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1.
Growth and characterization of epithelial cells from normal human uterine ectocervix and endocervix 总被引:3,自引:0,他引:3
M. E. Turyk T. R. Golub N. B. Wood J. L. Hawkins G. D. Wilbanks 《In vitro cellular & developmental biology. Plant》1989,25(6):544-556
Summary The human uterine cervix consists of an endocervical canal lined with a single layer of columnar mucus-secreting cells and
an outer ectocervix covered by a stratified squamous epithelium. We report here the culture of human endocervical epithelial
cells (HEnE) and human ectocervical epithelial cells (HEcE) in serum-free medium (KGM). Both HEnE and HEcE cultures were composed
of keratinocytelike cells which formed desmosomal contacts and stratified in the presence of high concentrations of calcium
ions. Cells with a pleomorphic epithelial morphology were observed in HEnE cultures, but not in HEcE cultures. Keratin 18,
which is characteristic of endocervix in vivo, was detected by indirect immunofluorescent staining in all HEnE cells but was
never detected in cultured HEcE. HEcE expressed keratin 13 which is characteristic of ectocervix in vivo. Although keratin
13 was never detected in primary HEnE cultures, it was expressed in passaged HEnE cultures grown in medium with high concentrations
of calcium and in late passage HEnE cultures. HEnE underwent an average of 15.1 population doublings during serial culture.
Mean colony-forming efficiency during Passages 2 to 3 was 14.7% and mean population doubling time was 17.8 h. HEcE cultures
underwent significantly more population doublings (29.0) than HEnE cultures, whereas colony-forming efficiencies and doubling
times were similar to those determined for HEnE. HEnE and HEcE cells may be useful in developing in vitro models of cervical
squamous metaplasia and for exploring the interactions between target cell differentiation, carcinogens, and papillomaviruses
in the development of cervical neoplasia.
This study was supported in part by the Rush University Committee on Research and by the Lester B. and Francis Knight and
the S. Charles and Marsha Papageorge research funds. 相似文献
2.
Neisseria gonorrhoeae (GC) establishes infection in women from the cervix, lined with heterogeneous epithelial cells from non-polarized stratified at the ectocervix to polarized columnar at the endocervix. We have previously shown that GC differentially colonize and transmigrate across the ecto and endocervical epithelia. However, whether and how GC invade into heterogeneous cervical epithelial cells is unknown. This study examined GC entry of epithelial cells with various properties, using human cervical tissue explant and non-polarized/polarized epithelial cell line models. While adhering to non-polarized and polarized epithelial cells at similar levels, GC invaded into non-polarized more efficiently than polarized epithelial cells. The enhanced GC invasion in non-polarized epithelial cells was associated with increased ezrin phosphorylation, F-actin and ezrin recruitment to GC adherent sites, and the elongation of GC-associated microvilli. Inhibition of ezrin phosphorylation inhibited F-actin and ezrin recruitment and microvilli elongation, leading to a reduction in GC invasion. The reduced GC invasion in polarized epithelial cells was associated with non-muscle myosin II-mediated F-actin disassembly and microvilli denudation at GC adherence sites. Surprisingly, intraepithelial GC were only detected inside epithelial cells shedding from the cervix by immunofluorescence microscopy, but not significantly in the ectocervical and the endocervical regions. We observed similar ezrin and F-actin recruitment in exfoliated cervical epithelial cells but not in those that remained in the ectocervical epithelium, as the luminal layer of ectocervical epithelial cells expressed ten-fold lower levels of ezrin than those beneath. However, GC inoculation induced F-actin reduction and myosin recruitment in the endocervix, similar to what was seen in polarized epithelial cells. Collectively, our results suggest that while GC invade non-polarized epithelial cells through ezrin-driven microvilli elongation, the apical polarization of ezrin and F-actin inhibits GC entry into polarized epithelial cells. 相似文献
3.
