Extracellular Matrix-related Gene Expression Research Articles

To elucidate the bioactive components of Lamiophlomis herba in the treatment of liver fibrosis via plasma pharmacochemistry and network pharmacology

Lamiophlomis Herba (LH) is a traditional Chinese and Tibetan dual-use herb with hemostatic and analgesic effects, and is widely used in the clinical treatment of traumatic bleeding and pain. In recent years, LH has been proven to treat liver fibrosis (LF), but the chemical components related to the pharmacological properties of LH in the treatment of LF are still unclear. Based on the theory of plasma pharmachemistry, the characteristic components in water extract and drug-containing plasma samples of LH were qualitatively analyzed by UPLC-Q-TOF-MS. The chemical components in plasma were screened and the targets were predicted by network pharmacology. Then, the predicted components and targets were verified in vitro by Elisa and qRT-PCR technology. Finally, the pharmacological effects of LH and its monomeric components were determined by hematoxylin-eosin staining of rat liver. A total of 50 chemical constituents were identified in LH, of which 12 were blood prototypes and 9 were metabolites. In vitro experiments showed that LH and its monomeric components luteolin, shanzhiside methyl ester, loganic acid, loganin, 8-O-acetyl shanzhiside methyl ester could increase the expression of antioxidant genes (NQO-1, HO-1) and decrease the expression of inflammatory genes (IL-6, IL-18), thereby reducing the expression of extracellular matrix-related genes and proteins (COL1A1, COL3A1, LN, α-sma, PC-III, Col-IV). In vivo experiments showed that LH could reduce the area of LF in rats in a dose-dependent manner, and shanzhiside methyl ester and 8-O-acetyl shanzhiside methyl ester may be the main components in pharmacodynamics. These effects may be mediated by LH-mediated Nrf2/NF-κB pathway. This study explored the potential pharmacodynamic components of LH in the treatment of LF, and confirmed that shanzhiside methyl ester and 8-O-acetyl shanzhiside methyl ester play a key role in the treatment of LF with LH.

Abstract 15180: Inhibition of Tgfβ Signaling Enables Durable Ventricular Pacing by TBX18 Gene Transfer

Introduction: Somatic gene transfer of an embryonic transcription factor, TBX18, could directly reprogram ventricular myocytes to induced pacemaker myocytes. We and others have previously reported that direct myocardial gene delivery of TBX18 created ventricular pacing in heart block animals in vivo. However, the de novo biological pacing was short-lived, diminishing its translational value. Hypothesis: Our preliminary RNAseq and mass spec data indicate pronounced Tgfβ signaling in TBX18-transduced cardiomyocytes. Tgfβ signaling induces fibrosis in multiple organs including the heart. We hypothesized that TBX18 triggers Tgfβ signaling and results in fibrosis, leading to loss of durable ventricular pacing. Methods: Adenoviral vectors expressing TBX18 or GFP was delivered to neonatal rat ventricular myocytes (NRVMs) in vitro or to the left ventricular apex of adult rats with complete atrioventricular block (CAVB). Results: In vitro experiments showed increased population of vimentin+ fibroblasts and alpha smooth muscle actin+ myofibroblasts in TBX18-NRVMs compared to GFP-NRVMs at 2 weeks after gene transfer. Single cell transcriptome data indicated that TBX18 triggered substantial changes in extracellular matrix-related gene expression and Tgfβ signaling. Quantitative RT-PCR and ELISA validated scRNA-seq data, indicating fibroblast activation. TBX18-NRVMs showed faster automaticity than that control did, but their automaticity began wavering after day 7. Treatment of TBX18-NRVMs with a Tgfβ signaling inhibitor, A83-01, mitigated fibroblast activation by TBX18 and preserved their automaticity. Direct delivery of TBX18 into ventricular myocardium resulted in focal fibrosis as well as expansion of vimentin+ fibroblasts at day 7. TBX18 gene transfer initiated ventricular pacing in a rat model of CAVB, but ventricular pacing diminished after one week. In contrast, treating TBX18-injected animals with A83-01 enabled sustained biological pacing beyond three weeks of after gene delivery, accompanied by reduced fibrosis in situ. Conclusions: Inhibition of Tgfβ signaling is sufficient to achieve durable cardiac pacing by TBX18-induced biological pacemakers.

