Recent Articles

3D Cell CultureBelow are the titles and abstracts from the most recent articles published in In Vitro Cellular and Developmental Biology – Animal. Click on the title to view the full article at Springerlink.


  • Cathepsin K degrades osteoprotegerin to promote osteoclastogenesis in vitro
    on January 23, 2023 at 12:00 am

    Abstract Osteoblasts produce the receptor activator of nuclear factor-kappa B ligand (RANKL) and osteoprotegerin, the inducer and the suppressor of osteoclast differentiation and activation. We previously proposed that the degradation of osteoprotegerin by lysine-specific gingipain of Porphyromonas gingivalis and neutrophil elastase is one of the mechanisms of bone resorption associated with infection and inflammation. In the present study, we found that cathepsin K (CTSK) also degraded osteoprotegerin in an acidic milieu and the buffer with a pH of 7.4. The 37 k fragment of osteoprotegerin produced by the reaction with CTSK was further degraded into low molecular weight fragments, including a 13 k fragment, depending on the reaction time. The N-terminal amino acid sequence of the 37 k fragment matched that of the intact osteoprotegerin, indicating that CTSK preferentially hydrolyzes the death domain-like region of osteoprotegerin, not its RANKL-binding […]

  • Mitochondrial transplantation against gentamicin-induced toxicity on rat renal proximal tubular cells: the higher activity of female rat mitochondria
    on January 11, 2023 at 12:00 am

    Abstract Mitochondrial dysfunction is a fundamental mechanism leading to drug nephrotoxicity, such as gentamicin-induced nephrotoxicity. Mitochondrial therapy (mitotherapy) or exogenous mitochondria transplantation is a method that can be used to replace dysfunctional mitochondria with healthy mitochondria. This method can help in the treatment of diseases related to mitochondria. In this research, we studied the transplantation effect of freshly isolated mitochondria on the toxicity induced by gentamicin on renal proximal tubular cells (RPTCs). Furthermore, possible gender-related effects on supplying exogenous rat kidney mitochondria on gentamicin-induced RPTCs were investigated. At first, the normality and proper functioning of fresh mitochondria were assessed by measuring mitochondrial succinate dehydrogenase activity (SDH) and changes in mitochondrial membrane potential (MMP). Then, the protective effects of mitochondrial transplantation against […]

  • SOX2 suppresses osteoblast differentiation of MC3T3-E1 cells through activating the transcription of LGR4
    on December 22, 2022 at 12:00 am

    Abstract Osteogenic differentiation is a crucial process of new bone formation. This study aimed to explore the roles and mechanism of SRY-Box Transcription Factor 2 (SOX2) on proliferation and osteogenic differentiation of MC3T3-E1 cells. Bone morphogenetic protein 2 (BMP2) was used to induce the osteogenic differentiation of MC3T3-E1 cells. The expression of SOX2 was determined by quantitative real-time PCR (RT-PCR) at different time points after induction. The SOX2 overexpression plasmids were constructed and transfected into MC3T3-E1 cells. Osteogenic differentiation was evaluated by Alizarin Red S staining and alkaline phosphatase (ALP) assay. The expressions of osteogenic differentiation markers including runt-related transcription factor 2 (Runx2), osteopontin (OPN), and osteocalcin (OCN) were detected by western blot assay. Luciferase reporter and CHIP assays were used to confirm that SOX2 regulated the transcriptional activation of leucine-rich […]

  • The immunotoxicity of ten insecticides against insect hemocyte cells in vitro
    on December 1, 2022 at 12:00 am

    Abstract Hemocytes in the hemolymph of insects perform innate immunity, but systematic studies to compare immunotoxicity of pesticides on hemocytes are still few. In this study, an insect hemocyte system was used to assess the impact of pesticides with different modes of action, which included loss of cell viability, inhibition of hemophagocytosis, and reduction of nitric oxide synthase (NOS) activity. Results showed that piericidin A was the most cytotoxic to hemocytes, chlorfluazuron and hexaflumuron were the next. Also, piericidin A, chlorfenapyr, and fipronil had strong inhibitory effects on hemophagocytosis, and the effects of piericidin A and chlorfenapyr were persistent, while that of fipronil was short-lived. Moreover, fenoxycarb and hexaflumuron selectively inhibited granulocyte phagocytosis, tebufenozide only showed inhibition on plasmatocyte phagocytosis, but both inhibitory effects were transient. Furthermore, fenoxycarb and hexaflumuron […]

