Biomedical subjects

N Forest

Publications and source records attributed to N Forest.

At least 19 recordsLinked to original sources

PubMed1998

H-7 stimulates desmosome formation and inhibits growth in KB oral carcinoma cells.

Protein kinase inhibitor H-7 was reported to stimulate desmosome formation in normal keratinocytes and to inhibit proliferation of neural cell lines. In the present study, the effects of this inhibitor on adhesion and growth of KB human oral carcinoma cells were investigated. H-7 was found to enhance desmosome assembly, as evidenced by an increased punctate labeling for the major desmosomal markers. Immunogold labeling confirmed the formation of desmosomes both at the cell surface and in the cytoplasm. In order to assess cell proliferation and possible correlation with adhesion, confluent cultures were treated and both adherert and detached cell fractions were counted. Under serum-free conditions, H-7 significantly reduced cell detachment. In contrast, EGF stimulated cell detachment, and this effect was abolished when cells were simultaneously treated with both EGF and H-7. Total cell counts were also significantly reduced by H-7, both in the presence and absence of EGF. Using the TUNEL technique, labeled cells were increased after H-7 treatment, thus implicating protein kinase inhibition in cell death. These results indicate that H-7 inhibits growth and stimulates adhesion of KB carcinoma cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1998

Cartilage formation by fetal rat chondrocytes cultured in alginate beads: a proposed model for investigating tissue-biomaterial interactions.

Chondrocytes from 21-day-old rat fetal nasal cartilage were cultured in alginate beads for up to 20 days. It was found that chondrocytes retained their spherical shape and typical chondrocytic appearance. During the culture time, chondrocytes underwent differentiation, as demonstrated by the alkaline phosphatase-specific activity and rate of proteoglycan synthesis. Morphological data confirmed chondrocyte differentiation with the appearance of hypertrophic chondrocytes scattered in the alginate gel and a dense extracellular matrix containing filamentous structures and matrix vesicles. In addition, Northern blot analysis performed on day 8 of culture showed that chondrocytes cultured in alginate beads expressed type II collagen mRNA. The alginate bead method also appeared to be suitable for testing biomaterials, and the ready dissolution of the alginate beads by chelating agents provided a simple means for the rapid recovery of encapsulated chondrocytes. Powdered glass-ceramic particles entrapped in the alginate gel were colonized by chondrocytes, which then proliferated and formed a tissue similar to a true calcified cartilaginous structure. These results indicate that the alginate system represents a relevant model for studies of chondrogenesis and endochondral ossification. Furthermore, the encapsulation method could prove useful for studies of tissue-biomaterial interactions in an in vitro environment which more closely mirrors the cartilage matrix than other culture methods.

Animals↗

PubMed1998

Gene and protein expression during differentiation and matrix mineralization in a chondrocyte cell culture system.

Endochondral bone formation occurs through a series of developmentally regulated cellular stages, from initial formation of cartilage tissue to calcified cartilage, resorption, and replacement by bone tissue. Nasal cartilage cells isolated by enzymatic digestion from rat fetuses were seeded at a final density of 10(5) cell/cm2 and cultured in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal calf serum in the presence of ascorbic acid and beta-glycerophosphate. First, cells lost their phenotype but in this condition they rapidly reexpressed the chondrocyte phenotype and were able to form calcified cartilaginous nodules with the morphological appearance of cartilage mineralization that occurs in vivo during endochondral ossification. In this mineralizing chondrocyte culture system, we investigated, between day 3 and day 15, the pattern expression of types II and X collagen, proteoglycan core protein, characteristic markers of chondrocyte differentiation, as well as alkaline phosphatase and osteocalcin associated with the mineralization process. Analysis of labeled collagen and immunoblotting revealed type I collagen synthesis associated with the loss of chondrocyte phenotype at the beginning of the culture. However, our culture conditions promoted extracellular matrix mineralization and cell differentiation towards the hypertrophic phenotype. This differentiation process was characterized by the induction of type X collagen mRNA, alkaline phosphatase, and diminished expression of type II collagen and core protein of large proteoglycan after an increase in their mRNA levels before the mineralizing process. These results revealed distinct switches of the specific molecular markers and indicated a similar temporal expression to that observed in vivo recapitulating all stages of the differentiation program in vitro.

Aggrecans↗

PubMed1998

Retinoic acid suppresses the osteogenic differentiation capacity of murine osteoblast-like 3/A/1D-1M cell cultures.