Deciphering mechanisms of cardiomyocytes and non-cardiomyocyte transformation in myocardial remodeling of permanent atrial fibrillation

IntroductionAtrial fibrillation (AF) is the most prevalent cardiac arrhythmia, and it significantly increases the risk of cardiovascular complications and morbidity, even with appropriate treatment. Tissue remodeling has been a significant topic, while its systematic transcriptional signature remains unclear in AF. ObjectivesOur study aims to systematically investigate the molecular characteristics of AF at the cellular-level. MethodsWe conducted single-nuclei RNA-sequencig (snRNA-seq) analysis using nuclei isolated from the left atrial appendage (LAA) of AF patients and sinus rhythm. Pathological staining was performed to validate the key findings of snRNA-seq. ResultsA total of 30 cell subtypes were identified among 80, 592 nuclei. Within the LAA of AF, we observed a specific subtype of dedifferentiated cardiomyocytes (CMs) characterized by reduced expression of cardiac contractile proteins (TTN and TRDN) and heightened expression of extracellular-matrix related genes (COL1A2 and FBN1). Transcription factor prediction analysis revealed that gene expression patterns in dedifferentiated CMs were primarily regulated by CEBPG and GISLI. Additionally, we identified a distinct subtype of endothelial progenitor cells (EPCs) demonstrating elevated expression of PROM1 and KDR, a population decreased within the LAA of AF. Epicardial adipocytes disclosed a reduced release of the anti-inflammatory and anti-fibrotic factor PRG4, and an augmented secretion of VEGF signals targeting CMs. Additionally, we noted accumulation of M2-like macrophages and CD8+ T cells with high pro-inflammatory score in LAA of AF. Furthermore, the analysis of intercellular communication revealed specific pathways related to AF, such as inflammation, extracellular matrix, and vascular remodeling signals. ConclusionsThis study has discovered the presence of dedifferentiated CMs, a decrease in endothelial progenitor cells, a shift in the secretion profile of adipocytes, and an amplified inflammatory response in AF. These findings could offer crucial insights for future research on AF and serve as valuable references for investigating novel therapeutic approaches for AF.

Stromal cells-specific retinoic acid determines parturition timing at single-cell and spatial-temporal resolution

The underlying mechanisms governing parturition remain largely elusive due to limited knowledge of parturition preparation and initiation. Accumulated evidences indicate that maternal decidua plays a criticalrole in parturition initiation. To comprehensively decrypt the cell heterogeneity in decidua approaching parturition, we investigate the roles of various cell types in mouse decidua process and reveal previously unappreciated insights in parturition initiation utilizing single-cell RNA sequencing (scRNA-seq). We enumerate the cell types in decidua and identity five different stromal cells populations and one decidualized stromal cells. Furthermore, our study unravels that stromal cells prepare for parturition by regulating local retinol acid (RA) synthesis. RA supplement decreases expression of extracellular matrix-related genes invitro and accelerates the timing of parturition invivo. Collectively, the discovery of contribution of stromal cells in parturition expands current knowledge about parturition and opens up avenues for the intervention of preterm birth (PTB).

FGF9 Recruits β-Catenin to Increase Hepatic ECM Synthesis and Promote NASH-Driven HCC.

Most nonalcoholic steatohepatitis (NASH) patients develop severe fibrosis through extracellular matrix (ECM) accumulation, which can lead to hepatocellular carcinoma (HCC). Fibroblast growth factor 9 (FGF9)is involved in serial types of cancer; however, the specific role of FGF9 in NASH-driven HCC is not fully understood. This study finds that FGF9 is increased in patients with NASH-associated HCC. Furthermore, NASH-driven HCC mice models by feeding wildtype mice with high-fat/high-cholesterol (HFHC) diet and low dose carbon tetrachloride (CCl4 ) treatment is established; and identified that hepatic FGF9 is increased; with severe fibrosis. Additionally, AAV-mediated knockdown of FGF9 reduced the hepatic tumor burden of NASH-driven HCC mice models. Hepatocyte-specific FGF9 transgenic mice (FGF9Alb ) fed with a HFHC diet without CCl4 treatment exhibited an increased hepatic ECM and tumor burden. However, XAV-939 treatment blocked ECM accumulation and NASH-driven HCC in FGF9Alb mice fed with HFHC diet. Molecular mechanism studies show that FGF9 stimulated the expression of ECM related genes in a β-catenin dependent manner; and FGF9 exerts its effect on β-catenin stability via the ERK1/2-GSK-3β signaling pathway. In summary, the data provides evidence for the critical role of FGF9 in NASH-driven HCC pathogenesis; wherein it promotes the tumors formation through the ECM pathway.

