of the decline in abundance of β1-integrin (cell-extracellular matrix protein) and occludin (tight junction protein), and increase in abundance of the activated apoptotic marker signal transducer and activator of transcription factor-3 (pSTAT3) protein were also accelerated by sucrose infusion. Conclusion: Physical distension by sucrose infusion accelerated the onset of the first stage of mammary apoptosis and involution, supporting a role for mechanotransduction during these processes.

Transcriptional modulation of intestinal innate defense/inflammation genes by preterm infant microbiota in a humanized gnotobiotic mouse model.

Background and AimsIt is known that postnatal functional maturation of the small intestine is facilitated by microbial colonization of the gut. Preterm infants exhibit defects in gut maturation, weak innate immunity against intestinal infection and increased susceptibility to inflammatory disorders, all of which may be related to the inappropriate microbial colonization of their immature intestines. The earliest microbes to colonize the preterm infant gut encounter a naïve, immature intestine. Thus this earliest microbiota potentially has the greatest opportunity to fundamentally influence intestinal development and immune function. The aim of this study was to characterize the effect of early microbial colonization on global gene expression in the distal small intestine during postnatal gut development.MethodsGnotobiotic mouse models with experimental colonization by early (prior to two weeks of life) intestinal microbiota from preterm human infants were utilized. Microarray analysis was used to assess global gene expression in the intestinal epithelium.Results and ConclusionMultiple intestinal genes involved in metabolism, cell cycle regulation, cell-cell or cell-extracellular matrix communication, and immune function are developmental- and intestinal microbiota- regulated. Using a humanized gnotobiotic mouse model, we demonstrate that certain early preterm infant microbiota from prior to 2 weeks of life specifically induce increased NF-κB activation and a phenotype of increased inflammation whereas other preterm microbiota specifically induce decreased NF-κB activation. These fundamental differences correlate with altered clinical outcomes and suggest the existence of optimal early microbial communities to improve health outcomes.

Novel ITGB6 mutation in autosomal recessive amelogenesis imperfecta.

ObjectiveHereditary defects in tooth enamel formation, amelogenesis imperfecta (AI), can be non-syndromic or syndromic phenotype. Integrins are signaling proteins that mediate cell–cell and cell–extracellular matrix communication, and their involvement in tooth development is well known. The purposes of this study were to identify genetic cause of an AI family and molecular pathogenesis underlying defective enamel formation.Materials and MethodsWe recruited a Turkish family with isolated AI and performed mutational analyses to clarify the underlying molecular genetic etiology.ResultsAutozygosity mapping and exome sequencing identified a novel homozygous ITGB6 transversion mutation in exon 4 (c.517G>C, p.Gly173Arg). The glycine at this position in the middle of the βI-domain is conserved among a wide range of vertebrate orthologs and human paralogs. Clinically, the enamel was generally thin and pitted with pigmentation. Thicker enamel was noted at the cervical area of the molars.ConclusionsIn this study, we identified a novel homozygous ITGB6 mutation causing isolated AI, and this advances the understanding of normal and pathologic enamel development.

Role of Wnt signalling in early pregnancy.

The integration of a complex network of signalling molecules promotes implantation of the blastocyst and development of the placenta. These processes are crucial for a successful pregnancy and fetal growth and development. The signalling network involves both cell-cell and cell-extracellular matrix communication. The family of secreted glycoprotein ligands, the Wnts, plays a major role in regulating a wide range of biological processes, including embryonic development, cell fate, proliferation, migration, stem cell maintenance, tumour suppression, oncogenesis and tissue homeostasis. Recent studies have provided evidence that Wnt signalling pathways play an important role in reproductive tissues and in early pregnancy events. The focus of this review is to summarise our present knowledge of expression, regulation and function of the Wnt signalling pathways in early pregnancy events of human and other model systems, and its association with pathological conditions. Despite our recent progress, much remains to be learned about Wnt signalling in human reproduction. The advancement of knowledge in this area has applications in the reduction of infertility and the incidence and morbidity of gestational diseases.

MicroRNA identity and abundance in developing swine adipose tissue as determined by solexa sequencing

MicroRNAs (miRNAs) are small ∼22-nt regulatory RNAs that regulate the stability and translation of cognate mRNAs. MiRNAs participate in the regulation of adipogenesis, and identification of the full repertoire of miRNAs expressed in adipose tissue is likely to significantly increase our understanding of adipose tissue growth and development. Here, we adopted a deep sequencing approach to determine the identity and abundance of miRNAs in developing swine adipose tissue. Via this approach, we identified the sequences and relative expression levels of 227 conserved miRNAs (of which 59 were novel) and 66 potential porcine miRNAs. The expression levels displayed a large range, as reflected by the number of sequence reads, which varied from several counts for rare miRNAs to several million reads for the most abundant miRNAs. The abundant miRNAs principally belonged to 32 miRNA gene families, including miR-143, miR-103, let-7, and miR-148. Of the conserved miRNAs, 93 miRNAs were up-regulated and 33 miRNAs were down-regulated in the adult pig adipose tissue. Moreover, we observed sequence variants and seed edits of the miRNAs. KEGG pathway analysis and GO term enrichment suggested that highly expressed miRNAs are involved in adipose tissue development, signal transduction, cell-cell and cell-extracellular matrix communication, neural development and function, and lipid metabolism including carboxylic acid, oxoacid, fatty acid, steroid, glycerolipid, alcohol and phospholipid metabolism. Our results expand the number of known porcine miRNAs and provide a thorough account of the miRNA transcriptome in porcine adipose tissue.