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1.
Induction of transforming growth factor-beta 1 on dentine pulp cells in different culture patterns 总被引:5,自引:0,他引:5
Nie X Tian W Zhang Y Chen X Dong R Jiang M Chen F Jin Y 《Cell biology international》2006,30(4):295-300
Recent studies have documented that TGF-beta1 takes part in dental pulp tissue repair. Moreover, dental pulp cells have the potential to differentiate into odontoblast-like cells and produce reparative dentine in this process. However, the molecular mechanisms and potential interactions between TGF-beta1 and dental pulp cells are not clear due to the complexity of the pulp/dentine microenvironment. In this study, we investigated the induction of TGF-beta1 on the dental pulp cells in cell culture, tissue culture and three-dimensional culture patterns. These results demonstrated that TGF-beta1 significantly increased the proliferation of cells and activity of ALPase. Dental pulp cells cultured in the presence of TGF-beta1 formed mineralization nodules. In the organ culture, dental pulp cells treated with TGF-beta1 differentiated into odontoblast-like cells and formed a pulp-dentinal complex; and TGF-beta1 significantly induced synthesis of dentine relative proteins DSPP, DMP-1. The dental pulp cells share some characteristics of the odontoblast, such as a parallel arrangement with columnar form and a unilateral cell process. Together, these data indicate that TGF-beta1 can make dental pulp cells differentiated into odontoblast-like cells and form the pulp-dentinal complex. Moreover, these results suggest that TGF-beta1 is an important regulatory factor in odontoblast differentiation during tooth development and pulp repair. 相似文献
2.
Although the recent advances in stem cell engineering have gained a great deal of attention due to their high potential in clinical research, the applicability of stem cells for preclinical screening in the drug discovery process is still challenging due to difficulties in controlling the stem cell microenvironment and the limited availability of high-throughput systems. Recently, researchers have been actively developing and evaluating three-dimensional (3D) cell culture-based platforms using microfluidic technologies, such as organ-on-a-chip and organoid-on-a-chip platforms, and they have achieved promising breakthroughs in stem cell engineering. In this review, we start with a comprehensive discussion on the importance of microfluidic 3D cell culture techniques in stem cell research and their technical strategies in the field of drug discovery. In a subsequent section, we discuss microfluidic 3D cell culture techniques for high-throughput analysis for use in stem cell research. In addition, some potential and practical applications of organ-on-a-chip or organoid-on-a-chip platforms using stem cells as drug screening and disease models are highlighted. 相似文献
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P. Vuillez F. René M. Plante C. Hindelang M. J. Klein J. M. Félix M. E. Stoeckel 《Cell and tissue research》1992,267(1):169-183
Summary Organotypic cultures, in defined medium, of pituitary primordia obtained from 15-day-old rat fetuses were performed in order to study the in vitro differentiation of melanotrophic cells. The morphological and ultrastructural features of the transplants resembled those of the gland developing in vivo. In situ hybridization on semi-thin sections, using a 35S-labelled oligonucleotide probe, revealed pro-opiomelanocortin-mRNA-containing cells on the first day of culture in the anterior lobe and after 2–3 days in the intermediate lobe. Immunoperoxidase labelling of adjacent sections showed that the same cells reacted with antibodies against -melanocyte-stimulating hormone (MSH), 3 and adrenocorticotropic hormone in both lobes. The pro-opiomelanocortin-mRNA-containing cells formed progressively conspicuous areas in the intermediate lobe, which was almost uniformly labelled after 6 days. In the anterior lobe, these cells remained scattered in small cell groups, and colloidal gold immunolabelling showed the progressive disappearance of MSH labelling from the secretory vesicles in cells exhibiting morphological features of adult corticotrophic cells. Both the MSH content of the explants and MSH release into the culture medium increased with time. Treatment with the dopamine agonist bromocriptine induced a strong dose-dependent decrease in MSH secretion, which was significant after 3 days in culture, indicating that dopamine D2 receptors are able to regulate hormonal release of melanotrophic cells at early stages. This system constitutes a suitable model for further studies of factors controlling cell differentiation and cellular interactions involved in histogenesis. 相似文献
5.
