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Biomedical subjects

M Yaar

Publications and source records attributed to M Yaar.

At least 55 records · Page 3Linked to original sources
PubMed1994

Aging and photoaging affect gene expression in cultured human keratinocytes.

BACKGROUND: Aging has two components: changes that occur in all individuals with the passage of time alone and changes that occur to varying degrees in different individuals as a result of repeated environmental injury. In the skin, the major environmental influence is sun exposure, and the combined effect of aging and sun damage has been termed photoaging. To determine whether aging and photoaging have characteristic effects on gene expression, we performed Northern blot analysis of several genes in early-passage cultured keratinocytes derived from donors of different ages (newborn, young adult, and old adult) and from paired sun-exposed (photoaged) and sun-protected sites of old-adult donors. OBSERVATIONS: We examined genes involved in cell division, immunomodulation, or differentiation. Aging alone strikingly increased the baseline expression of the differentiation-associated SPR2 and interleukin 1 receptor antagonist (IL-1ra) genes but had relatively little effect on the UV inducibility of any genes studied. In contrast, photoaging markedly increased the inducibility of the c-fos proto-oncogene and decreased the baseline expression of SPR2 and IL-1ra relative to that in cells derived from sun-protected skin of the same donors. CONCLUSIONS: Both aging and photoaging alter the expression of selected genes that are implicated in growth, differentiation, immunomodulation, and UV response in human epidermis. This may explain, in part, the predisposition to photocarcinogenesis in chronically sun-exposed skin of older individuals.

Adult↗
PubMed1993

Gene modulation accompanying differentiation of normal versus malignant keratinocytes.

Using tetradecanoylphorbol 13-acetate (TPA), a known inducer of epithelial cell differentiation, and Northern blot analysis, we investigated in normal versus well-differentiated malignant keratinocytes the modulation of genes implicated in their growth or differentiated function. In normal keratinocytes transient c-fos induction was detected within 30 min after stimulation and was followed by rapid down regulation of the proto-oncogenes c-myc and epidermal growth factor receptor. Within hours after stimulation mRNA levels for three well-characterized differentiated keratinocyte products, involucrin, interleukin-1 beta (IL1-beta), and fibronectin, were induced, as were those for a growth arrest and DNA damage-inducible (GADD 153) gene and a small proline-rich (SPR1) gene, both known to be associated with differentiation but with of as yet unknown functions. Heat shock protein 70 gene was initially down regulated and was induced only after 48 h. The well-differentiated malignant keratinocyte cell line differed in that the c-fos, GADD, SPR1, and IL1-beta genes had several-fold higher induction, but involucrin mRNA was undetectable and fibronectin mRNA was only minimally induced after TPA stimulation. Malignant cells reached terminal differentiation faster than normal keratinocytes as measured by inability to exclude trypan blue dye, and in situ hybridization using a riboprobe for the differentiation-associated SPR1 gene showed that normal keratinocytes constitutively express this transcript while malignant keratinocytes with virtually identical morphology and growth rate do not. These studies greatly expand our understanding of gene activation and down regulation during normal keratinocyte differentiation and imply that a malignant cell line, even when retaining the phenotype of normal cells, differs in its response to outside stimuli, furnishing at best an imperfect model for investigating the molecular mechanisms of cellular differentiation.

CCAAT-Enhancer-Binding Proteins↗
PubMed1993

Effect of donor age on epidermal growth factor processing in man.

Epidermal growth factor (EGF) is a well-characterized mitogen whose effectiveness decreases with age both in vivo and in vitro. Previous studies utilizing the Hayflick aging model of early versus late-passage fibroblasts failed to demonstrate changes that might account for the unresponsiveness. In contrast, we now report striking differences in EGF receptor (EGFR) number, affinity, and rate of EGF/EGFR internalization in early-passage dermal fibroblasts derived from newborn versus young adult versus old adult donors. These data demonstrate critical differences between the two models of cellular aging, provide an explanation for the age-associated loss of EGF responsiveness, and may explain in part the tendency toward impaired wound healing in the elderly.

Adolescent↗
PubMed1993

Treatment of human melanocytes and S91 melanoma cells with the DNA repair enzyme T4 endonuclease V enhances melanogenesis after ultraviolet irradiation.

