Biomedical subjects

I C Mackenzie

Publications and source records attributed to I C Mackenzie.

At least 19 recordsLinked to original sources

PubMed2001

Keratinocyte growth factor expression in human gingival fibroblasts and stimulation of in vitro gene expression by retinoic acid.

BACKGROUND: Keratinocyte growth factor (KGF) is a stromally derived growth factor of the fibroblast growth factor (FGF) family with paracrine effects targeted to influence the growth and differentiation of epithelia. Regional and temporal changes in KGF expression play important roles in the development and maintenance of epithelial structures and in epithelial wound healing. Differing patterns of expression of KGF by fibroblasts in the gingival region could therefore be related to the observed regional variation in the differentiation and behavior of gingival epithelia. METHODS: The in vitro and in vivo patterns of expression of KGF mRNA in human gingival and periodontal fibroblasts were examined using reverse transcription polymerase chain reactions (RT-PCR) and in situ hybridization with digoxigenin-labeled riboprobes. The patterns observed for human gingiva were compared with those for human skin and for murine tissues. RESULTS: Gingival and periodontal fibroblasts showed expression of KGF transcripts in vitro, and the degree of expression was markedly influenced by the presence of retinoic acid, an agent known to influence patterns of epithelial differentiation. Sections of human and murine gingiva and skin showed regionally variable expression of transcripts with the cells expressing KGF in the subepithelial, rather than the deeper, connective tissues and periodontium. CONCLUSIONS: The results point to a role of KGF in the maintenance of normal growth and differentiation of gingival epithelia. A lack of KGF expression by periodontal fibroblasts in vivo is expected to hinder apical epithelial migration and thus stabilize the epithelial attachment. The effects of retinoic acid (RA) on KGF expression in vitro provide an indirect mechanism by which RA may regulate the growth and differentiation of gingival epithelia.

Animals↗

PubMed1997

Retroviral transduction of murine epidermal stem cells demonstrates clonal units of epidermal structure.

It has been suggested that the number and position of epidermal stem cells are related to the units of columnar structure in the upper epidermal strata and that the cells of each unit are derived from a single stem cell. Studies of cell lineage in developing tissues have been facilitated by the use of retroviral transduction to provide inherited expression of a histochemically demonstrable foreign gene product. To provide direct evidence about the clonal nature of epidermal units, murine epidermal keratinocytes were transduced with a replication-deficient retroviral vector carrying the beta-galactosidase gene. Subepidermal injection of virus in vivo led to infrequent transduction with only transient presence of beta-gal-staining keratinocytes within the epidermis. Transduction of keratinocytes in vitro and transplantation back to in vivo sites permitted demonstration of the transduced gene in clusters of cells within the reformed epidermis throughout a 12-wk period. The epidermis redeveloped an ordered columnar structure with restriction of transduced cells to individual columnar units. This clonal appearance is compatible with derivation of each epidermal unit from a single stem cell but is not compatible with a random pattern of cell proliferation. Transduced epidermal sheets that were recombined with oral mucosal connective tissue also redeveloped normal columnar structure with restriction of beta-gal staining to individual columnar units. These data suggest that the establishment of an epidermal stem cell pattern related to units of structure is an intrinsic property of the epithelium and is not dependent on regionally-specific connective tissue influences.

Animals↗

PubMed1997

Transforming growth factor-beta response and expression in junctional and oral gingival epithelial cells.

The junctional (JE) and oral gingival (OGE) epithelium show distinct morphological phenotypes and express different cell surface and keratin markers. Transforming growth factor-beta (TGF-beta) has been shown to stimulate extracellular matrix formation and inhibit proteolytic matrix degradation in periodontal wound healing. To elucidate potential roles of TGF-beta in gingival epithelial regeneration and reattachment, the present study examined the effects of TGF-beta on JE and OGE cell growth and determined the patterns of expression of mRNAs for the TGF-beta isotypes beta 1, beta 2 and beta 3 and TGF-beta receptor types I, II and III. Primary cell cultures were initiated from JE and OGE and the cell phenotypes confirmed using monoclonal antibodies to specific keratins. TGF-beta induced a significant growth inhibition in OGE cells derived from 6 different patients with a mean inhibition of 46% and a range of 16-70% (p = 0.031). Although responses varied between patients, in general maximum inhibition occurred at 10 ng/ml TGF-beta. JE cells from 5 patients showed no significant growth inhibition by TGF-beta (p = 0.125). Greater expression of TGF-beta 2 and receptor type I mRNA was found in OGE than JE cells and thus appeared to be associated with differentiating epithelial cells. JE cells expressed more TGF-beta type II receptor specific mRNA than did OGE cells, but TGF-beta 1 mRNA expression was similar in JE and OGE cells. JE or OGE cultures derived from 2 of 3 patients showed expression of mRNA for the TGF-beta type III receptor. TGF-beta 3 mRNA was not detected in any of the JE or OGE samples examined. The greater sensitivity of OGE than JE to the growth inhibiting effects of TGF-beta correlated with higher expression of receptor type I mRNA which, together with the type II receptor, is required for sensitivity to growth inhibition by TGF-beta. The results suggest that, in addition to structural differences, the development of functional differences in the responses of JE and OGE to TGF-beta may be associated with the formation of JE from OGE cells and the reformation of attachment after periodontal surgery.

