Histomorphological and biochemical differentiation capacity in organotypic co‐cultures of primary gingival cells

Tomakidi, Pascal; Fusenig, Norbert E.; Kohl, Annette; Komposch, Gerda

doi:10.1111/j.1600-0765.1997.tb00549.x

Cited by 65 publications

(79 citation statements)

References 33 publications

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“…Secondly, due to this spatial separation, the interactions between the epithelial keratinocytes and the connective-tissue fibroblasts resemble that under in vivo conditions. [20][21][22] In the case of controls with keratinocytes only, they were handled as described above, but the pillar microarrays were devoid of fibroblasts ( Fig. 3C and 3C1).…”

Section: The Impact Of Gctfs Established On Micropillar Interfaces Onmentioning

confidence: 99%

“…[20][21][22] Studies with an emphasis on biomechanics described lattice contraction by oral or dermal connective-tissue cells or in response to fibroblasts to various growth factors, [23] but usually gave no approximation of the fibroblast-exerted traction forces. In a recently published paper, modeling of a spongy collagen lattice using the Euler column buckling model yielded contractile forces of dermal fibroblasts in a range from 10-41 nN.…”

Section: Introductionmentioning

confidence: 99%

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Connective‐Tissue Fibroblasts Established on Micropillar Interfaces are Pivotal for Epithelial‐Tissue Morphogenesis

Müssig

Steinberg

Schulz

et al. 2008

Adv Funct Materials

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Polydimethylsiloxane (PDMS) pillar arrays are applied as a biomechanical microenvironment to establish gingival connective‐tissue fibroblasts (GCTFs) and to further analyze the pivotal role of GCTFs in epithelial‐tissue morphogenesis. GCTFs are known to exert successful adhesion and growth on fibronectin immobilized on pillar heads, over time, concomitant with the increased gene expression of vimentin and collagen type‐I. GCTF‐populated pillar arrays clearly reveal that epithelial‐tissue morphogenesis of immortalized human gingival keratinocytes (IHGKs), co‐cultured for 7 and 14 days, parallels the in vivo phenotype more closely, when compared with GCTF‐free control arrays. This in vivo‐like phenotype is substantiated by higher mRNA levels for keratin 1, involucrin and filaggrin differentiation markers. Furthermore, it is reflected by a tissue‐specific protein orientation of the aforementioned molecules, and also of the cell‐to‐cell contact forming desmoplakin and the basement membrane constituents, laminin‐5, laminin‐1/10, and collagen type‐IV. These experiments suggest that the in vivo‐like phenotype of the IHGK is governed by the GCTFs growing on the micropillar interfaces. Moreover, they form the basis for the optimization or neogeneration of biomaterials by varying predefined microenvironmetal parameters to achieve an in vivo‐like cell growth and differentiation, indispensable for tissue morphogenesis during regeneration.

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Section: The Impact Of Gctfs Established On Micropillar Interfaces Onmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Connective‐Tissue Fibroblasts Established on Micropillar Interfaces are Pivotal for Epithelial‐Tissue Morphogenesis

Müssig

Steinberg

Schulz

et al. 2008

Adv Funct Materials

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“…Bosc23, Phoenix, and HaCaT were grown in DMEM supplemented with 10% fetal calf serum. Primary human oral fibroblasts were isolated from oral mucosa (49,50) and cultured in the medium described above. Primary human keratinocytes were isolated from skin of adult individuals as previously described (8) and grown together with NIH 3T3 feeder layers in FAD medium containing 3 parts Ham's F12, 1 part DMEM, 5% fetal calf serum, insulin (5 g/ ml), epidermal growth factor (10 ng/ml), cholera toxin (8.4 ng/ml), adenine (24 g/ml), and hydrocortisone (0.4 g/ml).…”

Section: Retroviral Expression Vectorsmentioning

confidence: 99%

The E6 and E7 Proteins of the Cutaneous Human Papillomavirus Type 38 Display Transforming Properties

Caldeira¹,

Zehbe²,

Accardi³

et al. 2003

J Virol

168

214

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Several studies have suggested the involvement of cutaneous human papillomaviruses (HPVs) in the development of nonmelanoma skin cancers. Here we have characterized the in vitro properties of E7 proteins of three cutaneous HPV types, 10, 20, and 38, which are frequently detected in skin specimens. We show that HPV38 E7 is able to inactivate the tumor suppressor pRb and induces loss of G 1 /S transition control, a key event in carcinogenesis. In contrast, HPV10 and HPV20 E7 proteins do not display these in vitro transforming activities. We also show that the two early proteins E6 and E7 of HPV38 are sufficient to corrupt the cell cycle and senescence programs in primary cells, inducing active and long-lasting proliferation of primary human keratinocytes, the natural host cells. Our study shows that E6 and E7 of this cutaneous HPV type have transforming activity in primary human cells, suggesting a role for HPV38 infection in skin carcinogenesis. In further support of such a role, we detected HPV38 DNA in approximately 50% of nonmelanoma skin cancers, but only in 10% of healthy skin specimens (P < 0.001).Nonmelanoma skin cancer is the most frequently occurring malignancy in the Caucasian population (34,38,47). Although these cancers have a good prognosis and are not normally associated with mortality, an increasing incidence of other invasive cancers and cancer mortality following nonmelanoma skin cancers has been reported (17,24,28,29). Several lines of evidence suggest the involvement of an infective agent in the etiology of this condition. Patients suffering from a rare genetic immune suppression termed epidermodysplasia verruciformis and individuals under long-lasting immunosuppression are prone to develop these cancers (21,22,30,37). Epidermodysplasia verruciformis patients are highly susceptible to human papillomavirus (HPV) infections by a specific subgroup of cutaneous HPVs, the so-called epidermodysplasia verruciformis types (e.g., HPV5 and HPV8), that lead to extensive verrucosis of confluent flat warts (22,30,37). In approximately 30% of cases, the HPV lesions develop into multifocal squamous cell carcinomas.Supporting the infectious role of cutaneous HPV types in the tumorigenesis of nonmelanoma skin cancers is the fact that other members of the papillomavirus family are clearly oncogenic (55). Indeed, clinical, epidemiological, and molecular data have demonstrated that mucosal high-risk HPV types (e.g., high-risk HPV16 and HPV18) are the etiological agents of anogenital cancers as well as a subgroup of head and neck cancers (55). The early region of these HPV types encodes two oncoproteins, E6 and E7, which associate with and neutralize the cellular tumor suppressors p53 and retinoblastoma (pRb), respectively (32,36).Independent studies suggest that cutaneous HPV types may also be involved in the development of squamous cell carcinoma and basal cell carcinomas in the general population (6,7,14,43). These indications are based only on studies assessing viral DNA presence in skin tumors by PCR, which ha...