Bone is a target tissue for action of retinoids though their precise role remains unclear. This study investigated the effects of retinoic acid (RA) on the marrow stromal 3/A/1D-1M osteoblast-like cells, derived from the in vivo transplantation of 3/A/1D-1 chondroprogenitor cells. Long-term treatment with 1 microM RA for 7 weeks induced a marked decrease in bone-like nodule number and ultrastructural alterations in the striation and the size of the collagen fibres. RA at concentrations varying from 10 nM to 3.16 microM had a dose-dependent inhibition effect on alkaline phosphatase (AP) activity with an IC50 of 0.7 microM. Treatment with 1 microM RA for up to 17 days induced a time-dependent inhibition of AP activity, while the beginning of RA treatment (4 or 52 h of culture) produced a differential magnitude of inhibition. These variations were unrelated to modifications of the expression of RAR receptor at the protein level, as assessed by Western blot analysis. Exposure to 1 microM RA for 6 or 24 h administered at day 14 produced an inhibition of AP activity, which reached a maximum after 48 h, with a recovery time of 8 days in both cases. Long-term treatment with RA at 1 microM completely abolished the level of osteocalcin mRNA on both days 12 and 16, as revealed by Northern blot analysis. However, such RA-treated cells retained the constitutive expression of type II procollagen transcripts. These results suggest that RA inhibits several aspects of osteogenic differentiation capacity, a loss of phenotype, which, in association with the maintenance of type II procollagen cartilage-related characteristic, could be a prerequisite step for cellular plasticity.

Alkaline Phosphatase↗

PubMed1998

Activated adult human alveolar bone cells: a new model of matrix mineralization.

Activated adult human alveolar bone cells were isolated from 2-wk-old osteogenic tissue recuperated from dental implant surgeries following a two-step procedure. Osteogenic tissues were cultured as explant for 2 months. Cells began to migrate in the first 3 d and were confluent in 3-4 wk. However, adjacent to the explants, multicellular cell layers began to form in 10 d, and matrix mineralization was observed by 4 wk in these areas. These formations enlarged and by the end of the culture period, large diffuse matrix mineralization areas were observed. Light and electron microscopic observations confirmed the presence of a collagen matrix undergoing a mineralization process but showing important differences with the mineralized matrix tissue formed with a rat embryo calvaria bone cell system. This new model, using activated human alveolar bone cells, may provide a tool to investigate alveolar bone development and physiology and to set up new therapeutic approaches.

Adult↗

PubMed1998

Desmosomes are regulated by protein kinase C in primary rat epithelial cells.

In the present study, we addressed the possible relevance of protein kinase C (PKC) in the regulation of intracytoplasmic desmosome assembly. Treatment of cultured rat lingual and epidermal keratinocytes with a potent and highly selective PKC inhibitor (GF109203X) induced an increase in granular labelling for major desmosomal proteins, desmoplakins, desmoglein and plakoglobin, both intracellularly and at the cell surface. This was associated with the formation of ultrastructurally recognizable desmosomes deep in the cytoplasm and increase in intercellular desmosome number. In contrast, PKC activation upon short exposure to 12-O-tetradecanoylphorbol 13-acetate (TPA) resulted in altered cell morphology, loss of intercellular contact and accumulation of desmosomal proteins in the juxtanuclear zone. On the other hand, PKC depletion by long term TPA treatment re-established cell-cell contact, where desmosomal markers were exclusively redistributed. Taken together, these results suggest that inhibition of PKC is required for intracytoplasmic as well as intercellular desmosome assembly, whereas its activation may regulate disassembly process.

Animals↗

PubMed1997

Behavior of fetal rat chondrocytes cultured on a bioactive glass-ceramic.

We examined the behavior of fetal rat chondrocytes cultured on a bioactive glass-ceramic containing apatite and wollastonite (A.W.G.C.). Biomaterial surface topography and profiles were evaluated by bidimensional profilometry and revealed a rough surface for the glass-ceramic compared to the plastic coverslips used as controls. Chondrocyte attachment was evaluated by measuring the number of attached cells after one day of culture and by morphological observations. Chondrocytes attached in great numbers to the material surface by means of focal contacts containing vinculin and beta1-integrin. Fluorescent labeling of actin and vimentin revealed a poor spreading of chondrocytes on the bioactive glass-ceramic compared to the plastic coverslips, where the cells appeared to adhere intimately to the surface and exhibited polygonal arrays of stress fibers. During the following days of culture, chondrocytes proliferated, colonized the surface of the material, and, finally, on day 10, formed nodular structures composed of round cells separated by a dense extracellular matrix. Furthermore, these clusters of round cells were positive for type II collagen and chondroitin sulfate, both hard markers of the chondrocyte phenotype. In addition, protein synthesis, alkaline phosphatase activity, and proteoglycan production were found to increase gradually during the culture period with a pattern similar to that observed on control cultures. These results demonstrate that the bioactive glass-ceramic tested in this study appears to be a suitable substrate for in vitro chondrocyte attachment, differentiation, and matrix production.