Antibodies, Monoclonal↗

PubMed1996

Influence of retinoic acid on the expression of cytokeratins, vimentin and ICAM-1 in human gingival epithelia in vitro.

Phenotypic differences exist in vivo between junctional (JE) and oral gingival (OGE) epithelia and an in vitro system has been developed that maintains phenotypic differences. This system, which permits in vitro studies of factors that may influence the epithelial phenotype, was used to investigate the effects of retinoic acid (RA) on epithelial expression of various markers known to distinguish JE from OGE. Primary cultures of JE and OGE were initiated from defined gingival regions and were subcultured and grown for 48 h in 96-well plates or on multiple-well slides. Control cultures were grown in medium supplemented with delipidized serum and all-trans RA was added to experimental groups. Other cultures were grown in a defined RA-free medium. Cultures were examined using monoclonal antibodies against cytokeratins, vimentin, and ICAM-1 and binding displayed by indirect immunocytochemical staining. Staining reactions were assessed by direct microscopic observation and assayed by spectrophotometric quantitation. The results showed that RA had minor effects on the marker expression of JE but markedly enhanced expression of cytokeratins 8, 18, 19, vimentin and ICAM-1 in OGE. These markers, which normally distinguish JE from OGE, were expressed at levels approaching or exceeding those of control JE cultures. These observations indicate that RA responsive mechanisms affect the phenotypes expressed by epithelia in vitro and suggest that such mechanisms may be related to the different phenotypic patterns expressed by gingival epithelia in vivo.

Antibodies, Monoclonal↗

PubMed1996

Expression of keratinocyte growth factor in periapical lesions.

The epithelial proliferation associated with inflammatory periapical lesions and with periapical cyst formation represents an interesting but poorly understood pathological change. Keratinocyte growth factor (KGF) is a recently identified growth factor that is produced by stromal fibroblasts and acts specifically to stimulate epithelial growth and differentiation. To investigate its possible role in the activation of the normally quiescent rests of Malassez, we examined the expression of KGF by in situ hybridization of sections of normal periodontal ligament (PDL) and of 12 periapical granulomas or cysts. Normal PDL and periapical granulomas with scant inflammatory infiltration showed few cells expressing message for KGF. However, KGF-expressing cells were found in the connective tissue stroma close to dense foci of inflammatory cells and to proliferating epithelial elements and cystic epithelial linings. Examination of tissues by the reverse-transcription polymerase chain reaction (RT-PCR) showed KGF expression in 4 specimens of periapical lesions but low or undetectable levels in normal PDL. These observations suggest that the induction of KGF expression in the stromal cells of periapical lesions may play an important role in stimulating the epithelial proliferation associated with cyst formation.

Base Sequence↗

PubMed1995

Expression of blood group-related glycoconjugates in the junctional and other oral epithelia of rodents.

BACKGROUND: The junctional epithelium (JE) attaches the gingiva to the non-vital tooth surface and has other unusual properties which protect the underlying periodontal tissues. The JE differs from other gingival and oral epithelia in its unusual expression of cytokeratins typical of both stratifying and of simple epithelia, a phenotypic pattern possibly related to its specialized functions. METHODS: The patterns of differentiation of rodent gingival and other epithelia were examined using monoclonal antibodies against various glycoconjugates which are expressed on epithelial cell surfaces and provide an alternative marker system for regionally-differing patterns of cell maturation. RESULTS: Markers that are typical of basal cells in other stratifying epithelia were expressed by all cell strata of JE. JE lacked differentiation markers typical of other stratifying oral epithelial but showed suprabasal expression of markers typically expressed by simple epithelia and specialized epithelia, such as taste buds. CONCLUSIONS: The phenotype of rodent JE differs from that of other oral epithelia and the pattern of differentiation assessed by its expression of glycoconjugates parallels that for other phenotypic markers, such as cytokeratins. Differentiation of rodent JE is similar to that of human JE. The functional significance of these patterns of expression is not yet clear but the markers characterizing this unusual epithelium in rodents may be associated with its behavior in periodontal disease and of value to experimental studies of its development.

