Rapid Evolution of Beta-Keratin Genes Contribute to Phenotypic Differences That Distinguish Turtles and Birds from Other Reptiles

PubMed Central

Li, Yang I.; Kong, Lesheng; Ponting, Chris P.; Haerty, Wilfried

2013-01-01

Sequencing of vertebrate genomes permits changes in distinct protein families, including gene gains and losses, to be ascribed to lineage-specific phenotypes. A prominent example of this is the large-scale duplication of beta-keratin genes in the ancestors of birds, which was crucial to the subsequent evolution of their beaks, claws, and feathers. Evidence suggests that the shell of Pseudomys nelsoni contains at least 16 beta-keratins proteins, but it is unknown whether this is a complete set and whether their corresponding genes are orthologous to avian beak, claw, or feather beta-keratin genes. To address these issues and to better understand the evolution of the turtle shell at a molecular level, we surveyed the diversity of beta-keratin genes from the genome assemblies of three turtles, Chrysemys picta, Pelodiscus sinensis, and Chelonia mydas, which together represent over 160 Myr of chelonian evolution. For these three turtles, we found 200 beta-keratins, which indicate that, as for birds, a large expansion of beta-keratin genes in turtles occurred concomitantly with the evolution of a unique phenotype, namely, their plastron and carapace. Phylogenetic reconstruction of beta-keratin gene evolution suggests that separate waves of gene duplication within a single genomic location gave rise to scales, claws, and feathers in birds, and independently the scutes of the shell in turtles. PMID:23576313

Autosomal-dominant Meesmann epithelial corneal dystrophy without an exon mutation in the keratin-3 or keratin-12 gene in a Chinese family.

PubMed

Cao, Wei; Yan, Ming; Hao, QianYun; Wang, ShuLin; Wu, LiHua; Liu, Qing; Li, MingYan; Biddle, Fred G; Wu, Wei

2013-04-01

Meesmann epithelial corneal dystrophy (MECD) is a dominantly inherited disorder, characterized by fragility of the anterior corneal epithelium and formation of intraepithelial microcysts. It has been described in a number of different ancestral groups. To date, all reported cases of MECD have been associated with either a single mutation in one exon of the keratin-3 gene (KRT3) or a single mutation in one of two exons of the keratin-12 gene (KRT12). Each mutation leads to a predicted amino acid change in the respective keratin-3 or keratin-12 proteins that combine to form the corneal-specific heterodimeric intermediate filament protein. This case report describes a four-generation Chinese kindred with typical autosomal-dominant MECD. Exon sequencing of KRT3 and KRT12 in six affected and eight unaffected individuals (including two spouses) did not detect any mutations or nucleotide sequence variants. This kindred demonstrates that single mis-sense mutations may be sufficient but are not required in all individuals with the MECD phenotype. It provides a unique opportunity to investigate further genomic and functional heterogeneity in MECD.

Identification of a feather β-keratin gene exclusively expressed in pennaceous barbule cells of contour feathers in chicken.

PubMed

Kowata, Kinue; Nakaoka, Minori; Nishio, Kaori; Fukao, Ayaka; Satoh, Akira; Ogoshi, Maho; Takahashi, Sumio; Tsudzuki, Masaoki; Takeuchi, Sakae

2014-05-25

Feathers are elaborate skin appendages shared by birds and theropod dinosaurs that have hierarchical branching of the rachis, barbs, and barbules. Feather filaments consist of β-keratins encoded by multiple genes, most of which are located in tandem arrays on chromosomes 2, 25, and 27 in chicken. The expansion of the genes is thought to have contributed to feather evolution; however, it is unclear how the individual genes are involved in feather formation. The aim of the present study was to identify feather keratin genes involved in the formation of barbules. Using a combination of microarray analysis, reverse-transcription polymerase chain reaction, and in situ hybridization, we found an uncharacterized keratin gene on chromosome 7 that was expressed specifically in barbule cells in regenerating chicken feathers. We have named the gene barbule specific keratin 1 (BlSK1). The BlSK1 gene structure was similar to the gene structure of previously characterized feather keratin genes, and consisted of a non-coding leader exon, an intron, and an exon with an open reading frame (ORF). The ORF was predicted to encode a 98 aa long protein, which shared 59% identity with feather keratin B. Orthologs of BlSK1 were found in the genomes of other avian species, including turkey, duck, zebra finch, and flycatcher, in regions that shared synteny with chromosome 7 of chicken. Interestingly, BlSK1 was expressed in feather follicles that generated pennaceous barbules but not in follicles that generated plumulaceous barbules. These results suggested that the composition of feather keratins probably varies depending on the structure of the feather filaments and, that individual feather keratin genes may be involved in building different portions and/or types of feathers in chicken. Copyright © 2014 Elsevier B.V. All rights reserved.

Structure and functions of keratin proteins in simple, stratified, keratinized and cornified epithelia

PubMed Central

Bragulla, Hermann H; Homberger, Dominique G

2009-01-01

Historically, the term ‘keratin’ stood for all of the proteins extracted from skin modifications, such as horns, claws and hooves. Subsequently, it was realized that this keratin is actually a mixture of keratins, keratin filament-associated proteins and other proteins, such as enzymes. Keratins were then defined as certain filament-forming proteins with specific physicochemical properties and extracted from the cornified layer of the epidermis, whereas those filament-forming proteins that were extracted from the living layers of the epidermis were grouped as ‘prekeratins’ or ‘cytokeratins’. Currently, the term ‘keratin’ covers all intermediate filament-forming proteins with specific physicochemical properties and produced in any vertebrate epithelia. Similarly, the nomenclature of epithelia as cornified, keratinized or non-keratinized is based historically on the notion that only the epidermis of skin modifications such as horns, claws and hooves is cornified, that the non-modified epidermis is a keratinized stratified epithelium, and that all other stratified and non-stratified epithelia are non-keratinized epithelia. At this point in time, the concepts of keratins and of keratinized or cornified epithelia need clarification and revision concerning the structure and function of keratin and keratin filaments in various epithelia of different species, as well as of keratin genes and their modifications, in view of recent research, such as the sequencing of keratin proteins and their genes, cell culture, transfection of epithelial cells, immunohistochemistry and immunoblotting. Recently, new functions of keratins and keratin filaments in cell signaling and intracellular vesicle transport have been discovered. It is currently understood that all stratified epithelia are keratinized and that some of these keratinized stratified epithelia cornify by forming a Stratum corneum. The processes of keratinization and cornification in skin modifications are

Introducing a Null Mutation in the Mouse K6α and K6β Genes Reveals Their Essential Structural Role in the Oral Mucosa

PubMed Central

Wong, Pauline; Colucci-Guyon, Emma; Takahashi, Kenzo; Gu, Changhong; Babinet, Charles; Coulombe, Pierre A.

2000-01-01

Mammalian genomes feature multiple genes encoding highly related keratin 6 (K6) isoforms. These type II keratins show a complex regulation with constitutive and inducible components in several stratified epithelia, including the oral mucosa and skin. Two functional genes, K6α and K6β, exist in a head-to-tail tandem array in mouse genomes. We inactivated these two genes simultaneously via targeting and homologous recombination. K6 null mice are viable and initially indistinguishable from their littermates. Starting at two to three days after birth, they show a growth delay associated with reduced milk intake and the presence of white plaques in the posterior region of dorsal tongue and upper palate. These regions are subjected to greater mechanical stress during suckling. Morphological analyses implicate the filiform papillae as being particularly sensitive to trauma in K6α/K6β null mice, and establish the complete absence of keratin filaments in their anterior compartment. All null mice die about a week after birth. These studies demonstrate an essential structural role for K6 isoforms in the oral mucosa, and implicate filiform papillae as being the major stress bearing structures in dorsal tongue epithelium. PMID:10953016

New consensus nomenclature for mammalian keratins

PubMed Central

Schweizer, Jürgen; Bowden, Paul E.; Coulombe, Pierre A.; Langbein, Lutz; Lane, E. Birgitte; Magin, Thomas M.; Maltais, Lois; Omary, M. Bishr; Parry, David A.D.; Rogers, Michael A.; Wright, Mathew W.

2006-01-01

Keratins are intermediate filament–forming proteins that provide mechanical support and fulfill a variety of additional functions in epithelial cells. In 1982, a nomenclature was devised to name the keratin proteins that were known at that point. The systematic sequencing of the human genome in recent years uncovered the existence of several novel keratin genes and their encoded proteins. Their naming could not be adequately handled in the context of the original system. We propose a new consensus nomenclature for keratin genes and proteins that relies upon and extends the 1982 system and adheres to the guidelines issued by the Human and Mouse Genome Nomenclature Committees. This revised nomenclature accommodates functional genes and pseudogenes, and although designed specifically for the full complement of human keratins, it offers the flexibility needed to incorporate additional keratins from other mammalian species. PMID:16831889

Sulfur mustard induces the formation of keratin aggregates in human epidermal keratinocytes.

PubMed

Dillman, James F; McGary, Kriston L; Schlager, John J

2003-12-01

The vesicant sulfur mustard is an alkylating agent that has the capacity to cross-link biological molecules. We are interested in identifying specific proteins that are altered upon sulfur mustard exposure. Keratins are particularly important for the structural integrity of skin, and several genetically inherited blistering diseases have been linked to mutations in keratin 5 and keratin 14. We examined whether sulfur mustard exposure alters keratin biochemistry in cultured human epidermal keratinocytes. Western blotting with specific monoclonal antibodies revealed the formation of stable high-molecular-weight "aggregates" containing keratin 14 and/or keratin 5. These aggregates begin to form within 15 min after sulfur mustard exposure. These aggregates display a complex gel electrophoresis pattern between approximately 100 and approximately 200 kDa. Purification and analysis of these aggregates by one- and two-dimensional gel electrophoresis and mass spectrometry confirmed the presence of keratin 14 and keratin 5 and indicate that at least some of the aggregates are composed of keratin 14-keratin 14, keratin 14-keratin 5, or keratin 5-keratin 5 dimers. These studies demonstrate that sulfur mustard induces keratin aggregation in keratinocytes and support further investigation into the role of keratin aggregation in sulfur mustard-induced vesication.

Keratin 5 overexpression is associated with serous ovarian cancer recurrence and chemotherapy resistance.

PubMed

Ricciardelli, Carmela; Lokman, Noor A; Pyragius, Carmen E; Ween, Miranda P; Macpherson, Anne M; Ruszkiewicz, Andrew; Hoffmann, Peter; Oehler, Martin K

2017-03-14

This study investigated the clinical significance of keratin 5 and 6 expression in serous ovarian cancer progression and chemotherapy resistance. KRT5 and KRT6 (KRT6A, KRT6B & KRT6C) gene expression was assessed in publically available serous ovarian cancer data sets, ovarian cancer cell lines and primary serous ovarian cancer cells. Monoclonal antibodies which detect both K5/6 or only K5 were used to assess protein expression in ovarian cancer cell lines and a cohort of high grade serous ovarian carcinomas at surgery (n = 117) and after neoadjuvant chemotherapy (n = 21). Survival analyses showed that high KRT5 mRNA in stage III/IV serous ovarian cancers was significantly associated with reduced progression-free (HR 1.38, P < 0.0001) and overall survival (HR 1.28, P = 0.013) whilst high KRT6 mRNA was only associated with reduced progression-free survival (HR 1.2, P = 0.031). Both high K5/6 (≥ 10%, HR 1.78 95% CI; 1.03-2.65, P = 0.017) and high K5 (≥ 10%, HR 1.90, 95% CI; 1.12-3.19, P = 0.017) were associated with an increased risk of disease recurrence. KRT5 but not KRT6C mRNA expression was increased in chemotherapy resistant primary serous ovarian cancer cells compared to chemotherapy sensitive cells. The proportion of serous ovarian carcinomas with high K5/6 or high K5 immunostaining was significantly increased following neoadjuvant chemotherapy. K5 can be used to predict serous ovarian cancer prognosis and identify cancer cells that are resistant to chemotherapy. Developing strategies to target K5 may therefore improve serous ovarian cancer survival.