Resistance training rejuvenates aging skin by reducing circulating inflammatory factors and enhancing dermal extracellular matrices

Aerobic training (AT) is suggested to be an effective anti-aging strategy for skin aging. However, the respective effects of resistance training (RT) have not been studied. Therefore, we compared the effects of AT and RT on skin aging in a 16-week intervention in 61 healthy sedentary middle-aged Japanese women. Data from 56 women were available for analysis. Both interventions significantly improved skin elasticity and upper dermal structure, and RT also improved dermal thickness. After the training intervention, expression of dermal extracellular matrix-related genes was increased in normal human primary dermal fibroblasts. AT and RT had different effects on circulating levels of factors, such as cytokines, hormones in serum, and metabolites, and RT increased dermal biglycan (BGN). To our knowledge, this is the first report to show different effects of AT and RT on skin aging and identify the key factors involved in RT-induced skin rejuvenation.

Expression of Adipose Tissue Extracellular Matrix-Related Genes Predicts Weight Loss after Bariatric Surgery.

We evaluated the association between white adipose tissue parameters before bariatric surgery (BS) and post-surgical weight loss, with an especial focus on extracellular matrix (ECM) gene expression. Paired samples from subcutaneous (SAT) and visceral adipose tissue (VAT) were obtained from 144 subjects undergoing BS. The association between total body weight loss (%TBWL) at 12 months after BS and the histological characteristics and gene expression of selected genes in SAT and VAT was analyzed. Fat cell area, size-frequency distribution, and fibrosis in SAT or VAT prior to surgery were not associated with %TBWL. On the contrary, the SAT expression of COL5A1 and COL6A3 was associated with %TBWL after BS (both p < 0.001), even after adjusting for age, gender, baseline BMI, and type 2 diabetes status (T2D). Furthermore, in logistic regression analyses, the expression of these genes was significantly associated with insufficient WL (IWL = TBWL < 20%) after BS (respectively, p = 0.030 and p = 0.031). Indeed, in ROC analysis, the prediction of IWL based on sex, age, BMI, T2D, and the type of surgery (AUC = 0.71) was significantly improved with the addition of SAT-COL5A1 gene expression (AUC = 0.88, Z = 2.13, p = 0.032). Our data suggest that the expression of SAT ECM-related genes may help explain the variability in TBWL following BS.

CD73-Positive Cell Spheroid Transplantation Attenuates Colonic Atrophy

The incidence of inflammatory bowel diseases (IBD) is increasing worldwide. Mesenchymal stem/stromal cells (MSCs) have immunomodulatory functions and are a promising source for cell transplantation therapy for IBD. However, owing to their heterogeneous nature, their therapeutic efficacy in colitis is controversial and depends on the delivery route and form of transplanted cells. Cluster of differentiation (CD) 73 is widely expressed in MSCs and used to obtain a homogeneous MSC population. Herein, we determined the optimal method for MSC transplantation using CD73+ cells in a colitis model. mRNA sequencing analysis showed that CD73+ cells exhibited a downregulation of inflammatory gene expression and an upregulation of extracellular matrix-related gene expression. Furthermore, three-dimensional CD73+ cell spheroids showed enhanced engraftment at the injured site through the enteral route, facilitated extracellular matrix remodeling, and downregulated inflammatory gene expression in fibroblasts, leading to the attenuation of colonic atrophy. Therefore, the interaction between intestinal fibroblasts and exogenous MSCs via tissue remodeling is one mechanism that can be exploited for colitis prevention. Our results highlight that the transplantation of homogeneous cell populations with well-characterized properties is beneficial for IBD treatment.

