PubMed Health⌕ Search

Biomedical subjects

L T Smith

Publications and source records attributed to L T Smith.

At least 19 recordsLinked to original sources
PubMed2006

DNA copy number gains in head and neck squamous cell carcinoma.

Gene amplification, a common mechanism for oncogene activation in cancer, has been used as a tag for the identification of novel oncogenes. DNA amplification is frequently observed in head and neck squamous cell carcinoma (HNSCC) and potential oncogenes have already been reported. We applied restriction landmark genome scanning (RLGS) to study gene amplifications and low-level copy number changes in HNSCC in order to locate previously uncharacterized regions with copy number gains in primary tumor samples. A total of 63 enhanced RLGS fragments, indicative of DNA copy number changes, including gains of single alleles, were scored. Enhanced sequences were identified from 33 different chromosomal regions including those previously reported (e.g. 3q26.3 and 11q13.3) as well as novel regions (e.g. 3q29, 8q13.1, 8q22.3, 9q32, 10q24.32, 14q32.32, 17q25.1 and 20q13.33). Furthermore, our data suggest that amplicons 11q13.3 and 3q26.3-q29 may be divided into possibly two and three independent amplicons, respectively, an observation supported by published microarray expression data.

Carcinoma, Squamous Cell↗
PubMed2003

Differential targets of CpG island hypermethylation in primary and metastatic head and neck squamous cell carcinoma (HNSCC).

Head and neck squamous cell carcinomas (HNSCC) often metastasise to the cervical lymph nodes. It is known for HNSCC as well as other cancers that progression from normal tissue to primary tumour and finally to metastatic tumour is characterised by an accumulation of genetic mutations. DNA methylation, an epigenetic modification, can result in loss of gene function in cancer, similar to genetic mutations such as deletions and point mutations. We have investigated the DNA methylation phenotypes of both primary HNSCC and metastatic tumours from 13 patients using restriction landmark genomic scanning (RLGS). With this technique, we were able to assess the methylation status of an average of nearly 1300 CpG islands for each tumour. We observed that the number of CpG islands hypermethylated in metastatic tumours is significantly greater than what is found in the primary tumours overall, but not in every patient. Interestingly, the data also clearly show that many loci methylated in a patient's primary tumour are no longer methylated in the metastatic tumour of the same patient. Thus, even though metastatic HNSCC methylate a greater proportion of CpG islands than do the primary tumours, they do so at different subsets of loci. These data show an unanticipated variability in the methylation state of loci in primary and metastatic HNSCCs within the same patient. We discuss two possible explanations for how different epigenetic events might arise between the primary tumour and the metastatic tumour of a person.

Adult↗
PubMed2001

Osmosensing and osmoregulatory compatible solute accumulation by bacteria.

Bacteria inhabit natural and artificial environments with diverse and fluctuating osmolalities, salinities and temperatures. Many maintain cytoplasmic hydration, growth and survival most effectively by accumulating kosmotropic organic solutes (compatible solutes) when medium osmolality is high or temperature is low (above freezing). They release these solutes into their environment when the medium osmolality drops. Solutes accumulate either by synthesis or by transport from the extracellular medium. Responses to growth in high osmolality medium, including biosynthetic accumulation of trehalose, also protect Salmonella typhimurium from heat shock. Osmotically regulated transporters and mechanosensitive channels modulate cytoplasmic solute levels in Bacillus subtilis, Corynebacterium glutamicum, Escherichia coli, Lactobacillus plantarum, Lactococcus lactis, Listeria monocytogenes and Salmonella typhimurium. Each organism harbours multiple osmoregulatory transporters with overlapping substrate specificities. Membrane proteins that can act as both osmosensors and osmoregulatory transporters have been identified (secondary transporters ProP of E. coli and BetP of C. glutamicum as well as ABC transporter OpuA of L. lactis). The molecular bases for the modulation of gene expression and transport activity by temperature and medium osmolality are under intensive investigation with emphasis on the role of the membrane as an antenna for osmo- and/or thermosensors.