  • Impact of repeated intravenous infusions of umbilical cord–derived versus bone marrow–derived mesenchymal stem cells on angiogenesis in a pregnant experimentally induced deep venous thrombosis rat model
    on December 1, 2022 at 12:00 am

    Abstract Deep venous thrombosis (DVT) therapy during pregnancy warrants special consideration for the woman and the fetus. This study aimed to evaluate the impact of umbilical cord–derived mesenchymal stem cells (UC-MSCs) and bone marrow–derived mesenchymal stem cells (BM-MSCs) in terms of pro-angiogenic capacity and amelioration of pregnancy outcomes. The pregnant DVT rat model was successfully established by the “stenosis” method. Three consecutive injections of both UC-MSCs and BM-MSCs improved angiogenesis and ameliorated the embryo absorption rate in pregnant SD rats with DVT, in which UC-MSCs promoted angiogenesis more significantly. Furthermore, the levels of serum vascular endothelial growth factor-A (VEGF-A) and epidermal growth factor (EGF) were significantly higher in the UC-MSC group compared to those of the BM-MSC group. Thereafter, differentially expressed genes (DEGs) in thrombosed inferior vena cava tissues in the UC-MSC and […]

  • IGF-1 increases survival of CD4+ lineage in a 2D model of thymocyte/thymic stromal cell co-culture
    on December 1, 2022 at 12:00 am

    Abstract Insulin-like growth factor-1 (IGF-1), in addition to its classic effects on cell proliferation and organism growth, has pleiotropic actions on the immune system, particularly on the thymus. Thus, the objective of this study was to evaluate the influence of IGF-1 on molecules involved in the survival of thymocytes in vitro using a co-culture system with thymic stromal cells obtained from C57BL/6 mice. The obtained thymic stroma has contained thymic epithelial cells, macrophages, dendritic cells, fibroblasts, and preserved the expression of the major histocompatibility complex (MHC) molecules. Fresh thymocytes were added to these cultures and the co-culture were treated daily with IGF-1 (100 ng/mL) for 3 days. In this scheme, the viability of the thymocytes was about 70%, either in the control (non-treated cells) or in the IGF-1-treated cultures. It was found that IGF-1 was able to increase the percentage of thymocytes from the CD4+ single-positive […]

  • The Berberis vulgaris L. extract berberine exerts its anti-oxidant effects to ameliorate cholesterol overloading–induced cell apoptosis in the primary mice hepatocytes: an in vitro study
    on December 1, 2022 at 12:00 am

    Abstract Cholesterol overloading stress damages normal cellular functions in hepatocytes and induces metabolic disorders to facilitate the development of multiple diseases, including cardiovascular diseases, which seriously degrades the life quality of human beings. Recent data suggest that the Berberis vulgaris L. extract berberine is capable of regulating cholesterol homeostasis, which is deemed as potential therapeutic drug for the treatment of cholesterol overloading–associated diseases, but its detailed functions and molecular mechanisms are still largely unknown. In the present study, we evidenced that berberine suppressed cell apoptosis in high-cholesterol-diet mice liver and cholesterol-overloaded mice hepatocytes. Also, cholesterol overloading promoted reactive oxygen species (ROS) generation to trigger oxidative damages in hepatocytes, which were reversed by co-treating cells with both berberine and the ROS scavenger N-acetylcysteine (NAC). Moreover, […]

  • Promotion of osteogenesis in BMSC under hypoxia by ATF4 via the PERK–eIF2α signaling pathway
    on December 1, 2022 at 12:00 am