Actins↗

PubMed1997

Glycoproteins expression in apical pathologic tissues: clinical incidences.

Indirect immunofluorescence was used for the localization of the primary collagens, fibronectin and laminin. Specimens were extracted from untreated teeth with periapical lesions from patients 20 to 30 years of age. An histological examination enabled the differentiation of granulomas and cysts, and 5 microns sections were used for the indirect immunofluorescence procedure. Antibodies against Type I, Type III, and Type V collagen and for fibronectin and laminin were obtained from glycoproteins of human cells. The antibody against Type IV collagen was prepared from Type IV collagen of beef retina. All the glycoproteins investigated were expressed in apical lesions. The intensity of the immunostaining appeared more positive at the external area compared with the center of the lesion. The type IV collagen was specific for the basement membrane of cysts. The immunofluorescence reactions of fibronectin and of laminin were similar in intensity in both granulomas and cysts.

Adult↗

PubMed1997

Rapid nodule evaluation computer-aided image analysis procedure for bone nodule quantification.

Using bone cell cultures, the effects of drugs on cell activities such as proliferation, differentiation, matrix formation, and mineralization can be explored. To quantify these parameters accurately and quickly, a kinetic reproducible computed image analysis procedure of culture dishes is proposed which could be conjointly used with biochemical analysis of the medium. In the present article, different mathematical procedures coupled either with or without histochemical staining are investigated and analyzed. Using serial cross sections and microradiographies of bone nodules, we demonstrated that the gray-level parameter is well correlated with bone mass and/or the mineralization status of the nodules. The procedure selected is a multistep procedure called rapid nodule evaluation (RNE), which uses a binary reconstruction program with different thresholds. To challenge this RNE procedure with the classical Von Kossa staining and quantification procedure, we cultured the cells in the presence of 10 nmol/L dexamethasone and compared the results using the two procedures. The RNE procedure appeared to be accurate and reproducible, and also has the advantage of speed and dynamic analysis over the classical Von Kossa quantification procedure.

Animals↗

PubMed1997

Sequential expression of bone matrix proteins during rat calvaria osteoblast differentiation and bone nodule formation in vitro.

We investigated the expression of osteocalcin (OC), bone sialoprotein (BSP), osteonectin (ON), and alkaline phosphatase (ALP) during cell differentiation and bone nodule formation by fetal rat calvaria cells, using immunofluorescent and immunogold techniques at light and electron microscopic levels. Six hours after plating all proteins were expressed in calvaria cells. However, expression was not detected during the proliferation phase after plating. Cell morphological modifications were observed in osteoblastic cells expressing ALP, OC, and BSP, but not ON. During the matrix formation phase, all proteins were expressed with various intensities and OC was limited to differentiated osteoblastic cells. EM observations demonstrated that BSP was selectively associated with clusters of needle-like crystals, but not with collagen fibers, in mineralization foci and in the mineralized matrix. OC was localized intracellularly and in all the extracellular compartments, and was concentrated at the mineralization front. ON was distributed uniformly throughout the osteoid and mineralized matrix, which was intensely labeled. The results show that the expression of bone matrix proteins during differentiation of calvaria cells and nodule formation in vitro duplicate what is observed during osteogenesis in vivo.

Alkaline Phosphatase↗

PubMed1996

Ecto-alkaline phosphatase considered as levamisole-sensitive phosphohydrolase at physiological pH range during mineralization in cultured fetal calvaria cells.