Bacteria↗
PubMed2001

Expression of a truncated keratin 5 may contribute to severe palmar--plantar hyperkeratosis in epidermolysis bullosa simplex patients.

Epidermolysis bullosa simplex are dominant disorders of skin fragility characterized by intraepidermal blistering upon mild mechanical trauma. Skin fragility is caused by expression of either an abnormal keratin 5 or an abnormal keratin 14 protein, which compromises the structure and function of the keratin cytoskeleton of basal cells. We report an epidermolysis bullosa simplex patient with a novel single base substitution (A-->T1414) that changes the lysine residue at amino acid 472 to a non-sense codon (K472X). This change predicts the synthesis of a truncated keratin 5, missing 119 amino acids, including the entire tail domain and the highly conserved KLLEGE motif at the carboxy terminus of the 2B domain of the central rod. Expression of an altered keratin 5, of predicted mass and pI for the product of the K472X allele, was documented by one- and two-dimensional western blots of protein extracts from patient skin. Ultrastructural analysis of the patient's nonhyperkeratotic skin was remarkable for basal keratinocytes with dense and irregular keratin filaments proximal to the basement membrane. Keratinocytes, transfected with a cDNA carrying the A-->T1414 non-sense mutation, overexpressed a truncated keratin 5, and showed a disorganized and collapsed keratin filament cytoskeleton. This is the second epidermolysis bullosa simplex patient reported with a premature termination mutation in the KLLEGE motif. The remarkable occurrence of severe palmar--plantar hyperkeratosis in both patients suggests that the keratin 5 tail domain may have unrecognized, but important, normal functions in palmar-plantar tissues.

Amino Acid Sequence↗
PubMed2000

Partial COL1A2 gene duplication produces features of osteogenesis imperfecta and Ehlers-Danlos syndrome type VII.

Type I collagen is the most abundant structural protein in the mammalian body. It exists as a heterotrimer of two subunits in the form [alpha1(I)]2alpha2(I). Pathogenic mutations in COL1A1 and COL1A2, the genes that encode the two subunits, cause a range of phenotypes including mild to lethal forms of osteogenesis imperfecta and a restricted set of Ehlers-Danlos syndrome phenotypes. Lethal mutations usually result from missense mutations that disrupt the normal triple helical structure of the molecule. Multi-exon duplication or deletion in type I collagen genes has rarely been observed and has generally resulted in a lethal or severe phenotype. We report a partial duplication in the COLIA2 gene that causes a relatively mild phenotype, despite the addition of 477 amino acids to the triple helical domain of the proalpha2(I) chain. The abnormal molecule is synthesized and secreted by cultured dermal fibroblasts in a normal fashion. Electron microscopy of dermal tissue reveals small but otherwise near normal collagen fibrils. The gene duplication occurred by mitotic sister chromatid exchange in the mother who is mosaic for the duplication allele. Examination of the abnormal sequence suggests a means by which the duplicated molecule could be processed and properly incorporated into mature collagen fibrils.

Base Sequence↗
PubMed2000

Expression of transglutaminase activity in developing human epidermis.

Epidermal transglutaminase (TGase) is known to catalyse cross-linking of several precursor proteins in the formation of cornified cell envelope at the terminal differentiation of keratinocytes. Expression of TGase activity was studied using an in situ TGase activity assay in human fetal skin samples of 49-163 days estimated gestational age (EGA). In the early two-layered epidermis (49-56 days EGA), in situ TGase activity was not observed in the periderm cells or the basal cells. In the late two-layered epidermis (57-65 days EGA), in situ TGase activity became weakly positive in the periderm cells, but not in the basal cells. In the three-layered (66-95 days EGA) and in four- or more layered (96-135 days EGA) stratified epidermis, in situ TGase activity was still restricted only to the periderm cells. After keratinization occurred in the interfollicular epidermis (163 days EGA), in situ TGase activity was expressed in the granular and cornified layers. This unique localization of TGase activity further support the hypothesis that periderm cells form cornified cell envelope during their regression process in human fetal skin development.