Keratin 17 null mice exhibit age- and strain-dependent alopecia

PubMed Central

McGowan, Kevin M.; Tong, Xuemei; Colucci-Guyon, Emma; Langa, Francina; Babinet, Charles; Coulombe, Pierre A.

2002-01-01

Onset of type I keratin 17 (K17) synthesis marks the adoption of an appendageal fate within embryonic ectoderm, and its expression persists in specific cell types within mature hair, glands, and nail. We report that K17 null mice develop severe alopecia during the first week postbirth, correlating with hair fragility, alterations in follicular histology, and apoptosis in matrix cells. These alterations are incompletely penetrant and normalize starting with the first postnatal cycle. Absence of a hair phenotype correlates with a genetic strain-dependent compensation by related keratins, including K16. These findings reveal a crucial role for K17 in the structural integrity of the first hair produced and the survival of hair-producing cells. Given that identical inherited mutations in this gene can cause either pachyonychia congenita or steatocystoma multiplex, the features of this mouse model suggest that this clinical heterogeneity arises from a cell type-specific, genetically determined compensation by related keratins. PMID:12050118

Keratin 17 null mice exhibit age- and strain-dependent alopecia.

PubMed

McGowan, Kevin M; Tong, Xuemei; Colucci-Guyon, Emma; Langa, Francina; Babinet, Charles; Coulombe, Pierre A

2002-06-01

Onset of type I keratin 17 (K17) synthesis marks the adoption of an appendageal fate within embryonic ectoderm, and its expression persists in specific cell types within mature hair, glands, and nail. We report that K17 null mice develop severe alopecia during the first week postbirth, correlating with hair fragility, alterations in follicular histology, and apoptosis in matrix cells. These alterations are incompletely penetrant and normalize starting with the first postnatal cycle. Absence of a hair phenotype correlates with a genetic strain-dependent compensation by related keratins, including K16. These findings reveal a crucial role for K17 in the structural integrity of the first hair produced and the survival of hair-producing cells. Given that identical inherited mutations in this gene can cause either pachyonychia congenita or steatocystoma multiplex, the features of this mouse model suggest that this clinical heterogeneity arises from a cell type-specific, genetically determined compensation by related keratins.

Hydrogels from feather keratin show higher viscoelastic properties and cell proliferation than those from hair and wool keratins.

PubMed

Esparza, Yussef; Bandara, Nandika; Ullah, Aman; Wu, Jianping

2018-09-01

Hydrogel prepared from keratin shows potential applications in tissue engineering. However, the importance of the keratin sources has not been considered. The objectives of this study were to characterize and compare the rheological (storage modulus), physical (porosity, pore size, swelling capacity, and water contact angle) and in vitro cell compatibility of hydrogel scaffolds prepared from various keratin sources. Keratins were characterized by means of their molecular weight, amino acid composition, thermal and conformational properties. Hydrogels from chicken feather keratins demonstrated substantially higher storage modulus (G') than hair and wool keratin hydrogels. However, higher swelling capacity (>3000%) was determined in hair and wool over feather keratin (1500%) hydrogels. Our results suggest that small molecular weight and β-sheet conformation of feather keratin (~10 kDa) facilitated the self-assembly of rigid hydrogels through disulfide bond re-oxidation. Whereas, high molecular weight (10-75 kDa) stretchable α-helix conformation in hair and wool keratins resulted in weaker hydrogels. The cell cultures using fibroblasts showed the highest proliferation rate on chicken feather keratin hydrogel scaffolds. After 15 days of culture, partial breakdown of keratin fibers was observed. Results indicate that stiffer avian keratins can be used to fabricate more mechanically robust biomaterials than mammalian keratins. Copyright © 2018 Elsevier B.V. All rights reserved.

Keratin capped silver nanoparticles - synthesis and characterization of a nanomaterial with desirable handling properties

USDA-ARS?s Scientific Manuscript database

Silver nanoparticles (NPs) were produced with keratin stabilizer and the NPs exhibited unimodal Gaussian distribution with average diameter of 3.5nm +/- 0.7 nm. The molecular mass of keratin stabilizer was 6-8 kDa. The mass of keratin capped NPs was >250 kDa to indicate the formation of crosslinked...

Soft epidermis of a scaleless snake lacks beta-keratin.

PubMed

Toni, M; Alibardi, L

2007-01-01

Beta-keratins are responsible for the mechanical resistance of scales in reptiles. In a scaleless crotalus snake (Crotalus atrox), large areas of the skin are completely devoid of scales, and the skin appears delicate and wrinkled. The epidermis of this snake has been assessed for the presence of beta-keratin by immunocytochemistry and immunoblotting using an antibody against chicken scale beta-keratin. This antibody recognizes beta-keratins in normal snake scales with molecular weights of 15-18 kDa and isoelectric points at 6.8, 7.5, 8.3 and 9.4. This indicates that beta-keratins of the stratum corneum are mainly basic proteins, so may interact with cytokeratins of the epidermis, most of which appear acidic (isoelectric points 4.5-5.5). A beta-layer and beta-keratin immunoreactivity are completely absent in moults of the scaleless mutant, and the corneous layer comprises a multi-layered alpha-layer covered by a flat oberhautchen. In conclusion, the present study shows that a lack of beta-keratins is correlated with the loss of scales and mechanical protection in the skin of this mutant snake.

Forty keratin-associated beta-proteins (beta-keratins) form the hard layers of scales, claws, and adhesive pads in the green anole lizard, Anolis carolinensis.

PubMed

Dalla Valle, Luisa; Nardi, Alessia; Bonazza, Giulia; Zucal, Chiara; Zuccal, Chiara; Emera, Deena; Alibardi, Lorenzo

2010-01-15

Using bioinformatic methods we have detected the genes of 40 keratin-associated beta-proteins (KAbetaPs) (beta-keratins) from the first available draft genome sequence of a reptile, the lizard Anolis carolinensis (Broad Institute, Boston). All genes are clustered in a single but not yet identified chromosomal locus, and contain a single intron of variable length. 5'-RACE and RT-PCR analyses using RNA from different epidermal regions show tissue-specific expression of different transcripts. These results were confirmed from the analysis of the A. carolinensis EST libraries (Broad Institute). Most deduced proteins are 12-16 kDa with a pI of 7.5-8.5. Two genes encoding putative proteins of 40 and 45 kDa are also present. Despite variability in amino acid sequences, four main subfamilies can be described. The largest subfamily includes proteins high in glycine, a small subfamily contains proteins high in cysteine, a third large subfamily contains proteins high in cysteine and glycine, and the fourth, smallest subfamily comprises proteins low in cysteine and glycine. An inner region of high amino acid identity is the most constant characteristic of these proteins and maps to a region with two to three close beta-folds in the proteins. This beta-fold region is responsible for the formation of filaments of the corneous material in all types of scales in this species. Phylogenetic analysis shows that A. carolinensis KAbetaPs are more similar to those of other lepidosaurians (snake, lizard, and gecko lizard) than to those of archosaurians (chick and crocodile) and turtles. (c) 2009 Wiley-Liss, Inc.

Keratins Are Altered in Intestinal Disease-Related Stress Responses

PubMed Central

Helenius, Terhi O.; Antman, Cecilia A.; Asghar, Muhammad Nadeem; Nyström, Joel H.; Toivola, Diana M.

2016-01-01

Keratin (K) intermediate filaments can be divided into type I/type II proteins, which form obligate heteropolymers. Epithelial cells express type I-type II keratin pairs, and K7, K8 (type II) and K18, K19 and K20 (type I) are the primary keratins found in the single-layered intestinal epithelium. Keratins are upregulated during stress in liver, pancreas, lung, kidney and skin, however, little is known about their dynamics in the intestinal stress response. Here, keratin mRNA, protein and phosphorylation levels were studied in response to murine colonic stresses modeling human conditions, and in colorectal cancer HT29 cells. Dextran sulphate sodium (DSS)-colitis was used as a model for intestinal inflammatory stress, which elicited a strong upregulation and widened crypt distribution of K7 and K20. K8 levels were slightly downregulated in acute DSS, while stress-responsive K8 serine-74 phosphorylation (K8 pS74) was increased. By eliminating colonic microflora using antibiotics, K8 pS74 in proliferating cells was significantly increased, together with an upregulation of K8 and K19. In the aging mouse colon, most colonic keratins were upregulated. In vitro, K8, K19 and K8 pS74 levels were increased in response to lipopolysaccharide (LPS)-induced inflammation in HT29 cells. In conclusion, intestinal keratins are differentially and dynamically upregulated and post-translationally modified during stress and recovery. PMID:27626448

Keratins Are Altered in Intestinal Disease-Related Stress Responses.

PubMed

Helenius, Terhi O; Antman, Cecilia A; Asghar, Muhammad Nadeem; Nyström, Joel H; Toivola, Diana M

2016-09-10

Keratin (K) intermediate filaments can be divided into type I/type II proteins, which form obligate heteropolymers. Epithelial cells express type I-type II keratin pairs, and K7, K8 (type II) and K18, K19 and K20 (type I) are the primary keratins found in the single-layered intestinal epithelium. Keratins are upregulated during stress in liver, pancreas, lung, kidney and skin, however, little is known about their dynamics in the intestinal stress response. Here, keratin mRNA, protein and phosphorylation levels were studied in response to murine colonic stresses modeling human conditions, and in colorectal cancer HT29 cells. Dextran sulphate sodium (DSS)-colitis was used as a model for intestinal inflammatory stress, which elicited a strong upregulation and widened crypt distribution of K7 and K20. K8 levels were slightly downregulated in acute DSS, while stress-responsive K8 serine-74 phosphorylation (K8 pS74) was increased. By eliminating colonic microflora using antibiotics, K8 pS74 in proliferating cells was significantly increased, together with an upregulation of K8 and K19. In the aging mouse colon, most colonic keratins were upregulated. In vitro, K8, K19 and K8 pS74 levels were increased in response to lipopolysaccharide (LPS)-induced inflammation in HT29 cells. In conclusion, intestinal keratins are differentially and dynamically upregulated and post-translationally modified during stress and recovery.

Genetic variants in pachyonychia congenita-associated keratins increase susceptibility to tooth decay

PubMed Central

Carlson, Jenna C.; Karacz, Chelsea M.; Schwartz, Mary E.; Cross, Michael A.; Marazita, Mary L.

2018-01-01

Pachyonychia congenita (PC) is a cutaneous disorder primarily characterized by nail dystrophy and painful palmoplantar keratoderma. PC is caused by mutations in KRT6A, KRT6B, KRT6C, KRT16, and KRT17, a set of keratin genes expressed in the nail bed, palmoplantar epidermis, oral mucosal epithelium, hair follicle and sweat gland. RNA-seq analysis revealed that all PC-associated keratins (except for Krt6c that does exist in the mouse genome) are expressed in the mouse enamel organ. We further demonstrated that these keratins are produced by ameloblasts and are incorporated into mature human enamel. Using genetic and intraoral examination data from 573 adults and 449 children, we identified several missense polymorphisms in KRT6A, KRT6B and KRT6C that lead to a higher risk for dental caries. Structural analysis of teeth from a PC patient carrying a p.Asn171Lys substitution in keratin-6a (K6a) revealed disruption of enamel rod sheaths resulting in altered rod shape and distribution. Finally, this PC-associated substitution as well as more frequent caries-associated SNPs, found in two of the KRT6 genes, that result in p.Ser143Asn substitution (rs28538343 in KRT6B and rs151117600 in KRT6C), alter the assembly of K6 filaments in ameloblast-like cells. These results identify a new set of keratins involved in tooth enamel formation, distinguish novel susceptibility loci for tooth decay and reveal additional clinical features of pachyonychia congenita. PMID:29357356

Genetic variants in pachyonychia congenita-associated keratins increase susceptibility to tooth decay.