Cell-mediated immunity (CMI) is key to defense against intracellular pathogens such as Chlamydia trachomatis and viruses that infect the lower female genital tract, but little is known about CMI at this site. Recent studies indicate that there are immunological microenvironments within the female genital tract, and that immune functions are affected by hormones as well as infections and inflammatory processes. To determine the distribution of mediators of CMI within the lower female genital tract, we have enumerated and characterized T-lymphocyte subsets and natural killer and antigen presenting cells (APCs; macrophages and dendritic cells) in the introitus, vagina, ectocervix, endocervix and cervical transformation zone (TZ) from healthy women, and have examined the effects of the menstrual cycle, menopause and inflammation on these parameters. In women without inflammation, T cells and APCs were most prevalent in the cervical TZ and surrounding tissue. Intraepithelial lymphocytes were predominantly CD8+ T cell+; most CD8+ cells in the TZ and endocervix, and a proportion of cells in the ectocervix, expressed T-cell internal antigen-1, a marker of cytotoxic potential. In contrast, the normal vaginal mucosa contained few T cells and APCs. Cervicitis and vaginitis cases had increased numbers of intraepithelial CD8+ and CD4+ lymphocytes and APCs. The menstrual cycle and menopause had no apparent effect on cellular localization or abundance in any of the lower genital tract tissues. These data indicate that the cervix, especially the TZ, is the major inductive and effector site for CMI in the lower female genital tract. Because CD4+ T cells and APCs are primary host cells for human immunodeficiency virus type 1 (HIV-1), these data also provide further evidence that the cervix is a primary infection site of HIV-1, and that inflammation increases the risk of HIV transmission. 相似文献
4.
5.
Simon-Assmann P Turck N Sidhoum-Jenny M Gradwohl G Kedinger M 《Cell biology and toxicology》2007,23(4):241-256
The intestinal epithelium is a particularly interesting tissue as (1) it is in a constant cell renewal from a stem cell pool
located in the crypts which form, with the underlying fibroblasts, a stem cell niche and (2) the pluripotent stem cells give
rise to four main cell types: enterocytes, mucus, endocrine, and Paneth cells. The mechanisms leading to the determination
of phenotype commitment and cell-specific expressions are still poorly understood. Although transgenic mouse models are powerful
tools for elucidating the molecular cascades implicated in these processes, cell culture approaches bring easy and elegant
ways to study cellular behavior, cell interactions, and cell signaling pathways for example. In the present review, we will
describe the major tissue culture technologies that allow differentiation of epithelial cells from undifferentiated embryonic
or crypt cells. We will point to the necessity of the re-creation of a complex microenvironment that allows full differentiation
process to occur. We will also summarize the characteristics and interesting properties of the cell lines established from
human colorectal tumors. 相似文献
6.
Subtyping of epithelial cells of normal and metaplastic human uterine cervix, using polypeptide-specific cytokeratin antibodies 总被引:10,自引:0,他引:10
Rivka Levy Bernard Czernobilsky Benjamin Geiger 《Differentiation; research in biological diversity》1988,39(3):185-196
The aim of the present study was to explore the histogenesis of metaplastic cells in the human uterine cervix. In a previous study we demonstrated that squamous cervical metaplasia expresses a unique set of cytokeratin polypeptides different from that expressed by the various normal epithelial elements of both the exo- and endocervix. It was thus proposed that the formation of squamous metaplasia represented a new route of differentiation. In the present study we further investigated this aspect by expanding the battery of monoclonal antibodies directed against specific cytokeratin epitopes used for immunohistochemical labelling. The antibodies used were: KS-1 A3, which specifically stains cytokeratin polypeptide no. 13; antibody KS-2.1, which is an anti-cytokeratin reacting with pseudostratified transitional and some simple epithelia; and antibody KS-B17.2 reacting with cytokeratin polypeptide no. 18. Examination of the staining patterns obtained with these antibodies revealed specific staining of ciliated cells with antibody KS-2.1 and of endocervical reserve cells with antibody KS-1A3. In 6 out of 19 cases tested reserve cells were also stained with antibody KS-2.1. These results enabled us to distinguish between at least four types of cells residing within the simple epithelium of the endocervix, namely columnar nonciliated cells, ciliated cells, and two subpopulations of reserve cells. Since metaplasia was positively stained by antibodies KS-1A3 and KS-2.1, we propose that the endocervical reserve cells that express cytokeratin polypeptide no. 13 are most probably the cells from which endocervical metaplasia is derived. 相似文献
7.