Cartilage tissue differentiation from mesenchymal cells derived from mature muscle in tissue culture 总被引:2,自引:0,他引:2
Marshall R. Urist Yoji Terashima M. Nakagawa Charles Stamos 《In vitro cellular & developmental biology. Plant》1978,14(8):697-706
Summary Under the influence of biochemical components of bone matrix gelatin (BMG), cartilage differentiates in tissue culture from
the connective tissue cell outgrowths of mature muscle. Proliferation and differentiation begin within 24 hr with synthesis
of hyaluronate, continue with high levels of synthesis of DNA and hyaluronidase, and culminate in production of large quantities
of chondroitin sulfate. The addition of hyaluronic acid to the culture medium during the first 48 hr of culture depresses,
whereas chondroitin sulfate enhances, subsequent production of cartilage. These observations on the cell biosynthetic products
prior to the appearance of mature cartilage suggest that the BMG-modified connective tissue outgrowths of mature muscle exhibit
the developmental potential of embryonic axial mesenchyme. Whether muscle harbors embryonic cells in a programmed but not
yet activated readiness (protodifferentiated state) to differentiate into cartilage, or simply contributes a population of
temporarily dedifferentiated fibroblasts, is not known, but in any event, BMG switches the pathway of further development
from fibrous connective tissue to cartilage.
These investigations were supported by grants-in-aid from the USPHS, National Institute of Dental Research (DE-2103-01). Drs.
Terashima and Nakagawa received a research fellowship from the Solo Cup Corporation. Charles Stamos was a Eugene and Marion
Bailey Summer Student Research Fellow. 相似文献
6.
牙本质涎磷蛋白(DSPP)的表达是细胞向成牙本质细胞分化的标志。试图分析人DSPP启动子及构建人DSPP启动子驱动的Lac Z基因表达的报告体系,从而方便快捷检测细胞是否向成牙本质细胞分化。为了建立能表达DSPP的细胞体系,分离了人牙胚间充质细胞,并用地塞米松诱导培养液进行诱导,结果显示,该诱导培养液能有效地诱导人牙胚间充质细胞DSPP基因的表达。利用双荧光素酶报告系统对4段人DSPP基因5′上游区域(-4 000-+54、-2 500-+54、-1 447-+54和-1 027-+54)进行分析,结果显示-2 500-+54区域的启动子活性最高。5′上游区从?2 500 bp延长到?4 000 bp时,启动子活性下降;5′上游区从-2 500 bp缩短至-1 447 bp时,启动子活性下降;再次将-1 447 bp缩短至-1 027 bp时,启动子活性进一步下降。结果暗示在-4 000 bp至-2 500 bp区域存在转录抑制元件,-2 500 bp至-1 027 bp区域存在转录激活元件。用-2 500-+54启动子区域和Lac Z基因构建ph DSPP-Lac Z慢病毒报告载体,并分别在人牙胚间充质细胞和永生化人牙胚间充质细胞系ih EDMC4上检测ph DSPP-Lac Z报告载体的功能,通过X-Gal染色,结果显示在2种细胞牙向分化过程中均可检测到Lac Z基因的表达。研究构建的ph DSPP-Lac Z慢病毒报告载体可为诱导人源细胞牙向分化、牙齿发育、牙齿再生工程等研究中DSPP的表达检测提供一种更加便捷的手段。 相似文献
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FABIANY DA COSTA GONÇALVES MICHELE ARAMBURU SERAFINI HELENA FLORES MELLO BIANCA PFAFFENSELLER ANELISE BERGMANN ARAÚJO FERNANDA VISIOLI ANA HELENA PAZ 《Cytotherapy》2018,20(12):1459-1471
Background aims
Although mesenchymal stromal cells (MSCs) have shown therapeutic potential in intestinal tissue repair, controversy concerning their short survival and poor biodistribution in recipient tissues still remains. Therefore, we investigated the paracrine role of MSC in three-dimensional culture of colon with experimental colitis.Methods
Colitis was induced in mice by oral administration of dextran sulfate sodium (DSS) for 7 days. Inflammatory responses were assessed on the basis of clinical signs, morphological, and histopathological parameters. On days 2 and 5, colonic explants were removed, and a three-dimensional culture was performed. The structural integrity of the intestinal mucosa was tested by treating the cultures with MSC or conditioned medium (CM) for 24 h, and then the colons were analyzed for histology/immunohistochemistry and interleukin (IL)-6 production.Results
Histological analysis demonstrated that both MSC and CM treatment reduced colon damage in organ culture. An increase in cell proliferation (Ki-67 staining) was observed after CM treatment. Additionally, MSC treatment was able to reduce CD3+ cells. The therapeutic effect of MSC and CM was mediated by the downregulation of IL-6.Discussion
The intestinal in vitro model has shown to be potentially useful for studying cellular interactions in a three-dimensional cell arrangement. Moreover, our results provide strong evidence that both MSC and CM treatments can alleviate colonic damage in organ culture. Importantly, these results suggest that MSC-secreted factors are able to protect the colon from inflammation caused by DSS-induced colitis independent of cell transplantation. 相似文献9.