PubMed

Duverger, Olivier; Carlson, Jenna C; Karacz, Chelsea M; Schwartz, Mary E; Cross, Michael A; Marazita, Mary L; Shaffer, John R; Morasso, Maria I

2018-01-01

Pachyonychia congenita (PC) is a cutaneous disorder primarily characterized by nail dystrophy and painful palmoplantar keratoderma. PC is caused by mutations in KRT6A, KRT6B, KRT6C, KRT16, and KRT17, a set of keratin genes expressed in the nail bed, palmoplantar epidermis, oral mucosal epithelium, hair follicle and sweat gland. RNA-seq analysis revealed that all PC-associated keratins (except for Krt6c that does exist in the mouse genome) are expressed in the mouse enamel organ. We further demonstrated that these keratins are produced by ameloblasts and are incorporated into mature human enamel. Using genetic and intraoral examination data from 573 adults and 449 children, we identified several missense polymorphisms in KRT6A, KRT6B and KRT6C that lead to a higher risk for dental caries. Structural analysis of teeth from a PC patient carrying a p.Asn171Lys substitution in keratin-6a (K6a) revealed disruption of enamel rod sheaths resulting in altered rod shape and distribution. Finally, this PC-associated substitution as well as more frequent caries-associated SNPs, found in two of the KRT6 genes, that result in p.Ser143Asn substitution (rs28538343 in KRT6B and rs151117600 in KRT6C), alter the assembly of K6 filaments in ameloblast-like cells. These results identify a new set of keratins involved in tooth enamel formation, distinguish novel susceptibility loci for tooth decay and reveal additional clinical features of pachyonychia congenita.

Complete Structure of an Epithelial Keratin Dimer: Implications for Intermediate Filament Assembly.

PubMed

Bray, David J; Walsh, Tiffany R; Noro, Massimo G; Notman, Rebecca

2015-01-01

Keratins are cytoskeletal proteins that hierarchically arrange into filaments, starting with the dimer sub-unit. They are integral to the structural support of cells, in skin, hair and nails. In skin, keratin is thought to play a critical role in conferring the barrier properties and elasticity of skin. In general, the keratin dimer is broadly described by a tri-domain structure: a head, a central rod and a tail. As yet, no atomistic-scale picture of the entire dimer structure exists; this information is pivotal for establishing molecular-level connections between structure and function in intermediate filament proteins. The roles of the head and tail domains in facilitating keratin filament assembly and function remain as open questions. To address these, we report results of molecular dynamics simulations of the entire epithelial human K1/K10 keratin dimer. Our findings comprise: (1) the first three-dimensional structural models of the complete dimer unit, comprising of the head, rod and tail domains; (2) new insights into the chirality of the rod-domain twist gained from analysis of the full domain structure; (3) evidence for tri-subdomain partitioning in the head and tail domains; and, (4) identification of the residue characteristics that mediate non-covalent contact between the chains in the dimer. Our findings are immediately applicable to other epithelial keratins, such as K8/K18 and K5/K14, and to intermediate filament proteins in general.

Spatially correlated phenotyping reveals K5-positive luminal progenitor cells and p63-K5/14-positive stem cell-like cells in human breast epithelium.

PubMed

Boecker, Werner; van Horn, Laura; Stenman, Göran; Stürken, Christine; Schumacher, Udo; Loening, Thomas; Liesenfeld, Lukas; Korsching, Eberhard; Gläser, Doreen; Tiemann, Katharina; Buchwalow, Igor

2018-05-09

Understanding the mechanisms regulating human mammary epithelium requires knowledge of the cellular constituents of this tissue. Different and partially contradictory definitions and concepts describing the cellular hierarchy of mammary epithelium have been proposed, including our studies of keratins K5 and/or K14 as markers of progenitor cells. Furthermore, we and others have suggested that the p53 homolog p63 is a marker of human breast epithelial stem cells. In this investigation, we expand our previous studies by testing whether immunohistochemical staining with monospecific anti-keratin antibodies in combination with an antibody against the stem cell marker p63 might help refine the different morphologic phenotypes in normal breast epithelium. We used in situ multilabel staining for p63, different keratins, the myoepithelial marker smooth muscle actin (SMA), the estrogen receptor (ER), and Ki67 to dissect and quantify the cellular components of 16 normal pre- and postmenopausal human breast epithelial tissue samples at the single-cell level. Importantly, we confirm the existence of K5+ only cells and suggest that they, in contrast to the current view, are key luminal precursor cells from which K8/18+ progeny cells evolve. These cells are further modified by the expression of ER and Ki67. We have also identified a population of p63+K5+ cells that are only found in nipple ducts. Based on our findings, we propose a new concept of the cellular hierarchy of human breast epithelium, including K5 luminal lineage progenitors throughout the ductal-lobular axis and p63+K5+ progenitors confined to the nipple ducts.

Sulfur mustard disrupts human α3β1-integrin receptors in concert with α6β4-integrin receptors and collapse of the keratin K5/K14 cytoskeleton

NASA Astrophysics Data System (ADS)

Werrlein, Robert J.; Braue, Catherine R.

2004-06-01

Sulfur mustard (SM; bis(2-chloroethyl) sulfide) is a chemical warfare agent that produces persistent, incapacitating blisters of the skin. The lesions inducing vesication remain elusive, and there is no completely effective treatment. Using mulitphoton microscopy and immunofluorescent staining, we found that exposing human epidermal keratinocytes (HEK) and intact epidermis to SM (400 μm for 5 min) caused progressive collapse of the keratin (K5/K14) cytoskeleton and depletion of α6β integrins. We now report that SM causes concomitant disruption nad collapse of the basal cell's α3β1-integrin receptors. At 1 h postexposure, images of Alexa488-conjugated HEK/α3β1 integrins showed almost complete withdrawal and disappearance of retraction fibers and a progressive loss of polarized mobility. With stero imaging, in vitro expression of this SM effect was characterized by collapse and abutment of adjacent cell membranes. At 2 h postexposure, there was an average 13% dorso-ventral collapse of HEK membranes that paralleled progressive collapse of the K5/K14 cytoskeleton. α3β1 integrin, like α6β4 integrin, is a regulator of cytoskeletal assembly, a receptor for laminin 5 and a mediator of HEK attachment to the basement membrane. Our images indicate that SM disrupts these receptors. We suggest that the progressive disruption destabilizes and potentiates blistering of the epidermal-dermal junction.

Simple Epithelial Keratins.

PubMed

Strnad, Pavel; Guldiken, Nurdan; Helenius, Terhi O; Misiorek, Julia O; Nyström, Joel H; Lähdeniemi, Iris A K; Silvander, Jonas S G; Kuscuoglu, Deniz; Toivola, Diana M

2016-01-01

Simple epithelial keratins (SEKs) are the cytoplasmic intermediate filament proteins of single-layered and glandular epithelial cells as found in the liver, pancreas, intestine, and lung. SEKs have broad cytoprotective functions, which are facilitated by dynamic posttranslational modifications and interaction with associated proteins. SEK filaments are composed of obligate heteropolymers of type II (K7, K8) and type I (K18-K20, K23) keratins. The multifaceted roles of SEKs are increasingly appreciated due to findings obtained from transgenic mouse models and human studies that identified SEK variants in several digestive diseases. Reorganization of the SEK network into aggregates called Mallory-Denk bodies (MDBs) is characteristic for specific liver disorders such as alcoholic and nonalcoholic steatohepatitis. To spur further research on SEKs, we here review the methods and potential caveats of their isolation as well as possibilities to study them in cell culture. The existing transgenic SEK mouse models, their advantages and potential drawbacks are discussed. The tools to induce MDBs, ways of their visualization and quantification, as well as the possibilities to detect SEK variants in humans are summarized. Copyright © 2016 Elsevier Inc. All rights reserved.

The X-Ray Crystal Structure of the Keratin 1-Keratin 10 Helix 2B Heterodimer Reveals Molecular Surface Properties and Biochemical Insights into Human Skin Disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bunick, Christopher G.; Milstone, Leonard M.

Keratins 1 (K1) and 10 (K10) are the primary keratins expressed in differentiated epidermis. Mutations in K1/K10 are associated with human skin diseases. We determined the crystal structure of the complex between the distal (2B) helices of K1 and K10 to better understand how human keratin structure correlates with function. The 3.3 Å resolution structure confirms many features inferred by previous biochemical analyses, but adds unexpected insights. It demonstrates a parallel, coiled-coil heterodimer with a predominantly hydrophobic intermolecular interface; this heterodimer formed a higher order complex with a second K1-K10-2B heterodimer via a Cys401K10 disulfide link, although the bond anglemore » is unanticipated. The molecular surface analysis of K1-K10-2B identified several pockets, one adjacent to the disulfide linkage and conserved in K5-K14. The solvent accessible surface area of the K1-K10 structure is 20–25% hydrophobic. The 2B region contains mixed acidic and basic patches proximally (N-terminal), whereas it is largely acidic distally (C-terminal). Mapping of conserved and nonconserved residues between K1-K10 and K5-K14 onto the structure demonstrated the majority of unique residues align along the outer helical ridge. Finally, the structure permitted a fresh analysis of the deleterious effects caused by K1/K10 missense mutations found in patients with phenotypic skin disease.« less

Ultrastructural localization of hair keratins, high sulfur keratin-associated proteins and sulfhydryl oxidase in the human hair.

PubMed

Alibardi, Lorenzo

2017-03-01

Hardening of the human hair shaft during cornification results from the bonding of keratins and keratin-associated proteins. In situ hybridization and light immunocytochemical studies have shown the general distribution of different keratins and some associated proteins but not determined their ultrastructural localization. I report here the localization of hair keratins, two high-sulfur keratin-associated proteins and sulfhydryl oxidase has been studied under the transmission electron microscope in the cornification zone of the human hair. The ultrastructural study on keratin distribution in general confirms previous light microscopic studies. Sulfur-rich KAP1 is mainly cortical but the labeling disappears in fully cornified cortical cells while a diffuse labeling is also present in differentiating cuticle cells. Sulfur-rich K26 immunolocalization is only detected in the exocuticle and endocuticle. Sparse labeling for sulfhydryl oxidase occurs in differentiating cortical cells but is weak and uneven in cuticle cells and absent in medulla and inner root sheath. Labeling disappears in the upper fully cornified cortex and cuticle. The observations indicate that sulfhydryl oxidase and keratin associated proteins are initially produced in the cytoplasm among keratin bundles accumulating in cortical and cuticle cells but these proteins undergo changes during the following cornification that alter the epitopes tagged by the antibodies.

Autosomal Recessive Hypotrichosis with Woolly Hair Caused by a Mutation in the Keratin 25 Gene Expressed in Hair Follicles.