8.
R. Niles K. C. Kim B. Hyman T. Christensen K. Wasano J. Brody 《In vitro cellular & developmental biology. Plant》1988,24(5):457-463
Summary Studies on the regulation of differentiation in airway epithelial cells have been hampered by the lack of cell culture systems
that differentiate in vitro. One such system that does exhibit differentiation is hamster tracheal epithelial cells (HTE).
A major problem with this system, however, is that at the time cells differentiate, they lyze the collagen gel upon which
they grow, resulting in termination of the culture. Here we report that by growing the HTE cells at 32° instead of 37°C we
can totally prevent lysis of the collagen gel. Cells grown at this lower temperature maintain their differentiated phenotype
as evidenced by abundant mucus granules and the secretion of authentic mucus glycoproteins into the culture media. We have
also developed a method for subculturing the primary cells which allows growth and differentiation in secondary culture. The
HTE cells were capable of being passaged at least three times and did not become transformed as judged by their inability
to grow in soft agar and to produce tumors in syngeneic animals. This improved HTE cell culture system will allow detailed
studies on the mechanisms which regulate growth, differentiation, and mucus secretion in surface airway epithelial cells.
This work was supported in part by grants HL-19717 and HL-36854 from the National Institutes of Health, Bethesda, MD. 相似文献
9.
Mammary epithelium differentiates in a stromal milieu of adipocytes and fibroblasts. To investigate cell-cell interactions that may influence mammary epithelial cell differentiation, we developed a co-culture system of murine mammary epithelium and adipocytes and other fibroblasts. Insofar as caseins are specific molecular markers of mammary epithelial differentiation, rat anti-mouse casein monoclonal antibodies were raised against the three major mouse casein components to study this interaction. Mammary epithelium from mid-pregnant mice was plated on confluent irradiated monolayers of 3T3-L1 cells, a subclone of the Swiss 3T3 cell line that differentiates into adipocytes in monolayer culture and other cell monolayers (3T3-C2 cells, Swiss 3T3 cells, and human foreskin fibroblasts). Casein was synthesized by mammary epithelium only in the presence of co-cultured cells and the lactogenic hormone combination of insulin, hydrocortisone, and prolactin. Synthesis and accumulation of alpha-, beta-, and gamma-mouse casein within the epithelium was shown by immunohistochemical staining of cultured cells with anti-casein monoclonal antibodies, and the specificity of the immunohistochemical reaction was demonstrated using immunoblots. A competitive immunoassay was used to measure the amount of casein secreted into the culture medium. In a 24-h period, mammary epithelium co-cultured with 3T3-L1 cells secreted 12-20 micrograms beta-casein per culture dish. There was evidence of specificity in the cell-cell interaction that mediates hormone-dependent casein synthesis. Swiss 3T3 cells, newborn foreskin fibroblasts, substrate-attached material ("extracellular matrix"), and tissue culture plastic did not support casein synthesis, whereas monolayers of 3T3-L1 and 3T3-C2 cells, a subclone of Swiss 3T3 cells that does not undergo adipocyte differentiation, did. We conclude that interaction between mammary epithelium and other specific nonepithelial cells markedly influences the acquisition of hormone sensitivity of the epithelium and hormone-dependent differentiation. 相似文献
10.