Alkaline phosphatase (ALP) activity expressed on the external surface of cultured fetal rat calvaria cells and its relationship with mineral deposition were investigated under pH physiological conditions. After replacement of culture medium by assay buffer and addition of p-nitrophenyl phosphate (pNPP), the rate of substrate hydrolysis catalyzed by whole cells remained constant for up to seven successive incubations of 10 min and was optimal over the pH range 7.6-8.2. It was decreased by levamisole by a 90% inhibition at 1 mM which was reversible within 10 min, dexamisole having no effect. Values of apparent Km for pNPP were close to 0.1 mM, and inhibition of pNPP hydrolysis by levamisole was uncompetitive (Ki = 45 microM). Phosphatidylinositol-specific phospholipase C (PI-PLC) produced the release into the medium of a p-nitrophenyl phosphatase (pNPPase) sensitive to levamisole at pH 7.8. The released activity whose rate was constant up to 75 min represented after 15 min 60% of the value of ecto-pNPPase activity. After 75 min of PI-PLC treatment the ecto-pNPPase activity remained unchanged despite the 30% decrease in Nonidet P-40-extractable ALP activity. High levels of 45Ca incorporation into cell layers used as index of mineral deposition were decreased by levamisole in a stereospecific manner after 4 h, an effect which was reversed within 4 h after inhibitor removal, in accordance with ecto-pNPPase activity variations. These results evidenced the levamisole-sensitive activity of a glycosylphosphatidylinositol-anchored pNPPase consistent with ALP acting as an ecto-enzyme whose functioning under physiological conditions was correlated to 45Ca incorporation and permit the prediction of the physiological importance of the enzyme dynamic equilibrium at the cell surface in cultured fetal calvaria cells.

4-Nitrophenylphosphatase↗

PubMed1996

Role of beta-GP-derived Pi in mineralization via ecto-alkaline phosphatase in cultured fetal calvaria cells.

The permissive effect of beta-GP on mineralization in cultured rat fetal calvaria cells was investigated in relationship with phosphohydrolase activity of ecto-ALP at physiological pH range. Beta-GP present in the culture medium for 8 days exerted a stimulatory effect on 45Ca incorporation into matrix cell layers while the ecto-ALP activity level measured on intact cells with a saturating concentration of pNPP was similar for cells grown either in the presence or absence of beta-GP. In both types of cultures, beta-GP addition inhibited pNPP hydrolysis in a competitive and reversible manner and increased Pi concentration in the medium. The dose dependency of the effect of beta-GP on 45Ca incorporation and generation of Pi was similar (k phi = 3 mM). Levamisole, but not dexamisole, inhibited both pNPP and beta-GP hydrolyses, which were likely catalyzed by the same ecto-enzyme. The rate of 45Ca incorporation into matrix cell layers, which was high (0.90 mumol/4h/mg cell protein) in cells grown in the absence of beta-GP, was inhibited by 50% by levamisole. In cells grown in the absence of beta-GP, the 45Ca incorporation rate increased progressively after beta-GP addition, reaching after 12 h the value of cultures grown in the presence of beta-GP, the increase being totally inhibited by levamisole. In both types of cells, addition of exogenous Pi at concentrations corresponding to medium levels of beta-GP-derived Pi rapidly led to high 45Ca incorporation rate which was unaffected by levamisole. beta-GP removal from cultures grown in its presence reduced by 50% the 45Ca incorporation rate which recovered the initial value after exogenous Pi addition independently of levamisole presence. Thus, mineral deposition did not affect the level and catalytic efficiency of ecto-ALP to hydrolyze beta-GP in cultured fetal calvaria cells, yet it influenced the beta-GP-stimulatory effect on mineralization so as to render this process not sensitive to high medium Pi levels.

Alkaline Phosphatase↗

PubMed1996

Cytoplasmic desmosome formation by H-7 and EGF treatment in cultured fetal rat keratinocytes.

Cytoplasmic desmosomes (CD) are classically found in dyskeratotic cells of many epithelial tumors. Their significance and mechanism of formation remain largely speculative. Recently, we have reported the induction of these structures in rat keratinocytes following a brief treatment with acrylamide, and proposed that protein kinase inhibition may be implicated in their formation. In the present study, we show that protein kinase inhibitor H-7 in the presence of EGF is able to induce CD in rat keratinocytes within half an hour. In serum free medium containing 20 ng/ml of EGF, desmosomal structures at different stages of assembly were obtained using H-7 at concentrations ranging between 20 and 80 microM. No such structures were found at lower concentrations. The plaque diameters were significantly small in comparison with plasma membrane plaques. EGF induced plakoglobin positive membrane invaginations and in the presence of H-7, desmosomal plaques assembled on these membranes as either half desmosomes or as symmetric ones. The present results implicate protein kinase inhibition in CD formation and suggest that EGF provides tubular membrane structures in the cytoplasm on which desmosomes may assemble.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1996

Phenotypic characterization of the 3/A/1D-1M osteogenic cell line derived from in vivo transplantation of 3/A/1D-1 chondroprogenitor murine teratocarcinoma cells.