    Abstract Mandibular distraction osteogenesis (MDO) is an endogenous tissue engineering technology in which bone marrow mesenchymal stem cells (BMSC) play a key role in MDO-related osteogenesis. Activating transcription factor 4 (ATF4) is involved in osteogenesis through activation of PERK (Protein kinase R-like endoplasmic reticulum kinase) in endoplasmic reticulum stress (ERS) condition under hypoxia. However, the specific role of ATF4 in MDO with BMSC remains unknown. The aim of this study was to explore the effects of ATF4 in MDO with BMSC under hypoxia. Briefly, canine BMSCs were cultured in a hypoxic chamber, and effects of hypoxia were evaluated using cell migration assay and Alizarin Red S staining. Expression levels of protein kinase R-like endoplasmic reticulum kinase, eukaryotic translation initiation factor 2α, ATF4, osteocalcin, and bone sialoprotein were evaluated using quantitative polymerase chain reaction and western blotting. BMSCs were transduced […]

  • Improved cryopreservation media formulation reduces costs of maintenance while preserving function of genetically modified insect cells
    on December 1, 2022 at 12:00 am

    Abstract Insect cell lines are an invaluable resource that facilitate various fundamental and applied research programs. Genetically engineered insect cell lines, in particular, serve as a platform through which the function of heterologously expressed proteins can be studied. However, a barrier to more widespread utilization and distribution of insect cell lines, genetically modified or not, is the technical and operational challenge associated with traditional cryopreservation methods, including their dependence on the use of liquid nitrogen facilities, animal or human serum products, and relatively high concentrations of permeating cryoprotectants (e.g., DMSO). Recent innovations in cryopreservation technologies have produced reagents with improved abilities to effectively preserve mammalian cell lines for long periods in regular laboratory deep freezers without using serum products, but their effectiveness in preserving genetically engineered insect cell lines […]

  • Benzophenones alter autophagy and ER stress gene expression in pancreatic beta cells in vitro
    on December 1, 2022 at 12:00 am

    Abstract Benzophenones (BPs) are endocrine disruptors frequently used in sunscreens and food packaging as UV blockers. Our goal was to assess the effect of benzophenone 2 (BP2) and 3 (BP3) on gene expression related to autophagy process and ER stress response in pancreatic beta cells. To that end, the mouse pancreatic beta cell line MIN6B1 was treated with 10 µM BP2 or BP3 in the presence or absence of the autophagy-inhibitor chloroquine (CQ, 10 µM) or the autophagy-inducer rapamycin (RAPA, 50 nM) during 24 h. BP3 inhibited the expression of the autophagic gene Ulk1, and additional effects were uncovered when autophagy was modified by CQ and RAPA. BP3 counteracted CQ-induced Lamp2 expression but did not compensate CQ-induced Sqstm1/p62 gene transcription, neither BP2. Nevertheless, the BPs did not alter the autophagic flux. In relation to ER stress, BP3 inhibited unspliced and spliced Xbp1 mRNA levels in the presence or absence of CQ, totally counteracted […]

  • A continuous myofibroblast precursor cell line from the tail muscle of Australasian snapper (Chrysophrys auratus) that responds to transforming growth factor beta and fibroblast growth factor
    on December 1, 2022 at 12:00 am

    Abstract Chrysophrys auratus (Australasian snapper) is one of the largest and most valuable finfish from capture fisheries in New Zealand, yet no cell lines from this species are reported in the scientific literature. Here, we describe a muscle-derived cell line initiated from the tail of a juvenile snapper which has been designated CAtmus1PFR (Chrysophrys auratus, tail muscle, Plant & Food Research). The cell line has been passaged over 100 times in 3 years and is considered immortal. Cells are reliant on serum supplementation for proliferation and exhibit a broad thermal profile comparable to the eurythermic nature of C. auratus in vivo. The impact of exogenous growth factors, including insulin-like growth factors I and II (IGF-I and IGF-II), basic fibroblast growth factor (bFGF), and transforming growth factor beta (TGFβ), on cell morphology and proliferation was investigated. Insulin-like growth factors acted as mitogens and had minimal […]

  • Cell line derived from muscle of Gymnocypris przewalskii, a species of Schizothoracinae in Qinghai Lake, Qinghai–Tibet Plateau
    on December 1, 2022 at 12:00 am