Sherwood Githens Jane A. Schexnayder Randy L. Moses Gerene M. Denning Jeffrey J. Smith Marsha L. Frazier 《In vitro cellular & developmental biology. Animal》1994,30(9):622-635
Summary Most of the pancreatic exocrine epithelium consists of acinar and intralobular duct (ductular) cells, with the balance consisting
of interlobular and main duct cells. Fragments of mouse acinar/ductular epithelium can be isolated by partial digestion with
collagenase and purified by Ficoll density gradient centrifugation. We investigated whether previously developed culture conditions
used for duct epithelium would result in the selective survival and proliferation of ductular cells from the acinar/ductular
fragments. The fragments were cultured on nitrocellulose filters coated with extracellular matrix. After 2 to 4 wk the filters
were covered with proliferating cells resembling parallel cultures of duct epithelium by the following criteria: protein/DNA
ratio, light and electron microscopic appearance, the presence of duct markers (carbonic anhydrase [CA] activity, CA II mRNA,
the cystic fibrosis transmembrane conductance regulator), the near absence of acinar cell markers (amylase and chymotrypsin),
a similar polypeptide profile after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the presence of spontaneous
and secretin-stimulated electrogenic ion transport. Both duct and ductular epithelia formed fluid-filled cysts in collagen
gels and both could be subcultured. We conclude that acinar/ductular tissue gives rise to ductular cells in culture by some
combination of acinar cell death and/or transdifferentiation to a ductular phenotype, accompanied by proliferation of these
cells and preexisting ductular cells. These cultures may be used to investigate the properties of this part of the pancreatic
duct system, from which most of the pancreatic juice water and electrolytes probably originates. 相似文献
11.
A method for the rapid establishment of normal adult mammalian colonic epithelial cell cultures 总被引:6,自引:0,他引:6
Alda Vidrich Rajeswari Ravindranath Kianbanoo Farsi Stephan Targan 《In vitro cellular & developmental biology. Plant》1988,24(3):188-194
Summary Normal colonic epithelial cell cultures of mammalian origin are required to facilitate the study of both normal cellular functions
as well as pathogenesis of certain (human) colonic diseases. To date, little information is available regarding the growth
requirements of colonic epithelial cells in culture of eitehr animal or human origin. Such data would enable the development
of a long-term culture system for these cells. In this study, we present methodology that results in the establishment of
homogeneous cultures of adult rabbit colonic epithelial reproducibly, quickly, and in quantity. The epithelial nature of the
cultures is unambiguously established by intermediate filament typing using antikeratin antibodies. Such culutres can now
be used for a variety of functional studies as well as to investigate the growth requirements of colonic epithelial in culture.
This work was supported by the Blinder Foundation for Crohn’s Disease Research, Harbor UCLA IBD Center (AM 36200) and grant
AM 27806 from the National Institutes of Health, Bethesda, MD. 相似文献
12.
Mechanisms controlling epithelial proliferation and differentiation in the prostate have been primarily investigated in mouse models. The regulation of proliferation and differentiation is poorly understood in human prostate epithelial cells. In vivo, the glandular prostate epithelium consists of a p63-positive proliferating basal cell layer and a post-mitotic p27-positive secretory cell layer. We have established an organized stratified culture system of human primary prostate epithelial cells to gain insight into mechanisms regulating proliferation and differentiation. In this system, expression of p63 is observed in the bottom layer. In addition, BrdU incorporation persists even though cells are confluent. In contrast, in the upper layer, p63 expression is greatly diminished, p27 is expressed, and the cells are growth arrested. Overexpression of cyclin D1 or knockdown of p27 does not increase proliferation. After inactivation of the nuclear phosphoprotein Rb, the cell layers remain organized and cell proliferation increases only in the bottom layer. Furthermore, the expression of p63 remains confined to the bottom layer after Rb inactivation. Altogether, this in vitro model recapitulates certain aspects of in vivo growth regulation and differentiation and suggests that the loss of Rb family proteins in human cells trigger hyperplasia but is not sufficient for transformation.This work was supported by the Departments of Pathology and Urology at Weill Medial College, by grants DAMD-17-02-1-0159, MEDC-GR-355, and P30 CA015704-30, and by grant RO1CA84069 to B.E.C. 相似文献
13.
14.