Bone cells involved in the replacement of cartilage by bone in the endochondral ossification process are known to enter via the medullar pathway. A hypothesis for the development of osteoblasts from chondroblasts was investigated by analyzing the phenotypic characteristics of the 3/A/1D-1M cell line derived from endochondral bone ossicle which was formed after in vivo transplantation of 3/A/1D-1 chondroprogenitor mouse teratocarcinoma cells. The 3/A/1D-1M cell cultures exhibited a triphasic evolution: after reaching confluence (day 3), cultures developed well-delimited cell clusters (days 6-8), which ultimately were organized into multilayered nodules (days 12-15). Electron-microscopic examination of such nodules at day 18 showed the presence of needle-shaped crystals associated with collagen fibrils in the extracellular space. The kinetics of collagen expression, investigated by an immunofluorescence staining procedure showed that, while confluent cultures mainly expressed type III collagen (70% of cells) with some type I (30-40% of cells) and V (30-40% of cells), the type I collagen became the major isoform beginning with day 6. From day 6 onwards, NP40-extracted alkaline phosphatase (AP) activity appeared concomitantly to cell cluster formation, and reached 160 nmol/min/mg of protein at the stage of nodule maturation (day 15). The strong inhibition of enzymatic activity by levamisole and L-homoarginine (IC50 = 0.9 microM and 5 mM, respectively) and its rapid heat inactivation at 56 degrees C (IT50 = 90 s), revealed the bone specificity of AP expressed by 3/A/1D-1M cells. In confluent cultures, brief exposure to parathyroid hormone (10 nM), known to be a bone-resorbing agent, showed a 60% increase in the intracellular cAMP level. In addition, while producing mRNA for the bone-specific protein osteocalcin, 3/A/1D-1M cells also produced type II procollagen mRNA, known to be the major cartilage-related characteristic. This in vitro study demonstrates that the 3/A/1D-1M clonal cell line, originating from 3/A/1D-1 chondroprogenitor cells after in vivo passage, was able to develop differentiated osteoblastic properties as well as the residual expression of the major chondrocytic RNA messenger.

Alkaline Phosphatase↗

PubMed1995

Osteoblast adherence and resorption activity of isolated osteoclasts on calcium sulphate hemihydrate.

The adherence of osteoblast-like cells and the resorption activity of isolated osteoclasts on calcium sulphate hemihydrate (CSH) was investigated. After a 24 h incubation period, alkaline phosphatase staining showed that rat osteoblast-like cells ROS 17/2.8 attached on CSH. Neutral red (NR) and tartrate-resistant acid phosphatase (TRAP) staining revealed that osteoclasts attached on CSH. Furthermore, osteoclasts formed lacunae as revealed by scanning electron microscopy. Although the lacunae formed by osteoclasts on CSH were diverse in shape and form, the most common type had approximately a circular outline, with a well-defined margin similar to those formed on dentine. Less commonly, excavations appeared as a discontinuous area of resorbed CSH with the presence of a circular zone around the non-resorbed area. Finally, by using 10(-9) M calcitonin, evidence was obtained that NR-positive cells were osteoclasts (58.3% and 57.66% decrease of NR-positive mouse and rat cells detected on CSH after 24 h incubation). However, no inhibition was obtained with 10(-11) M calcitonin. The overall number of NR-positive osteoclasts adherent on 256 mm2 CSH was 43 +/- 14 and 42 +/- 3 for mice and rat, respectively. The overall number of TRAP-positive mouse osteoclasts was 67 +/- 12. Acetazolamide (10(-5) M), a carbonic anhydrase inhibitor, inhibited the number of adherent NR osteoclasts on CSH by 50.42% and 41.6% for mouse and rat, respectively. These results indicate that osteoblasts attach on CSH and osteoclasts resorb CSH in vitro.

Acetazolamide↗

PubMed1995

Cytochalasin D induces changes in cell shape and promotes in vitro chondrogenesis: a morphological study.