PubMed

Zernov, Nikolay V; Skoblov, Mikhail Y; Marakhonov, Andrey V; Shimomura, Yutaka; Vasilyeva, Tatyana A; Konovalov, Fedor A; Abrukova, Anna V; Zinchenko, Rena A

2016-06-01

Hypotrichosis is an abnormal condition characterized by decreased hair density and various defects in hair structure and growth patterns. In particular, in woolly hair, hypotrichosis is characterized by a tightly curled structure and abnormal growth. In this study, we present a detailed comparative examination of individuals affected by autosomal-recessive hypotrichosis (ARH), which distinguishes two types of ARH. Earlier, we demonstrated that exon 4 deletion in the lipase H gene caused an ARH (hypotrichosis 7; MIM: 604379) in populations of the Volga-Ural region of Russia. Screening for this mutation in all affected individuals revealed its presence only in the group with the hypotrichosis 7 phenotype. Other patients formed a separate group of woolly hair-associated ARH, with a homozygous missense mutation c.712G>T (p.Val238Leu) in a highly conserved position of type I keratin KRT25 (K25). Haplotype analysis indicated a founder effect. An expression study in the HaCaT cell line demonstrated a deleterious effect of the p.Val238Leu mutation on the formation of keratin intermediate filaments. Hence, we have identified a previously unreported missense mutation in the KRT25 gene causing ARH with woolly hair. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Genomic Organization, Transcriptomic Analysis, and Functional Characterization of Avian α- and β-Keratins in Diverse Feather Forms

PubMed Central

Fan, Wen-Lang; Yan, Jie; Chen, Chih-Kuan; Lai, Yu-Ting; Wu, Siao-Man; Mao, Chi-Tang; Chen, Jun-Jie; Lu, Mei-Yeh Jade; Ho, Meng-Ru; Widelitz, Randall B.; Chen, Chih-Feng; Chuong, Cheng-Ming; Li, Wen-Hsiung

2014-01-01

Feathers are hallmark avian integument appendages, although they were also present on theropods. They are composed of flexible corneous materials made of α- and β-keratins, but their genomic organization and their functional roles in feathers have not been well studied. First, we made an exhaustive search of α- and β-keratin genes in the new chicken genome assembly (Galgal4). Then, using transcriptomic analysis, we studied α- and β-keratin gene expression patterns in five types of feather epidermis. The expression patterns of β-keratin genes were different in different feather types, whereas those of α-keratin genes were less variable. In addition, we obtained extensive α- and β-keratin mRNA in situ hybridization data, showing that α-keratins and β-keratins are preferentially expressed in different parts of the feather components. Together, our data suggest that feather morphological and structural diversity can largely be attributed to differential combinations of α- and β-keratin genes in different intrafeather regions and/or feather types from different body parts. The expression profiles provide new insights into the evolutionary origin and diversification of feathers. Finally, functional analysis using mutant chicken keratin forms based on those found in the human α-keratin mutation database led to abnormal phenotypes. This demonstrates that the chicken can be a convenient model for studying the molecular biology of human keratin-based diseases. PMID:25152353

Keratins Regulate p38MAPK-Dependent Desmoglein Binding Properties in Pemphigus

PubMed Central

Vielmuth, Franziska; Walter, Elias; Fuchs, Michael; Radeva, Mariya Y.; Buechau, Fanny; Magin, Thomas M.; Spindler, Volker; Waschke, Jens

2018-01-01

Keratins are crucial for the anchorage of desmosomes. Severe alterations of keratin organization and detachment of filaments from the desmosomal plaque occur in the autoimmune dermatoses pemphigus vulgaris and pemphigus foliaceus (PF), which are mainly caused by autoantibodies against desmoglein (Dsg) 1 and 3. Keratin alterations are a structural hallmark in pemphigus pathogenesis and correlate with loss of intercellular adhesion. However, the significance for autoantibody-induced loss of intercellular adhesion is largely unknown. In wild-type (wt) murine keratinocytes, pemphigus autoantibodies induced keratin filament retraction. Under the same conditions, we used murine keratinocytes lacking all keratin filaments (KtyII k.o.) as a model system to dissect the role of keratins in pemphigus. KtyII k.o. cells show compromised intercellular adhesion without antibody (Ab) treatment, which was not impaired further by pathogenic pemphigus autoantibodies. Nevertheless, direct activation of p38MAPK via anisomycin further decreased intercellular adhesion indicating that cell cohesion was not completely abrogated in the absence of keratins. Direct inhibition of Dsg3, but not of Dsg1, interaction via pathogenic autoantibodies as revealed by atomic force microscopy was detectable in both cell lines demonstrating that keratins are not required for this phenomenon. However, PF-IgG shifted Dsg1-binding events from cell borders toward the free cell surface in wt cells. This led to a distribution pattern of Dsg1-binding events similar to KtyII k.o. cells under resting conditions. In keratin-deficient keratinocytes, PF-IgG impaired Dsg1-binding strength, which was not different from wt cells under resting conditions. In addition, pathogenic autoantibodies were capable of activating p38MAPK in both KtyII wt and k.o. cells, the latter of which already displayed robust p38MAPK activation under resting conditions. Since inhibition of p38MAPK blocked autoantibody-induced loss of

Model-based analysis of keratin intermediate filament assembly

NASA Astrophysics Data System (ADS)

Martin, Ines; Leitner, Anke; Walther, Paul; Herrmann, Harald; Marti, Othmar

2015-09-01

The cytoskeleton of epithelial cells consists of three types of filament systems: microtubules, actin filaments and intermediate filaments (IFs). Here, we took a closer look at type I and type II IF proteins, i.e. keratins. They are hallmark constituents of epithelial cells and are responsible for the generation of stiffness, the cellular response to mechanical stimuli and the integrity of entire cell layers. Thereby, keratin networks constitute an important instrument for cells to adapt to their environment. In particular, we applied models to characterize the assembly of keratin K8 and K18 into elongated filaments as a means for network formation. For this purpose, we measured the length of in vitro assembled keratin K8/K18 filaments by transmission electron microscopy at different time points. We evaluated the experimental data of the longitudinal annealing reaction using two models from polymer chemistry: the Schulz-Zimm model and the condensation polymerization model. In both scenarios one has to make assumptions about the reaction process. We compare how well the models fit the measured data and thus determine which assumptions fit best. Based on mathematical modelling of experimental filament assembly data we define basic mechanistic properties of the elongation reaction process.

Topographical mapping of α- and β-keratins on developing chicken skin integuments: Functional interaction and evolutionary perspectives

PubMed Central

Wu, Ping; Ng, Chen Siang; Yan, Jie; Lai, Yung-Chih; Chen, Chih-Kuan; Lai, Yu-Ting; Wu, Siao-Man; Chen, Jiun-Jie; Luo, Weiqi; Widelitz, Randall B.; Li, Wen-Hsiung; Chuong, Cheng-Ming

2015-01-01

Avian integumentary organs include feathers, scales, claws, and beaks. They cover the body surface and play various functions to help adapt birds to diverse environments. These keratinized structures are mainly composed of corneous materials made of α-keratins, which exist in all vertebrates, and β-keratins, which only exist in birds and reptiles. Here, members of the keratin gene families were used to study how gene family evolution contributes to novelty and adaptation, focusing on tissue morphogenesis. Using chicken as a model, we applied RNA-seq and in situ hybridization to map α- and β-keratin genes in various skin appendages at embryonic developmental stages. The data demonstrate that temporal and spatial α- and β-keratin expression is involved in establishing the diversity of skin appendage phenotypes. Embryonic feathers express a higher proportion of β-keratin genes than other skin regions. In feather filament morphogenesis, β-keratins show intricate complexity in diverse substructures of feather branches. To explore functional interactions, we used a retrovirus transgenic system to ectopically express mutant α- or antisense β-keratin forms. α- and β-keratins show mutual dependence and mutations in either keratin type results in disrupted keratin networks and failure to form proper feather branches. Our data suggest that combinations of α- and β-keratin genes contribute to the morphological and structural diversity of different avian skin appendages, with feather-β-keratins conferring more possible composites in building intrafeather architecture complexity, setting up a platform of morphological evolution of functional forms in feathers. PMID:26598683

Monoclonal Antibody Analysis of Keratin Expression in the Central Nervous System

NASA Astrophysics Data System (ADS)

Franko, Maryellen C.; Gibbs, Clarence J.; Rhoades, Dorothy A.; Carleton Gajdusek, D.

1987-05-01

A monoclonal antibody directed against a 65-kDa brain protein demonstrates an epitope found in keratin from human epidermis. By indirect immunofluorescence, the antibody decorates intracytoplasmic filaments in a subclass of astrocytes and Purkinje cells of adult hamster brain. Double-label immunofluorescence study using antibody to glial fibrillary acidic protein and this antibody reveals the 65-kDa protein to be closely associated with glial filaments in astrocytes of fetal mouse brain cultures. Immunoblot analysis of purified human epidermal keratin and hamster brain homogenate confirms the reactivity of this antibody to epidermal keratin polypeptides. All the major epidermal keratins were recognized by this antibody. It did not bind to the remaining major intermediate filament proteins. These findings suggest that monoclonal antibody 34C9 recognizes a cytoskeletal structure connected with intermediate filaments. In addition, the monoclonal antibody demonstrates that epidermal keratins share an epitope not only among themselves but also with a ``neural keratin.''

A novel role of the NRF2 transcription factor in the regulation of arsenite-mediated keratin 16 gene expression in human keratinocytes.

PubMed

Endo, Hitoshi; Sugioka, Yoshihiko; Nakagi, Yoshihiko; Saijo, Yasuaki; Yoshida, Takahiko

2008-07-01

Inorganic sodium arsenite (iAs) is a ubiquitous environmental contaminant and is associated with an increased risk of skin hyperkeratosis and cancer. We investigated the molecular mechanisms underlying the regulation of the keratin 16 (K16) gene by iAs in the human keratinocyte cell line HaCaT. We performed reverse transcriptase polymerase chain reaction, luciferase assays, Western blots, and electrophoretic mobility shift assays to determine the transcriptional regulation of the K16 gene by iAs. We used gene overexpression approaches to elucidate the nuclear factor erythroid-derived 2 related factor 2 (NRF2) involved in the K16 induction. iAs induced the mRNA and protein expression of K16. We also found that the expression of K16 was transcriptionally induced by iAs through activator protein-1-like sites and an antioxidant response element (ARE) in its gene promoter region. Treatment with iAs also enhanced the production and translocation of the NRF2 transcription factor, an ARE-binding protein, into the nucleus without modification of its mRNA expression. In addition, iAs elongated the half-life of the NRF2 protein. When overexpressed in HaCaT cells, NRF2 was also directly involved in not only the up-regulation of the detoxification gene thioredoxin but also K16 gene expression. Our data clearly indicate that the K16 gene is a novel target of NRF2. Furthermore, our findings also suggest that NRF2 has opposing roles in the cell--in the activation of detoxification pathways and in promoting the development of skin disorders.

Non-Coding Keratin Variants Associate with Liver Fibrosis Progression in Patients with Hemochromatosis

PubMed Central

Lunova, Mariia; Guldiken, Nurdan; Lienau, Tim C.; Stickel, Felix; Omary, M. Bishr

2012-01-01

Background Keratins 8 and 18 (K8/K18) are intermediate filament proteins that protect the liver from various forms of injury. Exonic K8/K18 variants associate with adverse outcome in acute liver failure and with liver fibrosis progression in patients with chronic hepatitis C infection or primary biliary cirrhosis. Given the association of K8/K18 variants with end-stage liver disease and progression in several chronic liver disorders, we studied the importance of keratin variants in patients with hemochromatosis. Methods The entire K8/K18 exonic regions were analyzed in 162 hemochromatosis patients carrying homozygous C282Y HFE (hemochromatosis gene) mutations. 234 liver-healthy subjects were used as controls. Exonic regions were PCR-amplified and analyzed using denaturing high-performance liquid chromatography and DNA sequencing. Previously-generated transgenic mice overexpressing K8 G62C were studied for their susceptibility to iron overload. Susceptibility to iron toxicity of primary hepatocytes that express K8 wild-type and G62C was also assessed. Results We identified amino-acid-altering keratin heterozygous variants in 10 of 162 hemochromatosis patients (6.2%) and non-coding heterozygous variants in 6 additional patients (3.7%). Two novel K8 variants (Q169E/R275W) were found. K8 R341H was the most common amino-acid altering variant (4 patients), and exclusively associated with an intronic KRT8 IVS7+10delC deletion. Intronic, but not amino-acid-altering variants associated with the development of liver fibrosis. In mice, or ex vivo, the K8 G62C variant did not affect iron-accumulation in response to iron-rich diet or the extent of iron-induced hepatocellular injury. Conclusion In patients with hemochromatosis, intronic but not exonic K8/K18 variants associate with liver fibrosis development. PMID:22412904

Vitamin K3 (menadione)-induced oncosis associated with keratin 8 phosphorylation and histone H3 arylation.