Latimer JJ Johnson JM Miles TD Dimsdale JM Edwards RP Kelley JL Grant SG 《Cell and tissue research》2008,333(3):461-467
DNA repair, a fundamental function of cellular metabolism, has long been presumed to be constitutive and equivalent in all cells. However, we have previously shown that normal levels of nucleotide excision repair (NER) can vary by 20-fold in a tissue-specific pattern. We have now successfully established primary cultures of normal ovarian tissue from seven women by using a novel culture system originally developed for breast epithelial cells. Epithelial cells in these cultures aggregated to form three-dimensional structures called "attached ovarian epispheres". The availability of these actively proliferating cell cultures allowed us to measure NER functionally and quantitatively by the unscheduled DNA synthesis (UDS) assay, a clinical test used to diagnose constitutive deficiencies in NER capacity. We determined that ovarian epithelial cells manifested an intermediate level of NER capacity in humans, viz., only 25% of that of foreskin fibroblasts, but still 2.5-fold higher than that of peripheral blood lymphocytes. This level of DNA repair capacity was indistinguishable from that of normal breast epithelial cells, suggesting that it might be characteristic of the epithelial cell type. Similar levels of NER activity were observed in cultures established from a disease-free known carrier of a BRCA1 truncation mutation, consistent with previous normal results shown in breast epithelium and blood lymphocytes. These results establish that at least three "normal" levels of such DNA repair occur in human tissues, and that NER capacity is epigenetically regulated during cell differentiation and development. 相似文献
15.
Vitamin D regulates human ectocervical epithelial cell proliferation and insulin-like growth factor-binding protein-3 level 总被引:2,自引:0,他引:2
The differentiation status of the cervical epithelial cell has an important influence on responsiveness to estrogens and progestins. Several agents, including glucocorticoids and retinoids, are known to influence cervical cell differentiation. However, the effects of vitamin D have not been examined. Vitamin D is known to regulate cell proliferation and gene expression in a variety of epithelial cells. In the present study we investigated the ability of 1alpha25-dihydroxyvitamin D3 (D3) to regulate cell proliferation and expression of insulin-like growth factor-binding protein-3 (IGFBP-3) in human ectocervical epithelial cells. ECE16-1, a non-tumorigenic cervical cell line, was growth inhibited by D3 with maximal inhibition at 1000 nM. IGFBP-3 levels increased in parallel with the growth inhibition. IGFBP-3 levels were half-maximally increased at approximately 10-100 nM and maximally increased (10- to 30-fold) at 1000 nM D3. These studies show that vitamin D regulates cervical epithelial cell gene regulation and cell proliferation and that IGFBP-3 may be an in vivo marker of vitamin D action in the cervix. 相似文献
16.
S V Petrov N T Ra?khlin R S Nizamov 《Biulleten' eksperimental'no? biologii i meditsiny》1992,113(5):534-536
Epitope localization reacting with mice monoclonal antibodies (Mabs) H 4 was investigated using the specimen of epithelium of skin, human uterine cervix as well as in the culture of epithelium cells of guinea-pig duct deferent. Mice monoclonal antibodies against antigen H 4 obtained by the hybridoma method after immunization of mice with rat colon epithelium cytoskeletal fractions were used. Mabs H 4 were shown to react with antigen of intermediate filaments of all studied normal epithelial, cancer cells as well as culture epithelial cells. Mabs H 4 are supposed to be used as a unique immunohistochemical marker of epithelium cells under normal and malignant growth conditions. 相似文献
17.
Terstegge S Laufenberg I Pochert J Schenk S Itskovitz-Eldor J Endl E Brüstle O 《Biotechnology and bioengineering》2007,96(1):195-201
Embryonic stem cell (ESC) technology provides attractive perspectives for generating unlimited numbers of somatic cells for disease modeling and compound screening. A key prerequisite for these industrial applications are standardized and automated systems suitable for stem cell processing. Here we demonstrate that mouse and human ESC propagated by automated culture maintain their mean specific growth rates, their capacity for multi-germlayer differentiation, and the expression of the pluripotency-associated markers SSEA-1/Oct-4 and Tra-1-60/Tra-1-81/Oct-4, respectively. The feasibility of ESC culture automation may greatly facilitate the use of this versatile cell source for a variety of biomedical applications. 相似文献
18.