Methylphenidate Promotes Premature Growth Plate Closure: In Vitro Evidence

It is well known that patients with attention deficit hyperactivity disorder treated with stimulants, such as methylphenidate hydrochloride (MPH), have reduced height and weight. Even though MPH has an anorexigenic effect, an additional impact of this drug on the growth plate cannot be discarded. In this study, we aimed to determine the cellular effect of MPH on an in vitro growth plate model. We tested the effects of MPH on the viability and proliferation of a prechondrogenic cell line via an MTT assay. In vitro differentiation of this cell line was performed, and cell differentiation was evaluated through the expression of cartilage- and bone-related genes as measured via RT-PCR. MPH did not alter the viability or proliferation of prechondrogenic cells. However, it reduced the expression of cartilage extracellular matrix-related genes (type II collagen and aggrecan) and increased the expression of genes involved in growth plate calcification (Runx2, type I collagen, and osteocalcin) at different phases of their differentiation process. Our results evidence that MPH upregulates genes associated with growth plate hypertrophic differentiation. This may induce premature closure of the growth plate, which would contribute to the growth retardation that has been described to be induced by this drug.

Unique ligand and kinase-independent roles of the insulin receptor in regulation of cell cycle, senescence and apoptosis

Insulin acts through the insulin receptor (IR) tyrosine kinase to exert its classical metabolic and mitogenic actions. Here, using receptors with either short or long deletion of the β-subunit or mutation of the kinase active site (K1030R), we have uncovered a second, previously unrecognized IR signaling pathway that is intracellular domain-dependent, but ligand and tyrosine kinase-independent (LYK-I). These LYK-I actions of the IR are linked to changes in phosphorylation of a network of proteins involved in the regulation of extracellular matrix organization, cell cycle, ATM signaling and cellular senescence; and result in upregulation of expression of multiple extracellular matrix-related genes and proteins, down-regulation of immune/interferon-related genes and proteins, and increased sensitivity to apoptosis. Thus, in addition to classical ligand and tyrosine kinase-dependent (LYK-D) signaling, the IR regulates a second, ligand and tyrosine kinase-independent (LYK-I) pathway, which regulates the cellular machinery involved in senescence, matrix interaction and response to extrinsic challenges.

Effects of zinc deficiency on the regeneration of olfactory epithelium in mice.

The olfactory epithelium can regenerate after damage; however, the regeneration process is affected by various factors, such as viral infections, head trauma, and medications. Zinc is an essential trace element that has important roles in organ development, growth, and maturation. Zinc also helps regulate neurotransmission in the brain; nevertheless, its relationship with olfactory epithelium regeneration remains unclear. Therefore, we used a severe zinc deficiency mouse model to investigate the effects of zinc deficiency on olfactory epithelium regeneration. Male wild-type C57BL/6 mice were divided into zinc-deficient and control diet groups at the age of 4 weeks, and methimazole was administered at the age of 8 weeks to induce severe olfactory epithelium damage. We evaluated the olfactory epithelium before and 7, 14, and 28 d after methimazole administration by histologically analyzing paraffin sections. RNA sequencing was also performed at the age of 8 weeks before methimazole administration to examine changes in gene expression caused by zinc deficiency. In the zinc-deficient group, the regenerated olfactory epithelium thickness was decreased at all time points, and the numbers of Ki-67-positive, GAP43-positive, and olfactory marker protein-positive cells (i.e., proliferating cells, immature olfactory neurons, and mature olfactory neurons, respectively) failed to increase at some time points. Additionally, RNA sequencing revealed several changes in gene expression, such as a decrease in the expression of extracellular matrix-related genes and an increase in that of inflammatory response-related genes, in the zinc-deficient group. Therefore, zinc deficiency delays olfactory epithelium regeneration after damage in mice.

Transcriptomic and physiological analyses reveal temporal changes contributing to the delayed healing response to arterial injury in diabetic rats