    Abstract Gymnocypris przewalskii (naked carp), a native teleost, plays an important role in the ecosystem of Qinghai Lake (altitude, 3.2 km) on the Qinghai–Tibet Plateau in China. We developed a new cell line from the muscle of G. przewalskii using the explant technique and named the cell line GPM. This cell line was maintained in DMEM medium (high glucose) supplemented with 15% fetal bovine serum (FBS). The cell line was successfully subcultured up to 32 passages and was authenticated by immunofluorescence assay, sequencing the mitochondrial cytochrome C oxidase subunit I (COI) and 16S rRNA genes, and by chromosome analysis. In the medium containing 15% FBS, the cell line could be passaged stably at 25 °C. The GPM cell line could express green fluorescent protein (GFP) with a CMV promoter with about 5% transfection efficiency. MTT tests showed that Clostridium botulinum toxin (BTX) was toxic to the cell line. The cell line could be successfully […]

  • RTG-TOF, a rainbow trout (Oncorhynchus mykiss) cell line with an inducible gene expression system
    on December 1, 2022 at 12:00 am
  • Establishment and characterization of urothelial carcinoma cell lines with and without BRAF mutation (V595E) in dogs
    on December 1, 2022 at 12:00 am

    Abstract Each 5 urothelial carcinoma (UC) cell lines with and without the v-Raf murine sarcoma virus oncogene homolog B (BRAF) gene mutation (V595E) were established and examined V595E-related tumorigenic characteristics in dogs. No typical morphological features were observed in cloned cells with and without V595E. The cell proliferation of both cloned cells showed logarithmic growth curve and those doubling time were 24.9 ± 4.1 h in V595E ( +) and 29.3 ± 11.3 h in V595E ( −). On the growth curve of xenotransplanted tumor in severe combined immunodeficiency mice, 3 out of 5 V595E ( +) and 2 out of 5 V595E ( −) cloned cells revealed gradually and remarkably increasing curve, indicating clearly tumorigenicity. The xenotransplanted tumors with V595E ( +) showed typical features of UC, such as solid proliferation of pleomorphic tumor cells, formation of papillary structure, and glandular structure. Additionally, various vascular […]

  • The preventive use of resveratrol increases its antioxidant effect by SIRT1 and subclinical anti-inflammatory action in Neuro-2A cells
    on December 1, 2022 at 12:00 am

    Abstract Currently, the important role of oxidative stress in the aging process and in neurodegenerative diseases has been highlighted, suggesting the beneficial effect of antioxidants as adjuvant therapy. Resveratrol (RSV) is a polyphenolic compound used in the clinic and has been shown as an antioxidant and anti-inflammatory. Therefore, the objective was to verify neuroprotective and modulating effects of RSV on N2-A cells, pre or post inserted into an oxidative stress environment. For this, two treatment conditions were established: pre-stimulus and post-stimulus. The analysis of AMPK and SIRT1 cell signaling pathways was performed through the chemiluminescence assay using the dorsomorphin and EX527 inhibitors, respectively. The inflammatory profile was also evaluated in these neural cells, through the levels of IL-6, TNF, and IL-10. We observed that RSV in N2-A cells has anti-inflammatory effect and antioxidant property and it mechanism is dependent on the […]

  • In vitro effects of H2O2 on neural stem cell differentiation
    on October 1, 2022 at 12:00 am

    Abstract The development of the CNS is a complex and well-regulated process, where stem cells differentiate into committed cells depending on the stimuli from the microenvironment. Alterations of oxygen levels were stated to be significant in terms of brain development and neurogenesis during embryonic development, as well as the adult neurogenesis. As a product of oxygen processing, hydrogen peroxide (H2O2) has been established as a key regulator, acting as a secondary messenger, of signal transduction and cellular biological functions. H2O2 is involved in survival, proliferation, and differentiation of neural stem cells into committed cells of the CNS. Effects of different concentrations of exogenous H2O2 on neuronal differentiation and the molecular pathways involved are yet to be clearly understood. Here, we investigated the concentration-dependent effects of H2O2 on differentiation of neural stem cells using CGR8 embryonic mouse stem cell line. We have […]