Embryonic stem (ES) cells have indefinite self-renewal ability and pluripotency, and can provide a novel cell source for tissue engineering applications. In this study, a murine CCE ES cell line was used to derive hematopoietic cells in a 3-D fibrous matrix. The 3-D matrix was found to maintain the phenotypes of undifferentiated ES cells as indicated by alkaline phosphatase (ALP) activity and stage specific embryonic antigen-1 (SSEA-1) expression. In hematopoietic differentiation, cells from 3-D culture exhibited similar cell cycle distribution and SSEA-1 expression to those in the initial cell population. The Oct-4 expression was significantly down-regulated, which indicated the occurrence of differentiation, although the level was slightly higher than that in Petri dish culture. The expression of c-kit, cell surface marker for hematopoietic progenitor, was higher in the 3-D culture, suggesting a better-directed hematopoietic differentiation. Cells in the 3-D matrix tended to form large aggregates associated with fibers. For large-scale processes, a perfusion bioreactor can be used for both maintenance and differentiation cultures. As compared to the static culture, a higher growth rate and final cell density were resulted from the perfusion bioreactor due to better control of the reactor environment. At the same time, the differentiation capacity of ES cells was preserved in the perfusion culture. The ES cell culture in the fibrous matrix thus can be used as a 3-D model system to study effects of extracellular environment and associated physico-chemical parameters on ES cell maintenance and differentiation. 相似文献
10.
Platelet-derived growth factor exerts disparate effects on odontoblast differentiation depending on the dimers in rat dental pulp cells 总被引:3,自引:0,他引:3
Yokose S Kadokura H Tajima N Hasegawa A Sakagami H Fujieda K Katayama T 《Cell and tissue research》2004,315(3):375-384
Platelet-derived growth factor (PDGF) has recently been demonstrated to control the expression of alkaline phosphatase and proteoglycan synthesis of odontoblastic cells in dental pulp tissues. Although PDGF appears to be closely related to dentinogenesis, much about the mode of action of PDGF on odontoblast differentiation remains unclear. In this study, we examined the effects of three PDGF dimers (PDGF AA, AB, and BB) on odontoblastic differentiation of dental pulp cells in long-term mineralized cultures. Dental pulp cells isolated from rat lower incisors were continuously treated with each of PDGF AA, AB, and BB in separate cultures for 20 days. The three PDGF dimers suppressed alkaline phosphatase activity, osteocalcin and calcium content, and the formation of dentin-like nodules. The expression of mRNA for dentin sialoprotein (DSP) in the cells was inhibited by PDGF AA treatment, whereas PDGF AB and BB treatment stimulated the expression of DSP, even though the dentin-like nodule formation was inhibited. Although the effects of PDGF on odontoblastic differentiation varied among the dimers, the cells expressed both PDGF and receptors, whose quantities were similar. These results suggest that PDGF exerts diverse effects on odontoblastic differentiation depending on its dimeric form. These in vitro findings explain, at least in part, the in vivo action of PDGF in dentinogenesis during the repair process of damaged dental pulp.This work was supported in part by grants-in-aid from the Ministry of Science, Education, and Culture of Japan 相似文献
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Summary To clarify proliferative activity in the cells of the Rathke's pouch of the rat, we studied the labeling index using bromodeoxyuridine-immunohistochemistry. Rat fetuses were removed 1 h after transplacental injection of bromodeoxyuridine on day 11.5–21.5 of gestation, and were subsequently used to examine cellular proliferation. Although the labeling index within the Rathke's pouch was 30% on day 12.5, it decreased with development and by day 18.5 of gestation had a value of about 5%. The labeling index within Rathke's pouch was not homogeneous throughout the entire pouch, but tended to be higher in regions where cells were more densely packed. This heterogeneous pattern of distribution of labeling index values continued until day 15. On that day, immunoreactive ACTH cells first appeared in the region where the labeling index was low. From day 17 of gestation, the uneven distribution of the labeling index became vague and, simultaneously, the distribution of ACTH cells became homogeneous throughout the pouch. It was concluded that proliferation of the epithelium of Rathke's pouch is heavily involved in the growth of the pouch until at least the appearance of ACTH cells. 相似文献
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Shinji Izumi Motohiro Takeya Katsumasa Takagi Kiyoshi Takahashi 《Cell and tissue research》1990,262(1):1-8
Summary Ontogenetic development of the synovial A cells in fetal rat knee joints was investigated by immunohistochemistry, immuno-electron microscopy, cultivation, and autoradiography. At day 17 of gestation, immature macrophages were first seen in the articular interzone, and thereafter they differentiated into macrophages (synovial A cells), which were found in the synovial intima. The degree of reactivity of macrophages with five monoclonal antibodies increased in the developing synovial membranes of fetal rats as shown by immunohistochemistry. Similar findings were obtained in organ cultures of fetal knee joints. A marked difference of proliferative potential was found between A and B cells during ontogeny. A cells after birth did not incorporate 3H-thymidine in contrast to B cells. Before birth, B cells had a labelling index which was at least five times larger than that of A cells. The results of this study indicate that the synovial A cells are derived from both monocytes and fetal macrophages circulating in peripheral blood and that they differ from the synovial B cells in morphology, differentiation, and proliferative potential. 相似文献
15.