ObjectiveAtherosclerosis is a leading cause of mortality in the rapidly growing population with diabetes mellitus. Vascular interventions in patients with diabetes can lead to complications attributed to defective vascular remodeling and impaired healing response in the vessel wall. In this study, we aim to elucidate the molecular differences in the vascular healing response over time using a rat model of arterial injury applied to healthy and diabetic conditions. MethodsWistar (healthy) and Goto-Kakizaki (GK, diabetic) rats (n = 40 per strain) were subjected to left common carotid artery (CCA) balloon injury and euthanized at different timepoints: 0 and 20 hours, 5 days, and 2, 4, and 6 weeks. Noninvasive morphological and physiological assessment of the CCA was performed with ultrasound biomicroscopy (Vevo 2100) and corroborated with histology. Total RNA was isolated from the injured CCA at each timepoint, and microarray profiling was performed (n = 3 rats per timepoint; RaGene-1_0-st-v1 platform). Bioinformatic analyses were conducted using R software, DAVID bioinformatic tool, online STRING database, and Cytoscape software. ResultsSignificant increase in the neointimal thickness (P < .01; two-way analysis of variance) as well as exaggerated negative remodeling was observed after 2 weeks of injury in GK rats compared with heathy rats, which was confirmed by histological analyses. Bioinformatic analyses showed defective expression patterns for smooth muscle cells and immune cell markers, along with reduced expression of key extracellular matrix-related genes and increased expression of pro-thrombotic genes, indicating potential faults on cell regulation level. Transcription factor–protein-protein interaction analysis provided mechanistic evidence with an array of transcription factors dysregulated in diabetic rats. ConclusionsIn this study, we have demonstrated that diabetic rats exhibit impaired arterial remodeling characterized by a delayed healing response. We show that increased contractile smooth muscle cell marker expression coincided with decreased matrix metalloproteinase expression, indicating a potential mechanism for a lack of extracellular matrix reorganization in the impaired vascular healing in GK rats. These results further corroborate the higher prevalence of restenosis in patients with diabetes and provide vital molecular insights into the mechanisms contributing to the impaired arterial healing response in diabetes. Moreover, the presented study provides the research community with the valuable longitudinal gene expression data bank for further exploration of diabetic vasculopathy.

A Novel Role for Biomechanical Forces in the Pathogenesis of Idiopathic Constipation

INTRODUCTION: Chronic constipation is a common gastrointestinal disorder, but an anatomic or physiologic cause is identified in only 5% of patients. Surgery to remove chronically dilated segments of intestine is offered as a last resort. Our goal is to understand how the intestinal microenvironment is affected by biomechanical forces such as chronic distention in idiopathic constipation. METHODS: To evaluate the effects of biomechanical force on the colon, we used 3 models: (1) colon from wildtype mice was cultured ex vivo for 24 hours with and without distention and extracellular matrix–related gene expression was assayed using reverse transcriptase-polymerase chain reaction; (2) an in vivo mouse model of enteric distention was assessed at 7 to 14 days by immunohistochemistry; and (3) human specimens from pediatric patients who underwent colon resections for chronic idiopathic constipation (n = 27) were histologically evaluated to clinically validate our findings (Figure).FigureRESULTS: Ex vivo and in vivo colon under tension demonstrated increased fibrosis compared with control. Tension resulted in significantly decreased expression of antifibrotic Gremlin-1 and increased expression of profibrotic interleukin-1β. Collagens and matrix metalloproteinases were downregulated while laminins were upregulated. Seventy-seven percent of patients with chronic idiopathic constipation displayed dilated and/or redundant colon on imaging. In surgical specimens, 26% had interstitial fibrosis, 33% had muscular hypertrophy, and 15% had myocyte vacuolization. These histologic findings were also seen in distended mouse colon. CONCLUSION: Chronic intestinal distention led to a profibrotic phenotype in all 3 models. We conclude that mechanically sensitive cells may be affected by chronic distention and contribute to intestinal dysfunction in idiopathic constipation.

TGF-β-Mediated Modulation of Cell-Cell Interactions in Postconfluent Maturing Corneal Endothelial Cells.

PurposeTransforming growth factor-beta (TGF-β) is known to influence many cell functions. In the corneal endothelium, TGF-β1 exerts contextual effects, promoting endothelial–mesenchymal transition in proliferating cells and enhancing barrier integrity in early confluent maturing cells. Herein, we studied how TGF-β isoforms participate in the formation of corneal endothelial intercellular junctions.MethodsCorneal endothelial cells (CECs) were cultured using a two-phase media approach. When CECs reached confluence, the proliferation medium was replaced with maturation medium, which was supplemented or not with TGF-β isoforms. The cell morphology (circularity index), intercellular junction protein expression, trans-endothelial electrical resistance (TEER), and permeability of 7-day postconfluent CECs were assessed. Gene transcription and signaling pathways that were activated following maturation in the presence of TGF-β2 were also studied. The beneficial effect of TGF-β2 on CEC maturation was evaluated using ex vivo corneas mounted on a corneal bioreactor.ResultsThe results showed increases in circularity index, membrane localization of junction-related proteins, and TEER when TGF-β isoforms were individually added during the maturation phase, and TGF-β2 was the most effective isoform. Gene profiling revealed an increase in extracellular matrix-related gene expression. In ex vivo cell adhesion experiments, CECs that were matured in the presence of TGF-β2 had a higher circularity index and cell density and exhibited cell membrane-localized junction-related protein expression at earlier time points.ConclusionsThese results suggest that TGF-β2 can strengthen cell–cell and cell–substrate adhesion, which accelerates barrier integrity establishment and thus enhances CEC functionality.