Tanning is a protective response of ultraviolet (UV)-irradiated skin that decreases damage from subsequent sun exposures by increasing the epidermal content of melanin, a brown-black pigment that absorbs light energy throughout the UV and visible portions of the electromagnetic spectrum. The melanin pigment is made by epidermal melanocytes and transferred to surrounding keratinocytes. The action spectrum, time course, and histologic features of tanning are well studied, but the initiating molecular events are unknown. Previous work has shown that T4 endonuclease V, a prokaryotic DNA repair enzyme that catalyzes the first and rate-limiting step in repair of UV-induced pyrimidine dimers, delivered in carrier liposomes (T4N5), enhances repair of UV-induced DNA damage in cultured human cells and protects against photocarcinogenesis in an animal model. We now report that T4N5 treatment enhances UV-induced melanogenesis, as measured by melanin content, tyrosinase activity, 14C-dopa incorporation, and visual assessment in both S91 murine melanoma cells and human melanocytes. T4N5 treatment also increases cell yields following UV irradiation. These data suggest that tanning can be stimulated through enhanced DNA repair.

Cell Division↗
PubMed1992

Effect of aging and habitual sun exposure on the genetic response of cultured human keratinocytes to solar-simulated irradiation.

Aging and chronic sun exposure are known to be associated with decreased cutaneous immune function, changes in the balance between epidermal proliferation and differentiation, and a greatly enhanced risk of photocarcinogenesis. However, their specific effects on the response of human keratinocytes to ultraviolet (UV) irradiation are unknown. We therefore asked whether aging and photoaging modulate the response at the mRNA level to UV-inducible genes implicated in immunomodulation and/or growth control. Cultured human keratinocytes derived from newborn, young adult, and old adult donors were exposed to a single physiologic dose of solar-simulated UV or sham irradiation and harvested at 1, 4, 8, 24, and 48 h post-irradiation for northern blot analysis. Specific mRNA was detected using cDNA probes encoding the proto-oncogenes c-fos and c-myc and the growth-arrest and DNA damage (GADD153) gene, all recently shown by our laboratory to be modulated by UV in newborn keratinocytes; interleukin (IL)-1a, IL-1b, and the IL-1 receptor antagonist (IL-1ra), two keratinocyte cytokines and their competitive inhibitor, implicated in the immunomodulatory effect of UV; and SPR2, a recently cloned gene known to be induced during normal keratinocyte differentiation and by lethal UV-C irradiation. Our data suggest that aging alone strikingly increases the baseline expression of SPR2 and IL-1ra but has relatively little effect on the response to UV for the other genes examined. In contrast, the combination of aging and habitual sun exposure, so-called photoaging, markedly increases c-fos inducibility and decreases baseline expression of SPR2 and IL-1ra relative to that in cells from sun-protected skin of the same donors. The implied alterations in signal transduction and differentiation state observed in cells derived from habitually sun-exposed sites of old adults may explain in part the predisposition to photocarcinogenesis in photoaged skin.

Adult↗
PubMed1992

Ageing and photoageing of the skin: observations at the cellular and molecular level.

It is now well established that ageing occurs at the level of individual cells in the skin and other organ systems. Changes in cell behaviour, protein production and gene expression in response to standardized stimuli are readily observed in cultured cells derived from young vs old donors and from photoaged vs sun-protected body sites. Whether these changes are best viewed as a cause or a consequence of ageing cannot be determined at present. Nevertheless, available data now provide cellular and molecular correlates for the well-known differences in clinical responsiveness between newborn, adult and photoaged skin. From this basis, it will hopefully be possible to develop a more comprehensive understanding of cutaneous ageing processes.

Adult↗
PubMed1992

Evidence that filopodia outgrowth is a common final pathway for fibroblast growth inhibition in vitro.

To identify events associated with fibroblast growth inhibition, the effect of two known inhibitors, interferon-alpha and all-trans retinoic acid, on the growth and surface morphology of cultured fibroblasts was examined. Interferon-alpha administered at seeding reduced both growth rate and saturation density; all-trans retinoic acid reduced only saturation density. However, both negative growth modulators were associated with an increase in filopodia outgrowth and an increase in intracellular filamentous actin in a time course corresponding to onset of growth inhibition by these agents. In combination with earlier findings, these data suggested that, regardless of etiology, cultured fibroblast growth restriction is mediated in part by an actin-dependent outgrowth of filopodia that augment intercellular contact.

Cell Division↗
PubMed1991

Oncogenes. The growth control genes.

Genes that are involved in the development of cancer have been identified. These genes, called "oncogenes," are related to normal cellular genes called "proto-oncogenes," which code for proteins involved in the regulation of cellular growth and differentiation. Through the study of these genes, we now begin to assess prognosis of several malignant neoplasms. Moreover, specific inhibitors of oncogene activity have potential use in treatment regimens for cancer.

Cell Division↗
PubMed1991

Evidence for nerve growth factor-mediated paracrine effects in human epidermis.