Initial responses of odontoblasts and immunocompetent cells to cavity preparation by laser ablation were investigated in rat molars. In untreated control teeth, intense heat shock protein (Hsp) 25 immunoreactivity was found in the cell bodies of odontoblasts, whereas cells immunopositive for the class II major histocompatibility complex (MHC) antigen were predominantly located beneath the odontoblast layer in the dental pulp. Cavity preparation caused the destruction of the odontoblast layer and the shift of most class-II-MHC-positive cells from the pulp-dentin border toward the pulp core at the affected site. Twelve hours after cavity preparation, numerous class-II-MHC-positive cells appeared along the pulp-dentin border and extended their processes deep into the exposed dentinal tubules, but subsequently disappeared from the pulp-dentin border together with Hsp-25-immunopositive cells by 24 h after the operation. By 3–5 days postoperation, distinct abscess formation consisting of polymorphonuclear leukocytes was found in the dental pulp. The penetration of masses of oral bacteria was recognizable in the dentinal tubules beneath the prepared cavity. These findings indicate that cavity preparation by laser ablation induces remarkable inflammation by continuous bacterial infections via dentinal tubules in this experimental model, thereby delaying pulpal regeneration.This work was supported by Grant-in-Aid for Scientific Research to promote 2001-Multidisciplinary Research Projects in 2001–2005, and KAKENHI (C) (nos. 12671765 and 14571727 to H.O.) from MEXT 相似文献
16.
Summary The proliferation rate of differentiating fetal rat adrenocortical cells was studied in primary culture. In this system, stimulation with ACTH induces differentiation of zona glomerulosa-like cortical cells into zona fasciculata-like cells. Incorporation of bromodeoxyuridine (BrdU) was studied immunocytochemically by use of anti-BrdU antibody, and the proliferation rate was counted from the monolayer colonies of adrenocortical cells. After 21 days of cultivation in the absence of ACTH, the proliferation rate of zona glomerulosa-like cells was 10%. The rate slowly declined to 1% at the age of 100 days during continuous cultivation in the absence of ACTH. Stimulation with ACTH induced a strong inhibition in the proliferation rate (down to 2% during the first 24 h). Treatment with ACTH during the following 48 h led to an extremely intense proliferation of adrenocortical cells at a proliferation rate of 25%. Continuous treatment with ACTH up to 100 days led to a persistent growth of adrenocortical cells, and a proliferation rate over 2-fold higher than in control cells cultivated in the absence of ACTH. Thus, ACTH is the principal growth-promoting factor also in vitro, as has been found in in vivo studies. This growth effect is mediated by a biphasic course; at the beginning of differentiation the effect is inhibitory and is followed by a persistent stimulation of the growth of adrenocortical cells. 相似文献
17.
Gregory J. Nierode Sneha Gopal Paul Kwon Douglas S. Clark David V. Schaffer Jonathan S. Dordick 《Biotechnology and bioengineering》2019,116(1):168-180
Identification of conditions for guided and specific differentiation of human stem cell and progenitor cells is important for continued development and engineering of in vitro cell culture systems for use in regenerative medicine, drug discovery, and human toxicology. Three-dimensional (3D) and organotypic cell culture models have been used increasingly for in vitro cell culture because they may better model endogenous tissue environments. However, detailed studies of stem cell differentiation within 3D cultures remain limited, particularly with respect to high-throughput screening. Herein, we demonstrate the use of a microarray chip-based platform to screen, in high-throughput, individual and paired effects of 12 soluble factors on the neuronal differentiation of a human neural progenitor cell line (ReNcell VM) encapsulated in microscale 3D Matrigel cultures. Dose–response analysis of selected combinations from the initial combinatorial screen revealed that the combined treatment of all-trans retinoic acid (RA) with the glycogen synthase kinase 3 inhibitor CHIR-99021 (CHIR) enhances neurogenesis while simultaneously decreases astrocyte differentiation, whereas the combined treatment of brain-derived neurotrophic factor and the small azide neuropathiazol enhances the differentiation into neurons and astrocytes. Subtype specification analysis of RA- and CHIR-differentiated cultures revealed that enhanced neurogenesis was not biased toward a specific neuronal subtype. Together, these results demonstrate a high-throughput screening platform for rapid evaluation of differentiation conditions in a 3D environment, which will aid the development and application of 3D stem cell culture models. 相似文献
18.