Initial assessment of gingival biotype as a potential source of variability in the migration, contraction and gene expression of fibroblasts

ObjectiveGingival biotype refers to the clinical classification of gingiva based on the thickness of the tissue, with thick gingival tissues more resistant to trauma and recession than the thin variant. However, to date there has never been an analysis of whether fibroblasts isolated from different biotypes possess inherent phenotypic differences. We hypothesized that gingival fibroblasts from thick and thin biotype would exhibit differences in migration, contraction and gene expression in vitro in the presence of either transforming growth factor beta one (TGF-β1) or tumor necrosis factor alpha (TNFα), two major cytokines involved in wound repair. DesignMigration was quantified using closure of scratch wound assays, contraction was assessed using attached and detached collagen lattices and extracellular matrix related gene expression using Taqman Realtime polymerase chain reaction. ResultsHuman gingival fibroblasts isolated from both biotypes showed similar rates of closure of scratch wounds, which was not influenced by the addition of TGF-β1 or TNFα. Fibroblasts from both biotypes contracted detached, but not attached, collagen gels to 50 % of their original weight although this contraction was not associated with incorporation of α-smooth muscle actin into stressfibres under any tested culture condition. Analysis of gene expression showed that POSTN, and ACTA2 mRNA levels did not significantly change, but CCN2 and COL1A2 mRNA levels were significantly higher in thick compared to thin fibroblasts in response to TGF-β1. ConclusionWhile supra-cellular factors influence the healing, esthetic outcomes and recession in thin gingival biotypes, differences in gingival fibroblast gene expression in response to growth factors may also play a role and warrants further investigation.

Chitosan/Hyaluronic Acid/MicroRNA-21 Nanoparticle-Coated Smooth Titanium Surfaces Promote the Functionality of Human Gingival Fibroblasts.

PurposeForming a compact biological seal between the gingiva and the implant interface around the percutaneous parts of an implant is one of the key issues in preventing peri-implantitis.MethodsIn this study, since microRNA-21 (miR-21) has been approved to promote fibroblast proliferation and collagen formation in skin fibrosis, we prepared miR-21-loaded chitosan (CS)/tripolyphosphate (TPP)/hyaluronic acid (HA) nanoparticles (CTH NPs) and cross-linked them to smooth Ti surfaces with 0.2% gel solution for reverse transfection, after which isolated human gingival fibroblasts were cultured on the miR-21-functionalized Ti substrates.ResultsAn optimal CS:TPP:HA ratio (1:0.15:0.1) and N/P ratio (20:1) were chosen to produce appropriate nanoparticles. Finally, the CTH/miR-21 nanoparticle-coated smooth Ti surfaces demonstrated increased fibroblast adhesion, proliferation and expression of extracellular matrix-related genes along with similar cytotoxicity and cell spreading on the miR-21-functionalized Ti surface and the unmodified smooth Ti surface.ConclusionThe chitosan-based nanoparticles might be an efficient nonviral miRNA vector to form a stable biological seal in percutaneous areas of Ti for clinical use.

A bioinformatics approach to study the role of calcium phosphate properties in bone regeneration

Calcium phosphates (CaPs) are extensively used as biomaterials for bone repair and regeneration and are considered an interesting synthetic alternative for natural bone grafts. However, the bone regenerative potential of many CaP bone graft substitutes requires further improvements. The bioactivity of CaPs is regulated by a variety of properties, including their chemical composition and (surface) structural properties. In this study, we performed a comprehensive characterization of six CaP ceramics that had been previously tested in several in vitro and in vivo studies, allowing us to in silico distinguish osteoinductive and non-osteoinductive CaP ceramics. The numerical parameters obtained from this characterization were further correlated with osteogenesis-associated gene expression and pathways of osteoblastic MG63 cells cultured on these six CaP ceramics. The results showed, among others, the influence of both chemical and structural parameters on cytoskeleton- and extracellular matrix-related gene expression and osteogenesis-associated pathways such as MAPK and BMP. The bioinformatics approach presented here allows exploring the transcriptional landscape induced by CaP ceramics and gaining fundamental knowledge about cell-biomaterial interactions.