Epidermis↗
PubMed2000

Changing patterns of localization of putative stem cells in developing human hair follicles.

In rodents, the hair follicle stem cells lie in a well-defined bulge in the outer root sheath; however, the bulge as a stem cell site of human hair follicle epithelium is still controversial. Epidermal stem cells are thought to express high levels of beta1 integrin and low levels of E-cadherin and beta- and gamma-catenin. In order to clarify the ontogenic distribution of possible stem cells during hair follicle development, the expression patterns of beta1 integrin subunits, E-cadherin, and beta- and gamma-catenins in the skin samples from human fetuses of a series of estimated gestational ages (EGA) were examined. beta1 integrin-rich, E-cadherin-, and beta- and gamma-catenin-poor cells, possible stem cells, were localized to the entire hair germ (65-84 d EGA) and later to the outermost cells of hair peg (85-104 d EGA). In the bulbous hair peg (105-135 d EGA) and in the differentiated lanugo hair follicle (>135 d EGA), they were settled in the bulge and the outermost layer of the outer root sheath. This sequential localization was similar to that of cells rich in epidermal growth factor receptor expression and positive with keratin 19, a putative marker of epidermal stem cells. In addition, these beta1 integrin-rich, E-cadherin-, and beta- and gamma-catenin-poor cells showed similar, undifferentiated morphologic features by electron microscopy. This information of ontogenic localization of possible hair follicle stem cells contributes to the further understanding of mechanisms of human hair follicle morphogenesis and supports the idea that the human fetal hair follicle bulge is a site of stem cells for follicular epithelium.

Cadherins↗
PubMed2000

Mitochondrial ultrastructure in embryos after implantation.

Information on the morphology of mitochondria during embryogenesis is scattered in the literature, but there appears to be a consistent pattern. During early organogenesis, the embryo is in a state of relative hypoxia associated with a major decrease in terminal electron transport system activity and a marked increase in anaerobic glycolysis. Ultrastructural studies of a 14-somite monkey embryo and day 10 and 12 rat embryos, together with a review of the literature, led us to determine that this hypoxic stage is characterized by vesiculation of the mitochondrial inner membranes, or cristae. Starting in the late morula stage and continuing during early postimplantation embryogenesis, the cristae increase but appear tubular or vesicular. After the end of neurulation, and with the onset of vascular perfusion of embryonic tissues, the cristae gradually become lamellated; by the limb bud stage they appear more mature. We suggest that new cristae derive from blebs of the inner mitochondrial membrane and that with maturation these blebs collapse, giving them a lamelliform appearance. The delamellated state of the cristae might inactivate oxidative phosphorylation to protect the embryo from toxic respiratory end-products that could accumulate in an embryo before there is vascular perfusion. Consistent with this hypothesis, mitochondrial diameters in the developing heart of monkey and rat embryos were approximately twice those found in skin and neural tube.

Animals↗
PubMed1999

Elevated retinoic acid receptor beta(4) protein in human breast tumor cells with nuclear and cytoplasmic localization.

The transcription factor retinoic acid receptor beta(2) (RARbeta(2)) is a potent inhibitor of breast cancer cells in vitro, and studies suggest that RARbeta expression is lost in primary breast cancer. Although RARbeta(2) is selectively down-regulated at the mRNA level in breast tumor cells, we show that expression of an RARbeta protein is elevated in five of five breast tumor cell lines relative to normal human mammary epithelial cells. Subsequent analysis identified this protein as the translation product of the human RARbeta(4) transcript. Unlike the previously characterized mouse RARbeta(4) isoform, the human RARbeta(4) retains only half of a DNA-binding domain and lacks a ligand-independent transactivation domain at its N terminus. The RARbeta(4) protein localizes to the cytoplasm and to subnuclear compartments that resemble nuclear bodies. The structure and preliminary characterizations of human RARbeta(4), coupled with the observation that its expression is greatly elevated in breast tumor cell lines, support the hypothesis that RARbeta(4) functions as a dominant-negative repressor of RAR-mediated growth suppression.