The effects of gonadotropin-releasing hormone (GnRH) on the development of gonadotropes were investigated by the use of organ culture and by means of immunocytochemistry and radioimmunoassay. Pituitary primordia from rat fetuses were cultured in a medium with or without 10-9 M GnRH during the first 24 h of culture. The ratio of the number of immunoreactive LH cells to the total number of cells in the explants derived from 13.5-day fetuses was increased by the GnRH treatment after 6 or 8 days of culture, while the total number of cells was not altered. LH released into the medium and LH content of explants were not affected by the GnRH treatment. Subsequent treatment with 10-9 M GnRH for 4 h after 7 days of culture resulted in a marked release of LH, accompanying a significant decline in LH content, in both explants exposed or unexposed to the first GnRH treatment. However, the former explants contained a lower amount of LH than the latter explants. The present results indicate that pituitary primordia at 13.5 days of gestation are capable to respond to GnRH, and that GnRH is effective in stimulating the responsiveness of gonadotropes to GnRH during early pituitary cytodifferentiation. 相似文献
19.
Colonization of rat thymic anlage by the first wave of hemopoietic precursor cells (HPc) was investigated by means of transmission
electron microscopy and immunocytochemistry. HPc began migration into the thymic anlage between 13 and 13.5 gestation days
(GD), terminated colonization at about GD 16, and migrated sequentially through the two compartments of the thymic anlage
under the control of typical populations of stromal cells. First, HPc migrated through the external compartment of the perithymic
mesenchyme, tightly interconnected with fibroblasts. The type of junctions between the cells indicated that the fibroblasts
played a role in the control of HPc trafficking and in their entrance to the epithelial compartment. The second stage of colonization
was initiated by the entrance of HPc to the epithelial compartment and their interaction with thymic epithelial cells (TECs).
Based on morphological criteria, two populations of HPc were distinguished that colonized the anlage at various stages of
its development. The predominant population with ultrastructural traits common to thymocytes “homed” into the epithelial type
primordium. A small number of HPc, identified by protein S-100 expression and by Birbeck’s granules as precursors of dendritic
cells, colonized lymphoepithelial anlage in which subsets of cortical and medullary TECs could be distinguished. Thymocyte
migration and their reciprocal interactions with cortical TECs differed from the trafficking of dendritic cells toward the
medulla. The results demonstrated the influence of maturing thymocytes on the development of cortical epithelial cells and
the dynamic organization of the medullary microenvironment with direct involvement of dendritic cells.
This study was supported by UMS grant 501-2-0003404. 相似文献
20.
Biochemical analysis of selenoprotein expression in brain cell lines and in distinct brain regions 总被引:1,自引:0,他引:1
Hoppe B Bräuer AU Kühbacher M Savaskan NE Behne D Kyriakopoulos A 《Cell and tissue research》2008,332(3):403-414
Selenium is present in various biologically important selenoproteins. The preferential incorporation of selenium into the brain indicates its significance for this organ, but so far knowledge concerning the cerebral selenoproteome is scarce. We therefore investigated the expression of selenoproteins in various regions of the rat brain, various subcellular fractions and several brain cell lines by (75)Se-labelling, gel electrophoretic separation and autoradiography, with the (75)Se tracer as the selenoprotein marker. Quantitative evaluation of the labelled proteins in selenium-deficient rats revealed information regarding preferentially supplied selenoproteins and their distribution; 21 selenoproteins could be distinguished, among them a novel or modified 15-kDa selenoprotein enriched in the cerebellum cytosol. The selenoproteins differed in the degree of their expression among the brain regions and within a region among the subcellular fractions. Some cell-type-specific selenium-containing proteins were found in the cell lines. Differences in the distribution patterns between mono-cultured and co-cultured endothelial cells and astrocytes showed that mediators produced by other cells could affect the selenoprotein expression of a specific cell-type. This effect might play a role in the uptake and distribution of selenium in the brain but could also be of significance in the selenium metabolism of other tissues. 相似文献