Nerve growth factor (NGF) is critical to the development and maintenance of the peripheral nervous system, but its possible roles in other organ systems are less well characterized. We have recently shown that human epidermal melanocytes, pigment cells derived from the neural crest, express the NGF receptor (p75 NGF-R) in vitro (Peacocke, M., M. Yaar, C. P. Mansur, M. V. Chao, and B. A. Gilchrest. 1988. Proc. Natl. Acad. Sci. USA. 85:5282-5286). Using cultured human skin-derived cells we now demonstrate that the melanocyte p75 NGF-R is functional, in that NGF stimulation modulates melanocyte gene expression; that exposure to an NGF gradient is chemotactic for melanocytes and enhances their dendricity; and that keratinocytes, the dominant epidermal cell type, express NGF messenger RNA and hence are a possible local source of NGF for epidermal melanocytes in the skin. These combined data suggest a paracrine role for NGF in human epidermis.

Base Sequence↗
PubMed1991

Immediate and delayed molecular response of human keratinocytes to solar-simulated irradiation.

The clinical and histologic events in human skin after a single ultraviolet exposure are well documented; and both in vitro and in vivo studies have established immunologic and mutagenic effects of ultraviolet rays on cells. However, little is known about the ultraviolet damage/repair response at the level of gene induction. This study examines the genetic response in cultured human keratinocytes exposed to a single solar-simulated ultraviolet dose and harvested at 1, 4, 8, 24, and 48 hours postirradiation for northern blot analysis using c-DNA probes to well studied stress related genes, heat shock protein 70 and ubiquitin; to a recently cloned gene associated with growth arrest and DNA damage (gadd 153); to c-myc and c-fos, two proto-oncogenes known to be associated with cell growth and differentiation; and to glyceraldehyde phosphate dehydrogenase, known to be involved in cellular intermediary metabolism, gadd 153 and c-fos mRNA levels are increased early (1-4 hours) in keratinocytes, at a time when the cells appear indistinguishable from nonirradiated controls. At the same early time points, the c-myc mRNA level is decreased. Heat shock protein 70 and ubiquitin transcripts are detectable under basal conditions but are not increased by ultraviolet exposure; although at later time points (24-48 hours), when there is morphologic evidence of cell damage and downregulation of the otherwise constitutively expressed gene glyceraldehyde phosphate dehydrogenase, their unchanged steady state levels of mRNA constitute relative over expression. Irradiation of cells with the same solar-simulated ultraviolet dose filtered to remove the shortest and most damaging wavelengths results in a similar pattern of early gene induction, even though cell growth and survival are unaffected. These data suggest that two classes of genes participate in the response to ultraviolet rays: early genes whose mRNA levels are increased (gadd 153, c-fos) or decreased (c-myc) before or independent of any morphologic evidence of cell damage and late genes that are relatively increased (heat shock protein 70, ubiquitin) or decreased (glyceraldehyde phosphate dehydrogenase) in expression when morphologic cell damage is present. The early events may be a direct response to ultraviolet irradiation, whereas the late events may represent a secondary response to stress.

Base Sequence↗
PubMed1990

Dissociation of proto-oncogene induction from growth response in normal human fibroblasts.

Proto-oncogenes are cellular homologues of viral oncogenes that are known to be associated with regulation of growth and differentiation. The c-myc and c-fos proto-oncogenes have been extensively studied by using established cell lines but to a lesser extent by using normal cells. Using physiologic growth modulators, we have shown that mitotic stimulation of normal human dermal fibroblasts is associated with induction of c-myc and c-fos, but that growth inhibition of these cells is not necessarily accompanied by their down-regulation. When treated with both serum and interferon-alpha during quiescence, fibroblasts were delayed in their progress into the S-phase of the cell cycle as compared to cells treated with serum alone and displayed substantial growth inhibition as measured by cell number at the end of 1 week. However, this growth inhibition was not preceded by down-regulation or delay in induction of c-myc and c-fos mRNA. The above studies suggest that in normal fibroblasts growth inhibition is not necessarily dependent on down-regulation of transcription of either c-myc or c-fos and that interferon may act to inhibit cell growth either through a post-transcriptional effect on cellular proto-oncogenes necessary for cell proliferation or through induction of other, as yet unrecognized gene(s) associated with growth arrest.

Cell Cycle↗
PubMed1990

Variable keratin polypeptide profile in human stratum corneum.

Stratum corneum samples obtained from 46 members in three generations of seven families were analyzed for keratin pattern by gel electrophoresis. All these samples of apparently normal upper arm skin expressed the 55 kDa, 56.5 kDa, and 65 kDa keratin proteins; while only 28%, 20% and 48% of the samples expressed the 50 kDa, 58 kDa, and 67 kDa proteins, respectively. The keratin phenotype was identical in all members of two families (9 individuals) and variable in members of five other families (37 individuals), in whom the patterns were consistent with autosomal dominant inheritance. These results demonstrate inter-individual variations in stratum corneum keratin pattern and may reflect either polymorphism of genes coding for the various keratin polypeptides or a post transcriptional modification of mature keratins by proteolytic digestion.