PubMed

Scott, Gary K; Atsriku, Christian; Kaminker, Patrick; Held, Jason; Gibson, Brad; Baldwin, Michael A; Benz, Christopher C

2005-09-01

The vitamin K analog menadione (K3), capable of both redox cycling and arylating nucleophilic substrates by Michael addition, has been extensively studied as a model stress-inducing quinone in both cell culture and animal model systems. Exposure of keratin 8 (k-8) expressing human breast cancer cells (MCF7, T47D, SKBr3) to K3 (50-100 microM) induced rapid, sustained, and site-specific k-8 serine phosphorylation (pSer73) dependent on signaling by a single mitogen activated protein kinase (MAPK) pathway, MEK1/2. Normal nuclear morphology and k-8 immunofluorescence coupled with the lack of DNA laddering or other features of apoptosis indicated that K3-induced cytotoxicity, evident within 4 h of treatment and delayed but not prevented by MEK1/2 inhibition, was due to a form of stress-activated cell death known as oncosis. Independent of MAPK signaling was the progressive appearance of K3-induced cellular fluorescence, principally nuclear in origin and suggested by in vitro fluorimetry to have been caused by K3 thiol arylation. Imaging by UV transillumination of protein gels containing nuclear extracts from K3-treated cells revealed a prominent 17-kDa band shown to be histone H3 by immunoblotting and mass spectrometry (MS). K3 arylation of histones in vitro followed by electrospray ionization-tandem MS analyses identified the unique Cys110 residue within H3, exposed only in the open chromatin of transcriptionally active genes, as a K3 arylation target. These findings delineate new pathways associated with K3-induced stress and suggest a potentially novel role for H3 Cys110 as a nuclear stress sensor.

Binding Interactions of Keratin-Based Hair Fiber Extract to Gold, Keratin, and BMP-2

PubMed Central

de Guzman, Roche C.; Tsuda, Shanel M.; Ton, Minh-Thi N.; Zhang, Xiao; Esker, Alan R.; Van Dyke, Mark E.

2015-01-01

Hair-derived keratin biomaterials composed mostly of reduced keratin proteins (kerateines) have demonstrated their utility as carriers of biologics and drugs for tissue engineering. Electrostatic forces between negatively-charged keratins and biologic macromolecules allow for effective drug retention; attraction to positively-charged growth factors like bone morphogenetic protein 2 (BMP-2) has been used as a strategy for osteoinduction. In this study, the intermolecular surface and bulk interaction properties of kerateines were investigated. Thiol-rich kerateines were chemisorbed onto gold substrates to form an irreversible 2-nm rigid layer for surface plasmon resonance analysis. Kerateine-to-kerateine cohesion was observed in pH-neutral water with an equilibrium dissociation constant (KD) of 1.8 × 10−4 M, indicating that non-coulombic attractive forces (i.e. hydrophobic and van der Waals) were at work. The association of BMP-2 to kerateine was found to be greater (KD = 1.1 × 10−7 M), within the range of specific binding. Addition of salts (phosphate-buffered saline; PBS) shortened the Debye length or the electrostatic field influence which weakened the kerateine-BMP-2 binding (KD = 3.2 × 10−5 M). BMP-2 in bulk kerateine gels provided a limited release in PBS (~ 10% dissociation in 4 weeks), suggesting that electrostatic intermolecular attraction was significant to retain BMP-2 within the keratin matrix. Complete dissociation between kerateine and BMP-2 occurred when the PBS pH was lowered (to 4.5), below the keratin isoelectric point of 5.3. This phenomenon can be attributed to the protonation of keratin at a lower pH, leading to positive-positive repulsion. Therefore, the dynamics of kerateine-BMP-2 binding is highly dependent on pH and salt concentration, as well as on BMP-2 solubility at different pH and molarity. The study findings may contribute to our understanding of the release kinetics of drugs from keratin biomaterials and allow for the

Rhomboid family member 2 regulates cytoskeletal stress-associated Keratin 16

PubMed Central

Maruthappu, Thiviyani; Chikh, Anissa; Fell, Benjamin; Delaney, Paul J.; Brooke, Matthew A.; Levet, Clemence; Moncada-Pazos, Angela; Ishida-Yamamoto, Akemi; Blaydon, Diana; Waseem, Ahmad; Leigh, Irene M.; Freeman, Matthew; Kelsell, David P.

2017-01-01

Keratin 16 (K16) is a cytoskeletal scaffolding protein highly expressed at pressure-bearing sites of the mammalian footpad. It can be induced in hyperproliferative states such as wound healing, inflammation and cancer. Here we show that the inactive rhomboid protease RHBDF2 (iRHOM2) regulates thickening of the footpad epidermis through its interaction with K16. K16 expression is absent in the thinned footpads of irhom2−/− mice compared with irhom2+/+mice, due to reduced keratinocyte proliferation. Gain-of-function mutations in iRHOM2 underlie Tylosis with oesophageal cancer (TOC), characterized by palmoplantar thickening, upregulate K16 with robust downregulation of its type II keratin binding partner, K6. By orchestrating the remodelling and turnover of K16, and uncoupling it from K6, iRHOM2 regulates the epithelial response to physical stress. These findings contribute to our understanding of the molecular mechanisms underlying hyperproliferation of the palmoplantar epidermis in both physiological and disease states, and how this ‘stress' keratin is regulated. PMID:28128203

Novel mutations in the helix termination motif of keratin 3 and keratin 12 in 2 Taiwanese families with Meesmann corneal dystrophy.

PubMed

Chen, Ying-Ting; Tseng, Sung-Huei; Chao, Sheau-Chiou

2005-11-01

To analyze mutations of the keratin 3 gene (KRT3) and keratin 12 gene (KRT12) in 2 Taiwanese families with Meesmann corneal dystrophy (MCD). Diagnosis of MCD was confirmed by slit-lamp examination of the cornea in 4 members of family 1 and 6 members of family 2. All exons and flanking intron boundaries of KRT3 and KRT12 were amplified by polymerase chain reaction (PCR), and products were subjected to direct sequencing. Restriction fragment length polymorphism analysis (RFLP) with created mismatch primers, Bst XI and Nsp I, was used to confirm the presence of the mutations in affected individuals in family 1 and family 2, respectively. A novel heterozygous missense mutation (1508G-->C), predicting the substitution of a proline for an arginine (R503P) was detected in the helix termination motif of the keratin 3 polypeptide in family 1. Another novel heterozygous missense mutation (1286A-->G), predicting the substitution of a cysteine for a tyrosine at codon 429 (Y429C) was detected in the helix termination motif of the keratin 12 polypeptide in family 2. These 2 mutations were excluded from 50 normal controls by RFLP analysis, indicating that they were not common polymorphisms. A novel missense mutation (R503P) in KRT3 and another novel missense mutation (Y429C) in KRT12 lead to MCD in 2 unrelated Taiwanese families. The mutant codons in our study are all located in the highly conserved alpha-helix-termination motif, which is essential for keratin filament assembly. Mutation at this area may account for the disruption of keratin filament assembly, leading to MCD.

Highly Conserved Keratin-Associated Protein 7-1 Gene in Yak, Taurine and Zebu Cattle.

PubMed

Arlud, S; He, N; Sari, E M; Ma, Z-J; Zhang, H; An, T-W; Han, J-L

2017-01-01

Keratin-associated proteins (KRTAPs) play a critical role in cross-linking the keratin intermediate filaments to build a hair shaft. The genetic polymorphisms of the bovine KRTAP7-1 gene were investigated for the first time in this study. The complete coding sequence of the KRTAP7-1 gene in 108 domestic yak, taurine and zebu cattle from China and Indonesia were successfully amplified using polymerase chain reaction and then directly sequenced. Only two single-nucleotide polymorphisms (one nonsynonymous at c.7C/G and another synonymous at c.21C/T) and three haplotypes (BOVIN-KRTAP7-1*A, B and C) were identified in the complete coding sequence of the bovine KRTAP7-1 gene among all animals. There was no polymorphism across three Chinese indigenous yak breeds and one Indonesian zebu cattle population, all sharing the BOVINKRTAP71*A haplotype. The four taurine cattle populations also had BOVIN-KRTAP7-1*A as the most common haplotype with a frequency of 0.80. The frequency of novel haplotype BOVIN-KRTAP7-1*B was only 0.07 present in one heterozygous animal in each of the four taurine cattle populations, while BOVINKRTAP7- 1*C was only found in a Simmental and a local Chinese Yellow cattle population with frequencies of 0.17 and 0.36, respectively. The monomorphic yak KRTAP7-1 gene in particular, and highly conserved bovine, sheep and goat KRTAP7-1 genes in general, demonstrated its unique intrinsic structural property (e.g., > 21% high glycine content) and primary functional importance in supporting the mechanical strength and shape of hair.

ERK2-mediated C-terminal serine phosphorylation of p300 is vital to the regulation of epidermal growth factor-induced keratin 16 gene expression.

PubMed

Chen, Yun-Ju; Wang, Ying-Nai; Chang, Wen-Chang

2007-09-14

We previously reported that the epidermal growth factor (EGF) regulates the gene expression of keratin 16 by activating the extracellular signal-regulated kinase 1 and 2 (ERK1/2) signaling which in turn enhances the recruitment of p300 to the keratin 16 promoter. The recruited p300 functionally cooperates with Sp1 and c-Jun to regulate the gene expression of keratin 16. This study investigated in detail the molecular events incurred upon p300 whereby EGF caused an enhanced interaction between p300 and Sp1. EGF apparently induced time- and dose-dependent phosphorylation of p300, both in vitro and in vivo, through the activation of ERK2. The six potential ERK2 phosphorylation sites, including three threonine and three serine residues as revealed by sequential analysis, were first identified in vitro. Confirmation of these six sites in vivo indicated that these three serine residues (Ser-2279, Ser-2315, and Ser-2366) on the C terminus of p300 were the major signaling targets of EGF. Furthermore, the C-terminal serine phosphorylation of p300 stimulated its histone acetyltransferase activity and enhanced its interaction with Sp1. These serine phosphorylation sites on p300 controlled the p300 recruitment to the keratin 16 promoter. When all three serine residues on p300 were replaced by alanine, EGF could no longer induce the gene expression of keratin 16. Taken together, these results strongly suggested that the ERK2-mediated C-terminal serine phosphorylation of p300 was a key event in the regulation of EGF-induced keratin 16 expression. These results also constituted the first report identifying the unique p300 phosphorylation sites induced by ERK2 in vivo.

A novel mutation of the Keratin 12 gene responsible for a severe phenotype of Meesmann's corneal dystrophy

PubMed Central

Sullivan, Lori S.; Baylin, Eric B.; Font, Ramon; Daiger, Stephen P.; Pepose, Jay S.; Clinch, Thomas E.; Nakamura, Hisashi; Zhao, Xinping C.

2007-01-01

Purpose To determine if a mutation within the coding region of the keratin 12 gene (KRT12) is responsible for a severe form of Meesmann's corneal dystrophy. Methods A family with clinically identified Meesmann's corneal dystrophy was recruited and studied. Electron microscopy was performed on scrapings of corneal epithelial cells from the proband. Mutations in the KRT12 gene were sought using direct genomic sequencing of leukocyte DNA from two affected and two unaffected family members. Subsequently, the observed mutation was screened in all available family members using polymerase chain reaction and direct sequencing. Results A heterozygous missense mutation (Arg430Pro) was found in exon 6 of KRT12 in all 14 affected individuals studied. Unaffected family members and 100 normal controls were negative for this mutation. Conclusions We have identified a novel mutation in the KRT12 gene that is associated with a symptomatic phenotype of Meesmann's corneal dystrophy. This mutation results in a substitution of proline for arginine in the helix termination motif that may disrupt the normal helix, leading to a dramatic structural change of the keratin 12 protein. PMID:17653038

Keratin 5/14‑mediated cell differentiation and transformation are regulated by TAp63 and Notch‑1 in oral squamous cell carcinoma‑derived cells.