ABO Blood-Group System↗

PubMed1995

Formation of normal gingival epithelial phenotypes around osseo-integrated oral implants in humans.

The oral, oral sulcular, and junctional epithelia of the natural gingiva each possess distinct patterns of differentiation that are demonstrable both ultrastructurally and by their individual patterns of macromolecular synthesis. The supracrestal tissues reformed around oral implants structurally resemble those of natural gingiva but little is known about phenotype changes occurring in the epithelia. To investigate whether peri-implant epithelia acquire similar patterns of differentiation to those of natural gingiva, biopsies from the supracrestal regions of five oral implants were examined by immunofluorescent methods using a panel of monoclonal antibodies with specificities for individual cytokeratins and ICAM-1, macromolecules which act as markers of the three gingival epithelial phenotypes. The observed staining patterns indicated the formation of oral, oral sulcular, and junctional epithelia which were phenotypically indistinguishable from those of natural gingival epithelia. This degree of reprogramming of epithelial gene expression is a surprising observation and the potential mechanisms leading to the development of those new epithelial phenotypes are discussed in the context of what is known about the development of natural gingiva, in terms of the possible effects of inflammation, and in relation to the known connective tissue influences on epithelial differentiation.

Cell Differentiation↗

PubMed1994

Rates of clearance of the epithelial surfaces of mouse oral mucosa and skin.

Cell desquamation, by removing material adherent to the epithelial surface, appears to play an important role in limiting bacterial colonization of epithelia. Data are available concerning rates of epidermal surface clearance but similar data for oral mucosal epithelia, which exist in an environment more conducive to colonization, have not been reported. The rates of surface clearance of six regions of murine oral mucosa and skin were assessed by two different methods: (a) EM autoradiography to examine the rate of passage of a 3H-histidine label through the stratum corneum, and (b) measurement of rates of cell proliferation and of the size of the superficial cells of the stratum corneum. The autoradiographic (direct) method indicated regionally-varying epidermal clearance rates (6.3-15.9 h) which compared well with published data. This method indicated considerably faster (2.3-3.4 h) clearance rates for the three oral mucosal regions. The rates of surface clearance that were calculated indirectly, from rates of cell proliferation and of cell size, similarly showed faster clearance rates for mucosa (2.0-3.9 h) than for skin (6.0-17.9 h) but differed in the relative rates calculated for the individual regions of mucosa and skin.

Animals↗

PubMed1993

Patterns of cytokeratin expression in the epithelia of inflamed human gingiva and periodontal pockets.

Fourteen specimens of periodontal pockets and the associated marginal gingiva were collected and either frozen for examination using antibodies against various defined cytokeratin specificities or processed for 2-dimensional gel electrophoresis. The epithelium forming the pocket lining typically extended into the connective tissue of the pocket wall in the form of a network of finger-like strips. Immunocytological staining indicated that keratins (K) 5, 6, 14 and 19 were expressed by almost all cells of the pocket lining and K13 and K16 by the suprabasal cells. The coronal region of the pocket lining showed some cells staining for K4. Staining for K8 and K18 was seen in the apical region of the pocket lining and in the finger-like extensions of epithelium into the connective tissue. Compared with normal gingiva, the sulcular and the oral gingival epithelia showed a marked increase in staining for K19. Surprisingly, the pattern of keratin expression of the epithelium of the pocket lining was found to be essentially similar to that of normal junctional epithelium and the anatomical position of the boundaries between each epithelial phenotype were not significantly altered. These patterns of keratin expression were confirmed by the 2D electrophoretic analyses of microdissected regions of epithelium. The potential significance of inflammation to the epithelial changes associated with pocket formation is discussed.

Adult↗

PubMed1993

Keratin expression in taste bud cells of the circumvallate and foliate papillae of adult mice.