Base Sequence↗
PubMed1999

Outcome after surgical repair of junctional epidermolysis bullosa-pyloric atresia syndrome: a report of 3 cases and review of the literature.

BACKGROUND: Junctional epidermolysis bullosa-pyloric atresia syndrome is recognized as a distinct autosomal recessive entity. Affected infants present with skin fragility and inability to feed due to intestinal obstruction. Despite successful surgical repair of the anatomical defect, the outcome is poor owing to poor feeding, malabsorption, failure to thrive, and sepsis. OBSERVATIONS: In 70 cases of intestinal obstruction and epidermolysis bullosa reported in the medical literature and the 3 reported here, surgical intervention was attempted 51 times. In all except 16 infants, death occurred before age 11 months (mean age, 70 days). Renal involvement and continued failure to thrive accompanied the skin disease in survivors, who ranged in age from 30 days to 16 years (mean age, 4.0 years). CONCLUSIONS: The poor prognosis of this condition must be considered when decisions are made regarding surgical correction. Attempting surgical correction may be warranted in individual circumstances, but withholding surgical intervention and providing palliative support is an acceptable alternative.

Epidermolysis Bullosa, Junctional↗
PubMed1999

Trichodysplasia spinulosa--a newly described folliculocentric viral infection in an immunocompromised host.

This is a case report of an immunocompromised individual who presented with progressive alopecia, friable follicular spinous processes, and erythematous, indurated papules. Examination of skin biopsies using light microscopy and immunohistochemistry revealed pathologic changes of the follicular inner root sheath epithelium with dystrophic trichohyaline granules. Electron microscopy of thin sections of tissue revealed intracellular viral particles with a size and appearance consistent with those in the Papovaviridae family. Electron microscopy of negatively stained extract from a homogenized lesion also demonstrated icosahedral viruses with papovavirus morphology. We believe this is a previously unreported folliculocentric viral infection in an immunosuppressed human host and have termed this entity "trichodysplasia spinulosa".

Adult↗
PubMed1999

Ontogeny and regional variability of keratin 2e (K2e) in developing human fetal skin: a unique spatial and temporal pattern of keratin expression in development.

Keratin 2e (K2e) is expressed in the upper spinous and granular cells of adult epidermis. A highly specific polyclonal antibody was made against a C-terminal peptide of K2e and used to observe K2e expression at different developmental stages. At 12.5 weeks estimated gestational age (EGA) K2e was detected in trunk skin in scattered cells in the intermediate layer. At 13.5 weeks EGA, greater numbers of intermediate cells were stained with variable intensity, and staining in this pattern increased with age. Epidermal sheets from 14 weeks EGA showed that K2e + cells were excluded from developing hair follicles. At 135 days EGA, the following regional patterns were observed: in cheek, trunk, dorsal and ventral knee, elbow and dorsal hand there was moderate to intense staining of upper intermediate keratinocytes excluding cells of the hair canals and sweat ducts. The periumbilical region distinctly lacked K2e staining, while more distal areas showed increasing numbers of K2e + cells. The earliest expression of K2e was at 10 weeks EGA in the presumptive nail bed of developing digits. By 13.5 weeks EGA this pattern had shifted to the proximal nail fold, and K2e was absent in the nail bed. K2e was excluded from developing sweat glands and ducts and from developing hair follicles at the hair germ and early peg stages. By 15 weeks EGA in the fetal hair follicle small groups of cells were K2e + and by 19 weeks K2e + cells were seen at the level of the matrix. Some overlap in staining was detected for K2e with K10, and in palmar skin with K9; however, mostly the filamentous staining patterns for these keratins were distinctive. This study shows that the complex patterns of temporal and regional expression of K2e differ from known patterns for other epidermal keratins and suggest different regulation and function for this epidermal keratin.

Embryonic and Fetal Development↗
PubMed1999

Epimorphin expression during human foetal hair follicle development.