S. J. Clayton F. Walker 《Virchows Archiv. B, Cell pathology including molecular pathology》1983,43(1):63-66
The distribution of amyloid P component in normal human adult cervix was studied using fluorescent immunohistochemical techniques on frozen sections. Amyloid P component is associated with elastic fibres which are particularly concentrated in a sub-epithelial plexus in the ectocervix. This plexus does not extend into the endocervix but terminates at, or just caudal to, the squamocolumnar junction. Amyloid P component was not demonstrated in any of the epithelial basement membranes. 相似文献
19.
OBJECTIVE: To investigate the possibility of a correct cytologic diagnosis of cervical and endometrial carcinoma with other genital organ involvement. STUDY DESIGN: From uteri removed during hysterectomy due to cervical (33 cases) and endometrial (44 cases) cancer, samples were taken by cytobrush or spatula from the ectocervix, endocervix and endometrium of uteri opened longitudinally. Smears and cytosediments were stained by the Papanicolaou polychrome method. Moreover, acid beta-galactosidase activity was demonstrated in serial cytosediments by the indigogenic method of Lojda. From quenched tissue samples taken from the same sites as those for cytology, a series of cryostat sections was prepared and stained by hematoxylin and eosin or azure A, or subjected to the reaction for acid beta-galactosidase. RESULTS: In 17 of 33 patients with cervical cancer, the same type of cancer was also found in smears of the endocervix and endometrium. In six patients the type of cancer was different. Of 44 patients with endometrial cancer, 16 had an endocervical malignancy of the same type. In seven cases the type of cancer was different. The reaction for acid beta-galactosidase helped in the differentiation between squamous (negative reaction in cancer cells) and cylindrocellular (positive reaction) cancer in cytologic preparations. CONCLUSION: Before treatment, it is necessary to determine if there is involvement of the endocervix in endometrial cancer and of the endometrium in cervical cancer. Routine cytologic examination supplemented by the reaction for acid beta-galactosidase proved to be useful for this purpose. 相似文献
20.
Kathryn A. Elliget Benjamin F. Trump 《In vitro cellular & developmental biology. Animal》1991,27(9):739-748
Summary Normal rat kidney proximal tubule epithelial cell cultures were obtained by collagenase digestion of cortex and studied for
10 days. To assess the purity of the seeding suspension, we histochemically demonstrated γ-glutamyltranspeptidase in >95%
of the starting material. To identify cell types in cultures, we investigated several markers. Cells stained positively for
lectinArachis hypogaea (rat proximal tubule) and negatively forLotus tetragonolobus (rat distal tubule). Intermediate filament expression of cytokeratin confirmed the epithelial differentiation of the cultured
cells. Using indirect immunofluorescence, we found that cultures were negative for vimentin and Factor VIII. Cells exhibited
activities of two brush border enzymes, γ-glutamyltranspeptidase and leucine aminopeptidase, and Na+-dependent glucose transport activity. Multicellular domes were evident in the Week 2 of culture. Proliferation was studied
by comparing growth factor-supplemented serum-free medium to cells grown in serum; growth enhancers included insulin, hydrocortisone,
transferrin, glucose, bovine albumin, and epidermal growth factor. Cells proliferate best in medium with 5 or 10% serum and
in serum-free medium supplemented with insulin, hydrocortisone, transferrin, glucose, and bovine albumin. Proliferation was
assessed by determining cell number (population doublings). By light microscopy, the cells were squamous with numerous mitochondria,
a central nucleus, and a rather well-defined homogeneous ectoplasm. By electron microscopy, the cells were polarized with
microvilli and cell junctions at the upper surface and a thin basal lamina toward the culture dish. These data show that the
proximal tubule epithelial cells retain a number of functional characteristics and that they represent an excellent model
for studies of normal and abnormal biology of the renal proximal tubule epithelium.
This project was supported by grant 2-R01-DK15440-16A1 from the National Institutes of Health, Bethesda, MD, and by grant
N0001 4-88-K-0427 from the Department of the Navy, Washington, DC. 相似文献