  • Development and characterization of Spodoptera mauritia ovarian primary cell culture and evaluation of fenoxycarb toxicity
    on October 1, 2022 at 12:00 am

    Abstract The rice swarming caterpillar, Spodoptera mauritia (Biosduval, 1833) (Lepidoptera: Noctuidae), has been considered a severe pest and has caused the extreme loss of rice paddies in the nursery stages. The massive reproductive efficiency of pests results from balanced endocrine functioning. Insect treatments with exogenous either juvenile hormones (JH) or juvenile hormone analogues (JHAs) during low endogenous JH titers disrupt metamorphosis and ovarian development. Hence, it was thought worthwhile to develop a primary ovarian cell culture of Spodoptera mauritia to study the biological efficiency of JHA, fenoxycarb, at the cellular level. The study envisioned the cell characteristics and growth properties and found that most cells were spherical. Spindle-shaped cells were also present during the initial stage of active cell division. The majority of the cells grew attached to the bottom of the culture plates, and a few grew in suspension. The cell […]

  • TRIM47 promotes glioma angiogenesis by suppressing Smad4
    on October 1, 2022 at 12:00 am

    Abstract    Angiogenesis is required for tumor progression; thus, its investigation can be useful to identify strategies for potential cancer treatments. Tripartite motif 47 (TRIM47) is involved in the progression of multiple cancers. However, its role in glioma angiogenesis is largely unknown. In this study, we first showed that TRIM47 is frequently upregulated in gliomas, and increased TRIM47 levels are correlated with microvascular density. We then examined the role of TRIM47 in cellular functions related to angiogenesis in vitro and observed that TRIM47 knockdown significantly reduced human umbilical vein endothelial cell proliferation, migration, and tube formation. We also found that TRIM47 silencing reduced vessel density and tumor volume in glioma xenografts. Mechanistically, TRIM47 negatively regulated Smad4 expression in glioma cells, and SMAD4 knockdown rescued the suppressive effects of TRIM47 silencing. Taken together, our results indicate that […]

  • Hace1 overexpression mitigates myocardial hypoxia/reoxygenation injury via the effects on Keap1/Nrf2 pathway
    on October 1, 2022 at 12:00 am

    Abstract HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1 (Hace1) is a crucial mediator of multiple pathological disorders. However, there are few studies regarding the role of Hace1 in myocardial ischemia/reperfusion injury. Here, we studied the functional role of Hace1 on myocardial ischemia/reperfusion injury using hypoxia/reoxygenation (H/R)-injured cardiac cells in vitro. Reduced levels of Hace1 were observed in H/R-exposed cardiac cells. Hace1-overexpressed cardiac cells were resistant to H/R injuries with reduced apoptosis, lowered oxidative stress, and a suppressed inflammatory response. Subsequent analysis revealed that Hace1 overexpression enhanced the activation of nuclear translocation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and increased the transcriptional activity of Nrf2 in H/R-exposed cardiac cells. The knockout of kelch-like ECH-associated protein 1 (Keap1) diminished the regulatory role of Hace1 on Nrf2 […]

  • Feeder cell–dependent primary culture of single blastula–derived embryonic cell lines from marine medaka (Oryzias dancena)
    on October 1, 2022 at 12:00 am

    Abstract Fish embryonic stem cells (ESCs) are derived from blastomeres that have been cultured from blastula embryos. The most widely used method for derivation of fish ESCs is the culture of blastomeres that have been isolated from approximately 10 blastula embryos under feeder-free conditions. However, this method leads to intercellular genetic heterogeneity among the cultured cells, which is a major obstacle to the development of stable ESC culture conditions. In this study, to establish ESC lines with intercellular genetic homogeneity at the early stage of culture, we attempted to derive embryonic cell lines from single blastula–derived blastomeres of marine medaka (Oryzias dancena) in a feeder cell culture system. Using basic fibroblast growth factor–expressing feeder cells during primary culture, we successfully established 22 single blastula–derived embryonic cell lines that could be subcultured more than 20 times. In contrast, we were unable to […]

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