Epimorphin is a mesenchymal protein expressed in several organs and known to have an essential role in epithelial tissue organization, including hair follicle morphogenesis, in mice. Although about 90% homology has been reported between human and mouse epimorphin exon sequences, there is no information about expression and function of epimorphin in hair follicle development in humans. In order to elucidate the expression pattern of epimorphin in human hair follicle morphogenesis and to compare it with the distribution of tenascin and neural cell adhesion molecule (NCAM), skin samples from human foetuses of a series of estimated gestational ages (EGAs) (46-168 days EGA) were studied using monoclonal anti-epimorphin antibody MC-1, anti-tenascin antibody and anti-human NCAM antibody. Epimorphin was detected in the mesenchymal cell condensation at the pregerm stage (< 75 days EGA), and there was strong expression of epimorphin in the perifollicular mesenchymal cells around the hair germ (75-84 days EGA). At the hair peg stage (85-104 days EGA), epimorphin was around the hair peg with the strongest staining in the neck portion. This sequence of staining patterns was similar to that of tenascin. In the bulbous hair peg (105-134 days EGA), the perifollicular dermal mesenchymal cells were evenly positive for epimorphin. Mesenchymal cells underneath the follicle bulb prior to formation of the dermal papilla were also positive for epimorphin. In the lanugo hair follicle (> 134 days EGA), dermal papilla cells expressed epimorphin as well as tenascin and NCAM. These results indicate that epimorphin expression is closely linked to developing hair follicles in human foetuses. This suggests that epimorphin may have an important part in induction of morphogenesis during human foetal hair follicle development.

Female↗
PubMed1999

The calcium-activated neutral protease calpain I is present in normal foetal skin and is decreased in neonatal harlequin ichthyosis.

Calcium concentration is a critical factor for epidermal differentiation and is implicated in the expression and post-translational modification of numerous proteins in suprabasal cells of the epidermis. Calpains (calcium-activated neutral proteases) are believed to participate in signal transduction via highly regulated cytoplasmic protease activity. Here we investigate the expression of calpain I in normal human skin development and in neonatal harlequin ichthyosis (HI), a disorder of altered epidermal differentiation, especially the transition from the granular to the fully differentiated cornified layer. Calpain I was detected in developing foetal epidermis at 54 days estimated gestational age in the basal layer and the periderm of the developing foetal epidermis. By 125 days, calpain I was also detected in the granular layer. This pattern was maintained in newborn skin, but expression was significantly weaker in HI biopsies (n = 7). Reduced expression of calpain was specific to HI and was not observed in other skin diseases. Calpain was also normally expressed in the outer root sheath of hair follicles, in sebaceous glands and in sweat ducts and glands. Immunoblots of epidermal and keratinocyte extracts showed that the 78-kDa and 76-kDa active forms were generated via limited proteolysis of the 80-kDa inactive subunit; however, all forms were diminished in HI, consistent with findings in tissue sections. Our results show that calpain is present throughout the epidermis and is expressed from the early stages of development. These findings implicate calcium-mediated signalling events in the alteration of differentiation that occurs in HI.

Biomarkers↗
PubMed1999

Periderm cells form cornified cell envelope in their regression process during human epidermal development.

Terminally differentiated stratified squamous epithelium forms a lining of the plasma membrane called the cornified cell envelope, a thick layer of several covalently cross-linked precursor proteins including involucrin, small proline-rich proteins, and loricrin. Their cross-linking isodipeptide bonds are formed by epidermal transglutaminases 1-3. Material from lamellar granules is attached on the extracellular surface of corneocytes during the keratinization process. The formation of cornified cell envelope and sequential expression of major cornified cell envelope precursor proteins, transglutaminases, and 25 kDa lamellar granule-associated protein were studied in human embryonic and fetal skin. Ultrastructurally, membrane thickening has already started in periderm cells of the two-layered epidermis and an electron-dense, thickened cell envelope similar to cornified cell envelope in adult epidermis is observed in periderm cells at the three-layered and later stages of skin development. In the two-layered epidermis (49-65 d estimated gestational age), immunoreactivities of involucrin, small proline-rich proteins, all the transglutaminases, and lamellar granule-associated protein were present only in the periderm. In the three-layered epidermis and thereafter (66-160 d estimated gestational age), loricrin became positive in the periderm cells, transglutaminases extended to the entire epidermis, and lamellar granule-associated protein was detected in intermediate cells as well as periderm cells. Immunoelectron microscopy demonstrated that both major cornified cell envelope precursor proteins, involucrin and loricrin, were restricted to the cornified cell envelope in periderm cells at this stage of development. After 160 d estimated gestational age, the periderm had disappeared and cornified cell envelope proteins and lamellar granule-associated proteins were expressed in the spinous, granular, and cornified cells and transglutaminases were detected in the entire epidermis. These findings indicate that cornified cell envelope precursor proteins, transglutaminases, and lamellar granule-associated proteins are expressed in coordination in periderm cells during human epidermal development and suggest that periderm cells form cornified cell envelope in the process of regression.