PubMed

Srivastava, Saumya S; Alam, Hunain; Patil, Sonam J; Shrinivasan, Rashmi; Raikundalia, Sweta; Chaudhari, Pratik Rajeev; Vaidya, Milind M

2018-05-01

Keratins 5/14 (K5/14) are intermediate filament proteins expressed in the basal layer of stratified epithelial cells and are known targets of p63. Previous research in our laboratory showed that upon K5/14 downregulation in oral squamous cell carcinoma (OSCC)‑derived cells, there was an increase in intracellular Notch‑1 levels and differentiation markers such as involucrin, keratin 1 and a decrease in tumorigenic potential in vivo. However, the molecules involved in the K14 regulated cell differentiation and transformation are not known to date. In order to understand the possible role of TAp63, we downregulated TAp63 in a K14‑knockdown background. We observed that there was a decrease in the expression of Notch‑1. Expression levels of differentiation markers such as involucrin, K1, loricrin and filaggrin were also decreased. Furthermore, TAp63 downregulation led to an increase in invasion, migration and in vivo tumorigenic potential of these cells. We observed a decrease in β‑catenin signaling in K14‑downregulated cells. Notably, when TAp63 was downregulated in K14‑knockdown cells, there was increase in non‑phospho β‑catenin levels. Hence, this study indicates that TAp63 plays an important role in K14‑downregulated cells possibly by regulating the Notch‑1 expression. K14 regulates the expression of TAp63 which in turn regulates expression of Notch‑1. The present study is a step forward in our quest to understand the functional significance of molecules that regulate the process of differentiation and tumorigenesis in stratified epithelial cells.

Interplay between Solo and keratin filaments is crucial for mechanical force–induced stress fiber reinforcement

PubMed Central

Fujiwara, Sachiko; Ohashi, Kazumasa; Mashiko, Toshiya; Kondo, Hiroshi; Mizuno, Kensaku

2016-01-01

Mechanical force–induced cytoskeletal reorganization is essential for cell and tissue remodeling and homeostasis; however, the underlying cellular mechanisms remain elusive. Solo (ARHGEF40) is a RhoA-targeting guanine nucleotide exchange factor (GEF) involved in cyclical stretch–induced human endothelial cell reorientation and convergent extension cell movement in zebrafish gastrula. In this study, we show that Solo binds to keratin-8/keratin-18 (K8/K18) intermediate filaments through multiple sites. Solo overexpression promotes the formation of thick actin stress fibers and keratin bundles, whereas knockdown of Solo, expression of a GEF-inactive mutant of Solo, or inhibition of ROCK suppresses stress fiber formation and leads to disorganized keratin networks, indicating that the Solo-RhoA-ROCK pathway serves to precisely organize keratin networks, as well as to promote stress fibers. Of importance, knockdown of Solo or K18 or overexpression of GEF-inactive or deletion mutants of Solo suppresses tensile force–induced stress fiber reinforcement. Furthermore, knockdown of Solo or K18 suppresses tensile force-induced RhoA activation. These results strongly suggest that the interplay between Solo and K8/K18 filaments plays a crucial role in tensile force–induced RhoA activation and consequent actin cytoskeletal reinforcement. PMID:26823019

Keratinizing dentigerous cyst

PubMed Central

Sivasankar, Vaishnavi; Ranganathan, Kannan; Praveen, B

2014-01-01

Keratinizing dentigerous cyst is a rare entity. This article reports a case of keratinizing dentigerous cyst associated with an impacted mandibular canine. Clinical and radiological features, cone-beam computed tomography findings and histological features of the case are reported along with a discussion on keratinizing odontogenic cysts and the need for follow-up. PMID:24808713

Keratins 17 and 19 expression as prognostic markers in oral squamous cell carcinoma.

PubMed

Coelho, B A; Peterle, G T; Santos, M; Agostini, L P; Maia, L L; Stur, E; Silva, C V M; Mendes, S O; Almança, C C J; Freitas, F V; Borçoi, A R; Archanjo, A B; Mercante, A M C; Nunes, F D; Carvalho, M B; Tajara, E H; Louro, I D; Silva-Conforti, A M A

2015-11-25

Five-year survival rates for oral squamous cell carcinoma (OSCC) are 30% and the mortality rate is 50%. Immunohistochemistry panels are used to evaluate proliferation, vascularization, apoptosis, HPV infection, and keratin expression, which are important markers of malignant progression. Keratins are a family of intermediate filaments predominantly expressed in epithelial cells and have an essential role in mechanical support and cytoskeleton formation, which is essential for the structural integrity and stability of the cell. In this study, we analyzed the expressions of keratins 17 and 19 (K17 and K19) by immunohistochemistry in tumoral and non-tumoral tissues from patients with OSCC. The results show that expression of these keratins is higher in tumor tissues compared to non-tumor tissues. Positive K17 expression correlates with lymph node metastasis and multivariate analysis confirmed this relationship, revealing a 6-fold increase in lymph node metastasis when K17 is expressed. We observed a correlation between K17 expression with disease-free survival and disease-specific death in patients who received surgery and radiotherapy. Multivariate analysis revealed that low expression of K17 was an independent marker for early disease relapse and disease-specific death in patients treated with surgery and radiotherapy, with an approximately 4-fold increased risk when compared to high K17 expression. Our results suggest a potential role for K17 and K19 expression profiles as tumor prognostic markers in OSCC patients.

Hair growth promoting activity of discarded biocomposite keratin extract.

PubMed

Akanda, Md Rashedunnabi; Kim, Hak-Yong; Park, Mira; Kim, In-Shik; Ahn, Dongchoon; Tae, Hyun-Jin; Park, Byung-Yong

2017-08-01

Keratin biomaterial has been used in regenerative medicine owing to its in-vivo and in-vitro biocompatibility. The present study was aimed to investigate the hair growth promoting activity of keratin extract and its mechanism of action. Keratin extract was topically applied on the synchronized depilated dorsal skin of telogenic C57BL/6 mice and promoted hair growth by inducing the anagen phase. The histomorphometric observation indicated significantly increases the number, shaft of hair follicles and deep subcutis area in the keratin extract treated group in contrast to the control group, which was considered an indication of anagen phase induction. Subsequently, the quantitative real-time polymerase chain reaction analysis revealed that fibroblast growth factor-10, vascular endothelial growth factor, insulin-like growth factor-1, β-catenin, and Shh were expressed earlier in the keratin extract-treated group than in the control group. Besides, keratin extract has been observed to be biocompatible when analyzed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 4',6-diamidino-2-phenylindole staining using immortalized human keratinocyte cells, showing more than 90% cell viability. Our study demonstrated that keratin extract stimulating hair follicle growth by inducing the growth phase; anagen in telogenic C57BL/6 mice and thus the topical application of keratin extract may represent a promising biomaterial for the management and applications of hair follicle disorder.

Keratin sponge/hydrogel part 1. fabrication and characterization

USDA-ARS?s Scientific Manuscript database

Keratin sponge/hydrogel products formed by either the oxidation or reduction of U.S. domestic fine- or coarse-grade wool exhibited distinctively different topologies and molecular weights of 6- 8 kDa and 40-60 kDa, each with unique macro-porous structure and microstructural behaviors. The sponge/ ...

Pure keratin membrane and fibers from chicken feather.

PubMed

Ma, Bomou; Qiao, Xue; Hou, Xiuliang; Yang, Yiqi

2016-08-01

In this research, keratin was extracted from the disposable chicken feather using l-cysteine as reducing agent. Then, it was re-dissolved in the sodium carbonate-sodium bicarbonate buffer, and the pure keratin membrane and fiber were fabricated by doctor-blade casting process and wet spinning method, respectively. Scanning electron microscopy (SEM), fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD) and thermogravimetric analysis (TGA) were used to characterize the chemical and physical properties of resulting powder, membrane and fiber. Compared with the raw chicken feather, the regenerated keratin materials retain its chemical structure and thermal stability, their relative crystallinity is a little different depend on the shaping method, which leads to the difference in moisture regain. The mechanical results show that tensile strength of the keratin membrane researches 3.5MPa, have potential application in biomedical fields. However, the keratin fiber presents low tenacity, i.e. 0.5cN/dtex, this problem should be solved in order to apply the new fiber in textile and material science. Copyright © 2016 Elsevier B.V. All rights reserved.

Keratin: dissolution, extraction and biomedical application.

PubMed

Shavandi, Amin; Silva, Tiago H; Bekhit, Adnan A; Bekhit, Alaa El-Din A

2017-08-22

Keratinous materials such as wool, feathers and hooves are tough unique biological co-products that usually have high sulfur and protein contents. A high cystine content (7-13%) differentiates keratins from other structural proteins, such as collagen and elastin. Dissolution and extraction of keratin is a difficult process compared to other natural polymers, such as chitosan, starch, collagen, and a large-scale use of keratin depends on employing a relatively fast, cost-effective and time efficient extraction method. Keratin has some inherent ability to facilitate cell adhesion, proliferation, and regeneration of the tissue, therefore keratin biomaterials can provide a biocompatible matrix for regrowth and regeneration of the defective tissue. Additionally, due to its amino acid constituents, keratin can be tailored and finely tuned to meet the exact requirement of degradation, drug release or incorporation of different hydrophobic or hydrophilic tails. This review discusses the various methods available for the dissolution and extraction of keratin with emphasis on their advantages and limitations. The impacts of various methods and chemicals used on the structure and the properties of keratin are discussed with the aim of highlighting options available toward commercial keratin production. This review also reports the properties of various keratin-based biomaterials and critically examines how these materials are influenced by the keratin extraction procedure, discussing the features that make them effective as biomedical applications, as well as some of the mechanisms of action and physiological roles of keratin. Particular attention is given to the practical application of keratin biomaterials, namely addressing the advantages and limitations on the use of keratin films, 3D composite scaffolds and keratin hydrogels for tissue engineering, wound healing, hemostatic and controlled drug release.

A homozygous missense variant in type I keratin KRT25 causes autosomal recessive woolly hair.

PubMed

Ansar, Muhammad; Raza, Syed Irfan; Lee, Kwanghyuk; Irfanullah; Shahi, Shamim; Acharya, Anushree; Dai, Hang; Smith, Joshua D; Shendure, Jay; Bamshad, Michael J; Nickerson, Deborah A; Santos-Cortez, Regie Lyn P; Ahmad, Wasim; Leal, Suzanne M

2015-10-01

Woolly hair (WH) is a hair abnormality that is primarily characterised by tightly curled hair with abnormal growth. In two unrelated consanguineous Pakistani families with non-syndromic autosomal recessive (AR) WH, homozygosity mapping and linkage analysis identified a locus within 17q21.1-q22, which contains the type I keratin gene cluster. A DNA sample from an affected individual from each family underwent exome sequencing. A homozygous missense variant c.950T>C (p.(Leu317Pro)) within KRT25 segregated with ARWH in both families, and has a combined maximum two-point LOD score of 7.9 at ϴ=0. The KRT25 variant is predicted to result in disruption of the second α-helical rod domain and the entire protein structure, thus possibly interfering with heterodimerisation of K25 with type II keratins within the inner root sheath (IRS) of the hair follicle and the medulla of the hair shaft. Our findings implicate a novel gene involved in human hair abnormality, and are consistent with the curled, fragile hair found in mice with Krt25 mutations, and further support the role of IRS-specific type I keratins in hair follicle development and maintenance of hair texture. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Keratins as components of the enamel organic matrix

PubMed Central

Duverger, Olivier; Beniash, Elia; Morasso, Maria I.

2016-01-01

Dental enamel is a hardest tissue in the human body, and although it starts as a tissue rich in proteins, by the time of eruption of the tooth in the oral cavity only a small fraction of the protein remains. While this organic matrix of enamel represents less than 1% by weight it plays essential roles in improving both toughness and resilience to chemical attacks. Despite the fact that the first studies of the enamel matrix began in the 19th century its exact composition and mechanisms of its function remain poorly understood. It was proposed that keratin or a keratin-like primitive epithelial component exists in mature enamel, however due to the extreme insolubility of its organic matrix the presence of keratins there was never clearly established. We have recently identified expression of a number of hair keratins in ameloblasts, the enamel secreting cells, and demonstrated their incorporation into mature enamel. Mutation in epithelial hair keratin KRT75 leads to a skin condition called pseudofollicularis barbae. Carriers of this mutation have an altered enamel structure and mechanical properties. Importantly, these individuals have a much higher prevalence of caries. To the best of our knowledge, this is the first study showing a direct link between a mutation in a protein-coding region of a gene and increased caries rates. In this paper we present an overview of the evidence of keratin-like material in enamel that has accumulated over the last 150 years. Furthermore, we propose potential mechanisms of action of KTR75 in enamel and highlight the clinical implications of the link between mutations in KRT75 and caries. Finally, we discuss the potential use of keratins for enamel repair. PMID:26709044

Interplay between Solo and keratin filaments is crucial for mechanical force-induced stress fiber reinforcement.