One of the initial events required for the expression of cartilage-specific macromolecules in monolayer cultures is the reversion to the initial round shape of chondrocytes. Thus, considerable research efforts have focused on developing reliable procedures to maintain a round morphology of cultured chondrocytes. Our study focuses on evaluating the response of dedifferentiated fetal rat chondrocytes to cytochalasin D, an actin-disrupting agent, with special emphasis on the morphological events. Immediately after exposure to the drug, cells round up but flatten again after removing the agent. However, immunocytochemical procedures revealed a disorganization of microfilaments and intermediate filaments. Phase-contrast and scanning electron microscopic observations revealed that on day 6 of culture, cells located at the top of the cell layer adopted a spherical morphology. Prominent differences were noted in control cultures where cells had to aggregate prior to overt chondrogenesis. Transmission electron microscopy confirmed the round morphology of the cells situated at the top layer but also revealed the presence of cell contacts between the cells. In addition, cells located at the central part of the cell layer displayed a typical morphology of mature chondrocytes, separated by an extensive extracellular matrix. These morphological changes occurred parallel to the expression of type II collagen and chondroitin sulfate, both hallmarks of the chondrocyte phenotype strong in experimental cultures, relatively weak in control cultures, and only restricted on areas of polygonal cellular aggregates. Furthermore, [35S]-sulfate incorporation into sulfated glycosaminoglycans increased rapidly with the period of culture to a maximum after 7 days and was then two-fold in treated cultures. Taken together, these findings indicated that cytochalasin D stimulates chondrogenesis in response to modification of cytoskeleton architecture and the subsequent rounding up of the cells.

Actin Cytoskeleton↗

PubMed1994

Bioactive glass-ceramic containing crystalline apatite and wollastonite initiates biomineralization in bone cell cultures.

Rat bone cells were cultured in the presence of bioactive glass-ceramic containing crystalline apatite and wollastonite. Scanning electron microscopy observations of the surface of the seeded ceramic disks revealed that cells attached, spread, and proliferated on the material surface. Soaking in cell-free culture medium showed that no change occurred in the surface structure. However, when cultured with bone cells and observed under a transmission electron microscope, an electron-dense layer was noted initially at the surface of the material, before bone formation occurred. In addition, energy-dispersive X-ray microanalysis demonstrated the presence of calcium and phosphorus in this layer. Progressively, during the following days of culture, active osteoblasts synthetized and laid down an osteoid matrix composed of numerous collagen fibrils arranged either parallel or perpendicularly to the first-formed electron-dense layer. Mineralization initiated on the ceramic surface dispersed then along the collagenous fibrils, leading to a mineralized matrix which surrounded the ceramic particles. These results demonstrate the capacity of apatite-wollastonite glass ceramic to initiate biomineralization in osteoblast cultures and to achieve a direct bond between the surface apatite layer of the bioactive glass-ceramic and the mineralized bone matrix.

Animals↗

PubMed1994

Cytoplasmic desmosomes and intermediate filament disturbance following acrylamide treatment in cultured rat keratinocytes.

The present paper describes disturbances in the organization of tonofilaments and desmosomes of rat lingual and epidermal keratinocytes after treatment of the cells with acrylamide in culture. This treatment induced changes in cell shape, reduction of intercellular adhesion and a perinuclear accumulation of cytoplasmic organelles. Using specific antibodies for cytokeratins, the filaments were disorganized particularly in the perinuclear region. In untreated cells, keratin filament labelling was very weak or absent above and below the nucleus thus leaving a black nuclear space in fluorescine microscopy. Following acrylamide treatment, the keratin filament labelling covered the nuclear space which indicated the accumulation of these filaments all around the nucleus. Furthermore, the desmosomal junctions were often associated with thick keratin bundles. Antibodies for desmoplakins revealed a reduction in intercellular labelling and stronger cytoplasmic labelling. Ultrastructurally, well-developed long tonofilaments were found to associate with large desmosomal junctions. Furthermore, small-sized desmosomal structures were identified within the cytoplasm. Morphologically, these were identical to cell surface desmosomes and were almost always associated with well-developed tonofilaments. The effect of acrylamide on the protein kinase A activity might be implicated in the disturbances of the desmosome-intermediate filament complex and in the initiation of contractile forces necessary for perinuclear accumulation of intermediate filaments and for the formation of intact cytoplasmic desmosomes. The acrylamide-induced intermediate filament and desmosomal changes may provide valuable information on the mechanism of intact cytoplasmic desmosome formation in several skin diseases and in squamous cell carcinoma.

Acrylamide↗