Adult↗
PubMed1999

Cyclic ichthyosis with epidermolytic hyperkeratosis: A phenotype conferred by mutations in the 2B domain of keratin K1.

Bullous congenital ichthyosiform erythroderma (BCIE) is characterized by blistering and erythroderma in infancy and by erythroderma and ichthyosis thereafter. Epidermolytic hyperkeratosis is a hallmark feature of light and electron microscopy. Here we report on four individuals from two families with a unique clinical disorder with histological findings of epidermolytic hyperkeratosis. Manifesting erythema and superficial erosions at birth, which improved during the first few months of life, affected individuals later developed palmoplantar hyperkeratosis with patchy erythema and scale elsewhere on the body. Three affected individuals exhibit dramatic episodic flares of annular, polycyclic erythematous plaques with scale, which coalesce to involve most of the body surface. The flares last weeks to months. In the interim periods the skin may be normal, except for palmoplantar hyperkeratosis. Abnormal keratin-filament aggregates were observed in suprabasal keratinocytes from both probands, suggesting that the causative mutation might reside in keratin K1 or keratin K10. In one proband, sequencing of K1 revealed a heterozygous mutation, 1436T-->C, predicting a change of isoleucine to threonine in the highly conserved helix-termination motif. In the second family, a heterozygous mutation, 1435A-->T, was found in K1, predicting an isoleucine-to-phenylalanine substitution in the same codon. Both mutations were excluded in both a control population and all unaffected family members tested. These findings reveal that a clinical phenotype distinct from classic BCIE but with similar histology can result from K1 mutations and that mutations at this codon give rise to a clinically unique condition.

DNA Mutational Analysis↗
PubMed1999

Identification of an ATP-driven, osmoregulated glycine betaine transport system in Listeria monocytogenes.

The ability of the gram-positive, food-borne pathogen Listeria monocytogenes to tolerate environments of elevated osmolarity and reduced temperature is due in part to the transport and accumulation of the osmolyte glycine betaine. Previously we showed that glycine betaine transport was the result of Na(+)-glycine betaine symport. In this report, we identify a second glycine betaine transporter from L. monocytogenes which is osmotically activated but does not require a high concentration of Na(+) for activity. By using a pool of Tn917-LTV3 mutants, a salt- and chill-sensitive mutant which was also found to be impaired in its ability to transport glycine betaine was isolated. DNA sequence analysis of the region flanking the site of transposon insertion revealed three open reading frames homologous to opuA from Bacillus subtilis and proU from Escherichia coli, both of which encode glycine betaine transport systems that belong to the superfamily of ATP-dependent transporters. The three open reading frames are closely spaced, suggesting that they are arranged in an operon. Moreover, a region upstream from the first reading frame was found to be homologous to the promoter regions of both opuA and proU. One unusual feature not shared with these other two systems is that the start codons for two of the open reading frames in L. monocytogenes appear to be TTG. That glycine betaine uptake is nearly eliminated in the mutant strain when it is assayed in the absence of Na(+) is an indication that only the ATP-dependent transporter and the Na(+)-glycine betaine symporter occur in L. monocytogenes.

Adenosine Triphosphate↗