PubMed

Fujiwara, Sachiko; Ohashi, Kazumasa; Mashiko, Toshiya; Kondo, Hiroshi; Mizuno, Kensaku

2016-03-15

Mechanical force-induced cytoskeletal reorganization is essential for cell and tissue remodeling and homeostasis; however, the underlying cellular mechanisms remain elusive. Solo (ARHGEF40) is a RhoA-targeting guanine nucleotide exchange factor (GEF) involved in cyclical stretch-induced human endothelial cell reorientation and convergent extension cell movement in zebrafish gastrula. In this study, we show that Solo binds to keratin-8/keratin-18 (K8/K18) intermediate filaments through multiple sites. Solo overexpression promotes the formation of thick actin stress fibers and keratin bundles, whereas knockdown of Solo, expression of a GEF-inactive mutant of Solo, or inhibition of ROCK suppresses stress fiber formation and leads to disorganized keratin networks, indicating that the Solo-RhoA-ROCK pathway serves to precisely organize keratin networks, as well as to promote stress fibers. Of importance, knockdown of Solo or K18 or overexpression of GEF-inactive or deletion mutants of Solo suppresses tensile force-induced stress fiber reinforcement. Furthermore, knockdown of Solo or K18 suppresses tensile force-induced RhoA activation. These results strongly suggest that the interplay between Solo and K8/K18 filaments plays a crucial role in tensile force-induced RhoA activation and consequent actin cytoskeletal reinforcement. © 2016 Fujiwara et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Keratin expression profiling of transitional epithelium in the painful bladder syndrome/interstitial cystitis.

PubMed

Laguna, Pilar; Smedts, Frank; Nordling, Jörgen; Horn, Thomas; Bouchelouche, Kirsten; Hopman, Anton; de la Rosette, Jean

2006-01-01

Painful bladder syndrome/interstitial cystitis (PBS/IC) is a severely debilitating condition. Its cause is poorly understood; therapy is symptomatic and often unsuccessful. To study urothelial involvement, we characterized the keratin phenotype of bladder urothelium in 18 patients with PBS/IC using a panel of 11 keratin antibodies recognizing simple keratins found in columnar epithelia (keratins 7, 8, 18, and 20) and keratins associated with basal cell compartments of squamous epithelia (keratins 5, 13, 14, and 17). We also tested 2 antibodies recognizing more than 1 keratin also directed against basal cell compartments of squamous epithelia (D5/16 B4 and 34betaE12). Bladder urothelium in PBS/IC showed distinct differences in the profiles of keratins 7, 8, 14, 17, 18, and 20 compared with literature reports for normal bladder urothelium. These were characterized by a shift from the normal bladder urothelial keratin phenotype to a more squamous keratin profile, despite the lack of morphologic evidence of squamous epithelial differentiation and a loss of compartmentalization of keratin expression. The severity of these changes varied between biopsy specimens. Whether these changes are primary or secondary to another underlying condition remains to be determined.

Dissolution and characterization of biofunctional keratin particles extracted from chicken feathers

NASA Astrophysics Data System (ADS)

Sharma, Swati; Gupta, Arun; Chik, Syed Mohd Saufi Bin Tuan; Yeo Gek Kee, Chua; Poddar, Pradeep Kumar

2017-04-01

In the present study chicken feathers were hydrolyzed in alkaline environment. The pH value of feather hydrolyzed solution was adjusted according to the principle of isoelectric precipitation. Three kinds of precipitates of keratin polypeptide were collected at pH of 3.5, 5.5 and 7.5 respectively. The keratin solution were freeze dried and denoted as FKP1, FKP2, FKP3 respectively. All keratin particles possessed smooth, uniform and round surface by scanning electron microscope (SEM). FKP1, FKP2 and FKP3 had higher glass transition temperature examined by thermogravimetry (TG). Fourier transform infrared spectroscopy (FTIR) revealed that the extracted keratin retained the most of protein backbone, with the breakage of disulfide cross-links and hydrogen bonds.

Genomic regression of claw keratin, taste receptor and light-associated genes provides insights into biology and evolutionary origins of snakes.

PubMed

Emerling, Christopher A

2017-10-01

Regressive evolution of anatomical traits often corresponds with the regression of genomic loci underlying such characters. As such, studying patterns of gene loss can be instrumental in addressing questions of gene function, resolving conflicting results from anatomical studies, and understanding the evolutionary history of clades. The evolutionary origins of snakes involved the regression of a number of anatomical traits, including limbs, taste buds and the visual system, and by analyzing serpent genomes, I was able to test three hypotheses associated with the regression of these features. The first concerns two keratins that are putatively specific to claws. Both genes that encode these keratins are pseudogenized/deleted in snake genomes, providing additional evidence of claw-specificity. The second hypothesis is that snakes lack taste buds, an issue complicated by conflicting results in the literature. I found evidence that different snakes have lost one or more taste receptors, but all snakes examined retained at least one gustatory channel. The final hypothesis addressed is that the earliest snakes were adapted to a dim light niche. I found evidence of deleted and pseudogenized genes with light-associated functions in snakes, demonstrating a pattern of gene loss similar to other dim light-adapted clades. Molecular dating estimates suggest that dim light adaptation preceded the loss of limbs, providing some bearing on interpretations of the ecological origins of snakes. Copyright © 2017 Elsevier Inc. All rights reserved.

Association of keratin 8/18 variants with non-alcoholic fatty liver disease and insulin resistance in Chinese patients: A case-control study.

PubMed

Li, Rui; Liao, Xian-Hua; Ye, Jun-Zhao; Li, Min-Rui; Wu, Yan-Qin; Hu, Xuan; Zhong, Bi-Hui

2017-06-14

To test the hypothesis that K8/K18 variants predispose humans to non-alcoholic fatty liver disease (NAFLD) progression and its metabolic phenotypes. We selected a total of 373 unrelated adult subjects from our Physical Examination Department, including 200 unrelated NAFLD patients and 173 controls of both genders and different ages. Diagnoses of NAFLD were established according to ultrasonic signs of fatty liver. All subjects were tested for population characteristics, lipid profile, liver tests, as well as glucose tests. Genomic DNA was obtained from peripheral blood with a DNeasy Tissue Kit. K8/K18 coding regions were analyzed, including 15 exons and exon-intron boundaries. Among 200 NAFLD patients, 10 (5%) heterozygous carriers of keratin variants were identified. There were 5 amino-acid-altering heterozygous variants and 6 non-coding heterozygous variants. One novel amino-acid-altering heterozygous variant (K18 N193S) and three novel non-coding variants were observed (K8 IVS5-9A→G, K8 IVS6+19G→A, K18 T195T). A total of 9 patients had a single variant and 1 patient had compound variants (K18 N193S+K8 IVS3-15C→G). Only one R341H variant was found in the control group (1 of 173, 0.58%). The frequency of keratin variants in NAFLD patients was significantly higher than that in the control group (5% vs 0.58%, P = 0.015). Notably, the keratin variants were significantly associated with insulin resistance (IR) in NAFLD patients (8.86% in NAFLD patients with IR vs 2.5% in NAFLD patients without IR, P = 0.043). K8/K18 variants are overrepresented in Chinese NAFLD patients and might accelerate liver fat storage through IR.

Androgen regulation of the human hair follicle: the type I hair keratin hHa7 is a direct target gene in trichocytes.

PubMed

Jave-Suarez, Luis F; Langbein, Lutz; Winter, Hermelita; Praetzel, Silke; Rogers, Michael A; Schweizer, Juergen

2004-03-01

Previous work had shown that most members of the complex human hair keratin family were expressed in terminal scalp hairs. An exception to this rule was the type I hair keratin hHa7, which was only detected in some but not all vellus hairs of the human scalp (Langbein et al, 1999). Here we show that hHa7 exhibits constitutive expression in medullary cells of all types of male and female sexual hairs. Medullated beard, axillary, and pubic hairs arise during puberty from small, unmedullated vellus hairs under the influence of circulating androgens. This suggested an androgen-controlled expression of the hHa7 gene. Further evidence for this assumption was provided by the demonstration of androgen receptor (AR) expression in the nuclei of medullary cells of beard hairs. Moreover, homology search for the semipalindromic androgen receptor-binding element (ARE) consensus sequence GG(A)/(T)ACAnnnTGTTCT in the proximal hHa7 promoter revealed three putative ARE motifs. Electrophoretic mobility shift assays demonstrated the specific binding of AR to all three hHa7 AREs. Their function as AR-responsive elements, either individually or in concert within the hHa7 promoter, could be further confirmed by transfection studies with or without an AR expression vector in PtK2 and prostate PC3-Arwt cells, respectively in the presence or absence of a synthetic androgen. Our study detected hHa7 as the first gene in hair follicle trichocytes whose expression appears to be directly regulated by androgens. As such, hHa7 represents a marker for androgen action on hair follicles and might be a suitable tool for investigations of androgen-dependent hair disorders.

Reactivation of larval keratin gene (krt62.L) in blastema epithelium during Xenopus froglet limb regeneration.

PubMed

Satoh, Akira; Mitogawa, Kazumasa; Saito, Nanami; Suzuki, Miyuki; Suzuki, Ken-Ichi T; Ochi, Haruki; Makanae, Aki

2017-12-15

Limb regeneration is considered a form of limb redevelopment because of the molecular and morphological similarities. Forming a regeneration blastema is, in essence, creating a developing limb bud in an adult body. This reactivation of a developmental process in a mature body is worth studying. Xenopus laevis has a biphasic life cycle that involves distinct larval and adult stages. These distinct developmental stages are useful for investigating the reactivation of developmental processes in post-metamorphic frogs (froglets). In this study, we focused on the re-expression of a larval gene (krt62.L) during Xenopus froglet limb regeneration. Recently renamed krt62.L, this gene was known as the larval keratin (xlk) gene, which is specific to larval-tadpole stages. During limb regeneration in a froglet, krt62.L was re-expressed in a basal layer of blastema epithelium, where adult-specific keratin (Krt12.6.S) expression was also observable. Nerves produce important regulatory factors for amphibian limb regeneration, and also play a role in blastema formation and maintenance. The effect of nerve function on krt62.L expression could be seen in the maintenance of krt62.L expression, but not in its induction. When an epidermis-stripped limb bud was grafted in a froglet blastema, the grafted limb bud could reach the digit-forming stage. This suggests that krt62.L-positive froglet blastema epithelium is able to support the limb development process. These findings imply that the developmental process is locally reactivated in an postmetamorphic body during limb regeneration. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Measuring the regulation of keratin filament network dynamics

PubMed Central

Moch, Marcin; Herberich, Gerlind; Aach, Til; Leube, Rudolf E.; Windoffer, Reinhard

2013-01-01

The organization of the keratin intermediate filament cytoskeleton is closely linked to epithelial function. To study keratin network plasticity and its regulation at different levels, tools are needed to localize and measure local network dynamics. In this paper, we present image analysis methods designed to determine the speed and direction of keratin filament motion and to identify locations of keratin filament polymerization and depolymerization at subcellular resolution. Using these methods, we have analyzed time-lapse fluorescence recordings of fluorescent keratin 13 in human vulva carcinoma-derived A431 cells. The fluorescent keratins integrated into the endogenous keratin cytoskeleton, and thereby served as reliable markers of keratin dynamics. We found that increased times after seeding correlated with down-regulation of inward-directed keratin filament movement. Bulk flow analyses further revealed that keratin filament polymerization in the cell periphery and keratin depolymerization in the more central cytoplasm were both reduced. Treating these cells and other human keratinocyte-derived cells with EGF reversed all these processes within a few minutes, coinciding with increased keratin phosphorylation. These results highlight the value of the newly developed tools for identifying modulators of keratin filament network dynamics and characterizing their mode of action, which, in turn, contributes to understanding the close link between keratin filament network plasticity and epithelial physiology. PMID:23757496

Solo and keratin filaments regulate epithelial tubule morphology.