Differential expression of extracellular matrix-related genes in tissue of stable and unstable atherosclerotic plaques

Differential expression of extracellular matrix-related genes in tissue of stable and unstable atherosclerotic plaques

Acoustically accelerated neural differentiation of human embryonic stem cells

Human embryonic stem cells (hESCs) and their derived products offer great promise for targeted therapies and drug screening, however, the hESC differentiation process of mature neurons is a lengthy process. To accelerate the neuron production, an acoustic stimulator producing surface acoustic waves (SAWs) is proposed and realized by clamping a flexible printed circuit board (PCB) directly onto a piezoelectric substrate. Neural differentiation of the hESCs is greatly accelerated after application of the acoustic stimulations. Acceleration mechanisms for neural differentiation have been explored by bulk RNA sequencing, quantitative polymerase chain reaction (qPCR) and immunostaining. The RNA sequencing results show changes of extracellular matrix-related and physiological activity-related gene expression in the low or medium SAW dose group and the high SAW dose group, respectively. The neural progenitor cell markers, including Pax6, Sox1, Sox2, Sox10 and Nkx2-1, are less expressed in the SAW dose groups compared with the control group by the qPCR. Other genes including Alk, Cenpf, Pcdh17, and Actn3 are also found to be regulated by the acoustic stimulation. Moreover, the immunostaining confirmed that more mature neuron marker Tuj1-positive cells, while less stem cell marker Sox2-positive cells, are presented in the SAW dose groups. These results indicate that the SAW stimulation accelerated neural differentiation process. The acoustic stimulator fabricated by using the PCB is a promising tool in regulation of stem cell differentiation process applied in cell therapy. Statement of significanceHuman embryonic stem cells (hESC) are used for investigating the complex mechanisms involved in the development of specialized biological cells and organs. Different types of hESCs derived cell products can be used for cell therapy procedures aiming to regenerate functional tissues in patients who suffer from various degenerative diseases. Accelerating the hESCs’ differentiation process can considerably benefit the clinical utilization of these cells. This study develops a highly effective acoustic stimulator working at ∼20 MHz to investigate what roles do acousto-mechanical stimuli play in the differentiation of hESCs. Our results show that acoustic dose alters the extracellular matrix and physiological activity-related gene expression, which indicates that the acoustic stimulation is an important tool for regulating the stem cells’ differentiation processes in cell therapy.

Temporal profile of angiogenesis and expression of extracellular matrix-related genes in rat brains following experimental intracerebral hemorrhage.

Angiogenesis is essential for the repair process after intracerebral hemorrhage (ICH). Given the importance of the extracellular matrix (ECM) in angiogenesis, we analysed the temporal profile of angiogenesis in rat brains on days 4, 7, and 21 after ICH. To this end, we compared the expression of ECM-related genes between ICH-induced and sham-operated groups using a complementary DNA (cDNA) array. We further measured protein expression using western blot and immunohistochemistry assays. Fluorescein isothiocyanate (FITC)-dextran was injected into the tail vein to examine the angioarchitecture in the perihematomal region. Among the 88 ECM-related genes, we identified 42, 50, and 38 genes that were significantly upregulated on days 4, 7, and 21 after ICH, respectively (P < 0.05). Particularly, collagens, integrins, and matrix metalloproteinases (MMPs) were significantly increased on day 4 post-ICH and continued to increase at the other time points. Western blot and immunohistochemistry analyses showed a comparable trend in the upregulation of MMPs. Compared to the sham group, FITC-dextran labelling demonstrated decreased perfusion and increased vascular permeability in the perihematomal region in the ICH group. Doxycycline, an MMP inhibitor, significantly reduced angiogenesis (P < 0.05). The results of this study indicate that MMPs are involved in modulating angiogenesis following ICH.