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“…Phoenix cells and human POFs were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum. POFs were purified from oral mucosa as previously described (21,22). High-titer retroviral supernatants were generated by transient transfection of Phoenix cells (amphotropic viruses) (16) and used to infect POFs as previously described (1).…”

Section: Retroviral Expression Vectorsmentioning

confidence: 99%

Human Papillomavirus Type 16 E6 Promotes Retinoblastoma Protein Phosphorylation and Cell Cycle Progression

Malanchi

Accardi

Diehl

et al. 2004

J Virol

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We show that E6 proteins from benign human papillomavirus type 1 (HPV1) and oncogenic HPV16 have the ability to alter the regulation of the G 1 /S transition of the cell cycle in primary human fibroblasts. Overexpression of both viral proteins induces cellular proliferation, retinoblastoma (pRb) phosphorylation, and accumulation of products of genes that are negatively regulated by pRb, such as p16INK4a , CDC2, E2F-1, and cyclin A. Hyperphosphorylated forms of pRb are present in E6-expressing cells even in the presence of ectopic levels of p16INK4a . The E6 proteins strongly increased the cyclin A/cyclin-dependent kinase 2 (CDK2) activity, which is involved in pRb phosphorylation. In addition, mRNA and protein levels of the CDK2 inhibitor p21 WAF1/CIP1 were strongly down-regulated in cells expressing E6 proteins. The down-regulation of the p21 WAF1/CIP1 gene appears to be independent of p53 inactivation, since HPV1 E6 and an HPV16 E6 mutant unable to target p53 were fully competent in decreasing p21 WAF1/CIP1 levels. E6 from HPV1 and HPV16 also enabled cells to overcome the G 1 arrest imposed by oncogenic ras. Immunofluorescence staining of cells coexpressing ras and E6 from either HPV16 or HPV1 revealed that antiproliferative (p16 INK4a ) and proliferative (Ki67) markers were coexpressed in the same cells. Together, these data underline a novel activity of E6 that is not mediated by inactivation of p53.Human papillomaviruses (HPVs) infect keratinocytes in the basal layer of stratified epithelia at a variety of anatomical sites (26). On the basis of their tissue tropism, the HPVs can be subdivided into cutaneous and mucosal types, which infect the skin and the mucosae, respectively. All HPV types are able to induce hyperproliferative benign lesions. Indeed, HPV-induced proliferation is an absolute requirement for completion of the viral life cycle. In addition, certain mucosal types, termed high risk, can promote transformation of a benign lesion into a malignant one (26). HPV16 is the high-risk type most frequently found in premalignant and malignant cervical lesions worldwide (2). Several studies have demonstrated that two viral early proteins, E6 and E7, play a key role in the induction of benign and malignant lesions (23) by associating with several cellular factors and altering their function (11, 13). The best-characterized property of HPV16 E6 is its binding to the p53 tumor suppressor, leading to degradation of the cellular protein via the ubiquitin pathway (20). The role of p53 is to safeguard the integrity of the genome by inducing cell cycle arrest or apoptosis in the presence of DNA damage. Therefore, its inactivation by the E6 protein leads to chromosomal instability and increases the probability of an HPV-infected cell evolving towards malignancy. Several studies have shown that HPV16 E6 is able to associate with other cellular proteins (11), indicating its involvement in additional pathways. Experiments with transgenic mice have shown that E6 can induce epithelial hyperplasia (14,19). It has recen...

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Histomorphological and biochemical differentiation capacity in organotypic co‐cultures of primary gingival cells

Cited by 65 publications

References 33 publications

Connective‐Tissue Fibroblasts Established on Micropillar Interfaces are Pivotal for Epithelial‐Tissue Morphogenesis

Connective‐Tissue Fibroblasts Established on Micropillar Interfaces are Pivotal for Epithelial‐Tissue Morphogenesis

The E6 and E7 Proteins of the Cutaneous Human Papillomavirus Type 38 Display Transforming Properties

Human Papillomavirus Type 16 E6 Promotes Retinoblastoma Protein Phosphorylation and Cell Cycle Progression

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