The patterns of keratin expression of taste buds in murine oral mucosa were examined using a panel of antibodies with various specificities for cytokeratins. The patterns for taste buds differed markedly from those of the surrounding epithelium but no regional differences in the staining patterns of the taste buds themselves were detected. The taste buds and the ducts of von Ebner's glands were strongly stained by monoclonal antibodies (mAbs) against cytokeratins (K) 8, 18 and 19, those typically expressed by simple epithelia. Merkel cells, which are also present in the oral epithelium and may correspond to the type III cells in taste buds, stained for K8 and K18 but not K19. None of the mAbs against simple epithelial keratins stained the stratifying epithelium of the trench wall or of the oral mucosa. Antibodies with specificity for keratins that are expressed as differentiation products of the mucosal epithelium did not stain taste buds. The staining pattern of the epithelium of the trench wall indicated that it expressed some keratins typical of stratifying epithelia but lacked the full pattern of differentiation of the adjacent mucosal epithelia. Staining within the taste buds was not homogeneous but no clear differences of keratin staining could be directly related to the subtypes of cells constituting them. The markedly differing patterns of keratin expression between taste buds and the adjacent epithelium raises questions about the type of inductive signals that produce and maintain these patterns.

Animals↗

PubMed1992

Patterns of phenotypic expression of human junctional, gingival and reduced enamel epithelia in vivo and in vitro.

Epithelia differ regionally in their patterns of phenotypic expression. The junctional epithelium (JE) that attaches the oral mucosa to the teeth is a unique tissue that shows a pattern of differentiation unlike other oral epithelia and forms basal lamina against the non-vital tooth surface. The mechanisms that establish this unusual phenotype and the developmental origin of this epithelium are both uncertain. The formation of JE by downgrowth of the oral gingival epithelium (OGE) during tooth eruption has been suggested but morphological studies indicate that it may be derived from the reduced enamel epithelium (REE) that covers the crown of the unerupted tooth. These epithelia of potential origin differ in their developmental histories: intrinsic differences between them could thus significantly influence the phenotype of an epithelium formed from them. The patterns of phenotypic expression of specimens of dissected JE, OGE and REE, and of cell cultures of these epithelia grown under standardized conditions, were examined (1) by immunocytochemistry using monoclonal antibodies with specificity for individual cytokeratins, vimentin and ICAM-1, and (2) by two-dimensional SDS-PAGE and immunoblotting. The results indicated that, in vivo, OGE expressed keratin markers typical of differentiating mucosal epithelium; JE and REE, in contrast, lacked expression of most such markers but expressed keratins typical of simple epithelia together with some undefined keratin peptides. All epithelia showed changes in vitro but OGE remained different from JE and REE. OGE lost expression of the differentiation markers K1, K10 and K13; it acquired some expression of K19, but less than JE and REE. Cultures of JE and REE retained some expression of ICAM-1 and K8 and K18, and consistently acquired high levels of vimentin expression. These findings indicate that differences persist in standardized culture conditions and that these are apparently of an intrinsic nature. They support a concept of the origins of JE from REE and suggest that the unusual in vivo phenotype of JE results partly from intrinsic differences acquired during its development.

Cell Differentiation↗

PubMed1991

The nature of the epithelium in acquired cholesteatoma. Part 2. Cell culture.

The exact nature and role of the epithelial layer in cholesteatoma remains undetermined. The aim of this study was to investigate cholesteatoma epithelium and normal aural epithelia in common cell culture conditions. Samples of cholesteatoma, external meatal epidermis and middle ear mucosa were obtained, successfully grown in cell culture, and subcultured. No significant morphological differences were found between cholesteatoma and aural epidermis. The only differences noted were delayed onset of colony formation, and the need to subculture prior to the cultures becoming confluent in the cholesteatoma cultures. Further research is required to account for these differences in growth patterns.

Cell Count↗

PubMed1991

The nature of the epithelium in acquired cholesteatoma.

Monoclonal antibodies with defined specifications for individual cytokeratins were used to stain the epithelia of the external auditory meatus, the middle ear and cholesteatoma. The observed staining indicated that the epithelium of the external auditory meatus has a pattern of keratin expression typical of epidermis in general and the epithelium of the middle ear resembles simple columnar epithelia. The pattern of staining of cholesteatoma closely resembled that of the skin of the external auditory meatus.

Antibodies, Monoclonal↗

PubMed1991

Epithelial lining of sinus tracts associated with periapical disease: an immunocytochemical study using monoclonal antibodies to keratins.

We examined 30 specimens of mucosal sinus tracts arising in association with periapical inflammation. Immunocytochemistry and a panel of monoclonal antibodies to keratins were used to demonstrate the epithelial lining and its pattern of keratin expression. Half of the specimens were found to have an epithelial lining that was continuous with the mucosal epithelium and showed various degrees of extension into the sinus. The presence of an epithelial lining appeared to correlate with the duration of the sinus tracts. The immunostaining patterns of the epithelial linings were similar to those of the mucosal epithelium near the sinus opening although some differences were found. Proliferating strands of epithelium, presumably derived from the rest of Malassez, were observed deep in the tissues but were not continuous with the epithelial lining. The results suggest that the epithelial lining of sinus tracts is derived not from epithelial rests in the periapical region but from the mucosal epithelium adjacent to the opening of the sinus tract and that the observed changes in the pattern of keratinization are due to the influence of inflammation or of the connective tissue substrate.