Adolescent↗
PubMed1990

Cellular and molecular mechanisms of cutaneous aging.

There are two major theories of aging: the programmatic theory states that aging is an inherent genetic process, and the stochastic theory states that aging represents random environmental damage. Processes that are associated with cellular damage and aging are the production of free radicals (a process much enhanced after ultraviolet irradiation) and an increasing number of errors during DNA replication. Cellular manifestations of intrinsic aging include decreased life span of cells, decreased responsiveness of cells to growth signals, which may reflect loss of cellular receptors to growth factors, and increased responsiveness to growth inhibitors. All these findings are more pronounced in cells derived from photodamaged skin. Molecular manifestations of intrinsic aging, studied mainly in fibroblasts, are altered membrane composition, adhesion properties, production of extracellular matrix, and activity of the enzyme catalase. Molecular changes as a result of photoaging are less well characterized; published studies mainly target differences in composition of the extracellular dermal matrix.

Adult↗
PubMed1989

Interferon and the epidermis: implications for cellular senescence.

Human keratinocytes are epithelial cells that cover the external surface of the body and function primarily to protect against physical, chemical and biological injury. Since 1975, serial cultivation of disaggregated human keratinocytes in vitro has been possible and these cultures have been utilized to assess in vitro physiological age. As has been demonstrated in human dermal fibroblasts harvested from donors of different ages, human epidermal keratinocytes from older donors manifest a decreased proliferative response to serum and to selected mitogens when compared to younger donors. Recent work suggests an increased sensitivity to negative growth modulators as well. The epidermis constantly renews itself. In normal skin, the majority of the cells in the germinative, basal cell layer are blocked in Go and do not cycle unless stimulated. Interferons (IFN) are a family of glycoproteins well known for their antiviral activity and their ability to inhibit growth and alter the behavior of various normal and transformed cell types, both in vitro and in vivo. As such, we examined intact skin for the presence of negative modulators of growth and demonstrated the presence, by immunofluorescence and Western blotting, of a protein of approximate molecular weight 40 kilodaltons with polyclonal antiserum directed against recombinant IFN-alpha. Extracts from healthy, non-virally infected keratinocyte cultures contained IFN activity as determined by viral plaque inhibition assays. Further, we have demonstrated that IFN inhibits the growth of the human keratinocyte in a non-cytotoxic, reversible manner and that the keratinocytes harvested from older adult donors are significantly more sensitive to the growth inhibitory effects of IFN than are the keratinocytes of young donors.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Induction of nerve growth factor receptors on cultured human melanocytes.

Normal differentiation and malignant transformation of human melanocytes involve a complex series of interactions during which both genetic and environmental factors play roles. At present, the regulation of these processes is poorly understood. We have induced the expression of nerve growth factor (NGF) receptors on cultured human melanocytes with phorbol 12-tetradecanoate 13-acetate and have correlated this event with the appearance of a more differentiated, dendritic morphology. Criteria for NGF receptor expression included protein accumulation and cell-surface immunofluorescent staining with a monoclonal antibody directed against the human receptor and induction of the messenger RNA species as determined by blot-hybridization studies. The presence of the receptor could also be induced by UV irradiation or growth factor deprivation. The NGF receptor is inducible in cultured human melanocytes, and we suggest that NGF may modulate the behavior of this neural crest-derived cell in the skin.

Cell Line↗
PubMed1988

An extract of bovine thymus stimulates human keratinocyte growth in vitro.

An extract prepared from newborn calf thymus stimulated proliferation of human keratinocytes cultured from newborn foreskins and from skin biopsies of 26 adult volunteers aged 19 to 70 years. Growth over the 7-day assay period in the basal medium was age-dependent, with newborn cultures achieving a 10-fold increase in cell number over seeding density, old adult cultures barely maintaining their seeding density and young adult cultures intermediate in proliferative capacity. Maximally stimulatory extract concentration was 5-fold higher for newborn than for adult keratinocytes, with adult cultures experiencing toxicity at doses still growth-promoting for newborn cultures. At optimal extract concentration the maximal average increase in cell yield (66.3% for newborn, 53.6% for young adult, and 18.1% for old) indicated decreased mitogen responsiveness or increased inhibitor sensitivity with increasing donor age. Stimulation of cholera toxin-treated cultures was equally high, ranging from 39.4% to 145.9%, suggesting that the extract acts through a cyclic AMP-independent pathway. Thymic extract did not increase colony forming efficiency. Our findings provide further support for the concept of functional interactions between the skin and the immune system, in addition to the recognized morphologic similarities between thymic cells and keratinocytes. Furthermore, these data confirm earlier findings of an inverse relationship between mitogen responsiveness and donor age for cultured cells.

Adult↗