Antibodies, Monoclonal↗

PubMed1991

Patterns of cytokeratin expression in human gingival epithelia.

Specimens of human gingiva were collected from teenage and adult subjects and frozen sections were stained with an extensive panel of monoclonal antibodies with defined specificities for individual cytokeratins. The results indicated different and distinctive patterns of keratin expression by the oral gingival, oral sulcular and the junctional epithelia. It was observed that epithelium with staining characteristics of sulcular epithelium extended over the gingival crest onto the oral surface of the gingiva. Junctional epithelium showed the unusual pattern of co-expression of keratins typical of the stratifying and of the simple epithelial phenotypes. The patterns of gingival keratin expression are compared with those of other mucosal epithelia. The findings are discussed in relation to mechanisms that may determine or influence the junctional epithelial phenotype.

Adolescent↗

PubMed1990

Antibody markers of basal cells in complex epithelia.

In the course of immunohistochemical studies it has become apparent that there is a distinct phenotype of keratin expression that is shared by basal epithelial cells in a variety of different tissues. A basal cell can be defined as a cell in contact with a basal lamina but with no free luminal surface; this distinguishes it from a simple epithelial cell, which has a free luminal surface as well as basal lamina contact, and from stratifying suprabasal keratinocytes, which have neither basal lamina contact nor free luminal surface. All basal cells, whether they are in glandular ductal or secretory epithelia, or in stratified squamous epithelia, express the keratin pair K5 and K14. In this paper we describe monoclonal and polyclonal antibodies that are monospecific for both keratins 14 and 5 or are specific for denaturation-sensitive epitopes unique to basal cells, including five new monoclonal antibodies: LL001 and LL002 (to keratin 14), 2.1.D7 (to keratins 5, 6 and 8), and LH6 and LH8 (conformation-specific basal cell markers). These antibodies have been used to monitor the distribution of the basal cell phenotype and to demonstrate the expression of keratins 5 and 14 in this cell type, in both stratified epithelia and mixed epithelial glands. The consistent association of this keratin pair with basal cells suggests a possible specific function for these keratin in reinforcing epithelia under physical stress, whilst expression of these keratins may conflict with the differentiated functions of most simple epithelial cells.

Amino Acid Sequence↗

PubMed1989

The influence of subepithelial connective tissues on epithelial proliferation in the adult mouse.

Subepithelial connective tissue is capable of modulating the pattern of histodifferentiation of stratified epithelia from adult animals, but it is not known whether the supporting connective tissue also influences epithelial proliferative activity. Epithelial and connective tissues of murine skin and oral mucosa, differing in their morphology and proliferative activity, were separated and heterotypically recombined prior to grafting to histocompatible hosts. After 3 or 8 weeks in situ, mitotic activity was determined following the administration of vinblastine sulfate. Although the mitotic activity in each of the epithelia could be modulated by some connective tissues, there was no distinct pattern of behavior. In combination with connective tissues from tongue or palate, the ear epidermis acquired a significantly increased mitotic activity. In contrast, when oral epithelia with high mitotic activity were recombined with dermal connective tissue, there was usually a significant reduction in proliferative activity. As there was no apparent association between mitotic activity and the induced changes in either organization or histodifferentiation, it is suggested that subepithelial connective tissue is capable of directly influencing the mitotic activity in the overlying epithelium.

Animals↗

PubMed1989

Isolation of subpopulations of murine epidermal cells using monoclonal antibodies against differentiation-related cell surface molecules.

Monoclonal antibodies (mAbs) against epithelial cells were prepared by immunization of rats with lyophilized murine epithelia. Screening against tissue sections and epithelial cell suspensions permitted identification of mAbs against surface molecules that are expressed early in cell differentiation. Staining with these mAbs followed by fluorescence-activated cell sorting enabled isolation of subpopulations of basal epithelial cells. Staining these subpopulations with antibodies against known differentiation markers (cytokeratins and bullous pemphigoid antigen) and measurements of cell size indicated that they represented fractions of the basal cell population in sequential stages of early differentiation. Labeling mice with bromodeoxyuridine at various times prior to cell isolation showed that the least-differentiated basal cells cycle more slowly than those at later stages, data which support the concept of a differentiation-related, hierarchical pattern of organization of the proliferative compartment.

Animals↗