PubMed

Nishimura, Ryosuke; Kato, Kagayaki; Fujiwara, Sachiko; Ohashi, Kazumasa; Mizuno, Kensaku

2018-04-28

Epithelial tubules, consisting of the epithelial cell sheet with a central lumen, are the basic structure of many organs. Mechanical forces play an important role in epithelial tubulogenesis; however, little is known about the mechanisms controlling the mechanical forces during epithelial tubule morphogenesis. Solo (also known as ARHGEF40) is a RhoA-targeting guanine-nucleotide exchange factor that is involved in mechanical force-induced RhoA activation and stress fiber formation. Solo binds to keratin-8/keratin-18 (K8/K18) filaments, and this interaction plays a crucial role in mechanotransduction. In this study, we examined the roles of Solo and K8/K18 filaments in epithelial tubulogenesis using MDCK cells cultured in 3D collagen gels. Knockdown of either Solo or K18 resulted in rounder tubules with increased lumen size, indicating that Solo and K8/K18 filaments play critical roles in forming the elongated morphology of epithelial tubules. Moreover, knockdown of Solo or K18 decreased the level of diphosphorylated myosin light chain (a marker of contractile force) at the luminal and outer surfaces of tubules, suggesting that Solo and K8/K18 filaments are involved in the generation of the myosin II-mediated contractile force during epithelial tubule morphogenesis. In addition, K18 filaments were normally oriented along the long axis of the tubule, but knockdown of Solo perturbed their orientation. These results suggest that Solo plays crucial roles in forming the elongated morphology of epithelial tubules and in regulating myosin II activity and K18 filament organization during epithelial tubule formation.

Surface active complexes formed between keratin polypeptides and ionic surfactants.

PubMed

Pan, Fang; Lu, Zhiming; Tucker, Ian; Hosking, Sarah; Petkov, Jordan; Lu, Jian R

2016-12-15

Keratins are a group of important proteins in skin and hair and as biomaterials they can provide desirable properties such as strength, biocompatibility, and moisture regaining and retaining. The aim of this work is to develop water-soluble keratin polypeptides from sheep wool and then explore how their surface adsorption behaves with and without surfactants. Successful preparation of keratin samples was demonstrated by identification of the key components from gel electrophoresis and the reproducible production of gram scale samples with and without SDS (sodium dodecylsulphate) during wool fibre dissolution. SDS micelles could reduce the formation of disulphide bonds between keratins during extraction, reducing inter-molecular crosslinking and improving keratin polypeptide solubility. However, Zeta potential measurements of the two polypeptide batches demonstrated almost identical pH dependent surface charge distributions with isoelectric points around pH 3.5, showing complete removal of SDS during purification by dialysis. In spite of different solubility from the two batches of keratin samples prepared, very similar adsorption and aggregation behavior was revealed from surface tension measurements and dynamic light scattering. Mixing of keratin polypeptides with SDS and C 12 TAB (dodecyltrimethylammonium bromide) led to the formation of keratin-surfactant complexes that were substantially more effective at reducing surface tension than the polypeptides alone, showing great promise in the delivery of keratin polypeptides via the surface active complexes. Neutron reflection measurements revealed the coexistence of surfactant and keratin polypeptides at the interface, thus providing the structural support to the observed surface tension changes associated with the formation of the surface active complexes. Copyright © 2016. Published by Elsevier Inc.

Preparation of keratin and chemically modified keratin hydrogels and their evaluation as cell substrate with drug releasing ability.

PubMed

Nakata, Ryo; Osumi, Yu; Miyagawa, Shoko; Tachibana, Akira; Tanabe, Toshizumi

2015-07-01

Keratin was extracted as a reduced form from wool, which was then subjected to acetamidation, carboxymethylation or aminoethylation at abundant free cysteine residues to give acetamidated keratin (AAK), carboxymethylated keratin (CMK) and aminoethylated keratin (AEK). Hydrogels were prepared from intact and three chemically modified keratins simply by concentrating their aqueous solution and subsequent cooling. The lowest concentration to form a hydrogel without fluidity was 110 mg/ml for AAK, 120 mg/ml for AEK, 130 mg/ml for keratin and 180 mg/ml for CMK. Comparing with a hydrogel just prepared (swelling ratio: 600-700), each hydrogel slightly shrank in an acidic solution. While AAK hydrogel little swelled in neutral and basic solutions, other hydrogels became swollen and CMK hydrogel reached to dissolution. Hydrogels of keratin, AAK and AEK were found to support cell proliferation, although cell elongation on AAK and AEK hydrogel was a little suppressed. On the other hand, CMK hydrogel did not seem to be suitable for a cell substrate because of its high swelling in culture medium. Evaluation of the hydrogels as a drug carrier showed that keratin and AAK hydrogels were good sustained drug release carriers, which showed the drug release for more than three days, while the release from AEK and CMK hydrogels completed within one day. Thus, keratin and chemically modified keratin hydrogels, especially keratin and AAK hydrogels, were promising biomaterials as a cell substrate and a sustained drug release carrier. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters.

PubMed

Tong, Yaw-Chong; Chang, Shwu-Fen; Liu, Chia-Yang; Kao, Winston W-Y; Huang, Chong Heng; Liaw, Jiahorng

2007-11-01

This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for beta-galactosidase (beta-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-Gal) color staining, 1,2-dioxetane beta-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, beta-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may

Isolation of a new class of cysteine-glycine-proline-rich beta-proteins (beta-keratins) and their expression in snake epidermis.

PubMed

Dalla Valle, Luisa; Nardi, Alessia; Alibardi, Lorenzo

2010-03-01

Scales of snakes contain hard proteins (beta-keratins), now referred to as keratin-associated beta-proteins. In the present study we report the isolation, sequencing, and expression of a new group of these proteins from snake epidermis, designated cysteine-glycine-proline-rich proteins. One deduced protein from expressed mRNAs contains 128 amino acids (12.5 kDa) with a theoretical pI at 7.95, containing 10.2% cysteine and 15.6% glycine. The sequences of two more snake cysteine-proline-rich proteins have been identified from genomic DNA. In situ hybridization shows that the messengers for these proteins are present in the suprabasal and early differentiating beta-cells of the renewing scale epidermis. The present study shows that snake scales, as previously seen in scales of lizards, contain cysteine-rich beta-proteins in addition to glycine-rich beta-proteins. These keratin-associated beta-proteins mix with intermediate filament keratins (alpha-keratins) to produce the resistant corneous layer of snake scales. The specific proportion of these two subfamilies of proteins in different scales can determine various degrees of hardness in scales.

Constitutive expression of the K-domain of a Vaccinium corymbosum SOC1-like (VcSOC1-K) MADS-box gene is sufficient to promote flowering in tobacco.

PubMed

Song, Guo-qing; Walworth, Aaron; Zhao, Dongyan; Hildebrandt, Britton; Leasia, Michael

2013-11-01

The K-domain of a blueberry-derived SOC1 -like gene promotes flowering in tobacco without negatively impacting yield, demonstrating potential for manipulation of flowering time in horticultural crops. The SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) and SOC1-likes, belonging to the MIKC(c) (type II) MADS-box gene subfamily, are major floral activators and integrators of plant flowering. Both MADS-domains and K (Keratin)-domains are highly conserved in MIKC(c)-type MADS proteins. While there are many reports on overexpression of intact MIKC(c)-type MADS-box genes, few studies have been conducted to investigate the effects of the K-domains. In this report, a 474-bp K-domain of Vaccinium SOC1-like (VcSOC1-K) was cloned from the cDNA library of the northern highbush blueberry (Vaccinium corymbosum L.). Functional analysis of the VcSOC1-K was conducted by ectopically expressing of 35S:VcSOC1-K in tobacco. Reverse transcription PCR confirmed expression of the VcSOC1-K in T0 plants. Phenotypically, T1 transgenic plants (10 T1 plants/event) flowered sooner after seeding, and were shorter with fewer leaves at the time of flowering, than nontransgenic plants; but seed pod production of transgenic plants was not significantly affected. These results demonstrate that overexpression of the K-domain of a MIKC(c)-type MADS-box gene alone is sufficient to promote early flowering and more importantly without affecting seed production.

Heterogeneity of keratin expression and actin distribution in benign and malignant mammary diseases.

PubMed

Wada, T; Yasutomi, M; Yamada, K; Hashimura, K; Kunikata, M; Tanaka, T; Huang, J W; Mori, M

1991-01-01

Immunoreactivity of monoclonal anti-cytokeratin KL1, PKK1, K8.12 and anti-actin antibodies in 101 cases of diseased human breast lesions showed irregular keratin distribution in luminal cells of terminal ductal-lobular unit and basal layer cells of the interlobular and main duct. Actin staining was confined to myoepithelial cells. Benign lesions showed great heterogeneity in luminal cells of the terminal ductal-lobular units. Breast carcinoma showed a reduced staining for keratins, heterogeneity of keratin expression was found in solid tubular carcinoma, and actin was usually absent: however, papillo-ductal or comedo type had actin positive myoepithelial cells around carcinoma foci.

Mechanistic investigation of a hemostatic keratin biomaterial

NASA Astrophysics Data System (ADS)

Rahmany, Maria Bahawdory

Traumatic injury leads to more productive years lost than heart disease, cancer and stroke combined. Trauma is often accompanied and complicated by uncontrolled bleeding. Human hair keratin biomaterials have demonstrated efficacy in controlling hemorrhage in both small and large animal models; however little is known about the mechanism by which these proteins aid in blood clotting. Inspection of the amino acid sequence of known keratins shows the presence of several cellular binding motifs, suggesting a possible mechanism and potentially eliminating the need to functionalize the material's surface for cellular interaction. In addition to small animal studies, the hemostatic activity of keratin hydrogels was explored through porcine hemorrhage models representing both a high flow and low flow bleed. In both studies, keratin hydrogels appeared to lead to a significant reduction in blood loss. The promising results from these in vivo studies provided the motivation for this project. The objective of this dissertation work was to assess the mechanism of action of a hemostatic keratin biomaterial, and more broadly assess the biomaterial-cellular interaction(s). It is our hypothesis that keratin biomaterials have the capacity to specifically interact with cells and lead to propagation of intracellular signaling pathway, specifically contributing to hemostasis. Through application of biochemical and molecular tools, we demonstrate here that keratin biomaterials contribute to hemostasis through two probable mechanisms; integrin mediated platelet adhesion and increased fibrin polymerization. Platelets are the major cell type involved in coagulation both by acting as a catalytic surface for the clotting cascade and adhering to extracellular matrix (ECM) proteins providing a soft platelet plug. Because keratin biomaterials have structural and biochemical characteristics similar to ECM proteins, we utilized several adhesion assays to investigate platelet adhesion to keratin

Carbon Fibers from Chicken Feather Keratin

NASA Astrophysics Data System (ADS)

Miller, Melissa E.; Wool, Richard

2006-03-01

As the availability of synthetic and fossil-fuel based resources is becoming limited, bio-based materials offer an environmentally friendly alternative. Chicken feathers remain a huge agricultural waste. The feathers are comprised of approximately 97% keratin, but are currently used only to enrich animal feed. However, this usage is becoming a problem with the spread of diseases such as Bovine Spongiform Encephalopathy, commonly called ``Mad Cow Disease.'' The hollow, microcrystalline, oriented keratin feather fibers offer a novel, low cost approach to producing carbon fibers through controlled pyrolysis. Carbonized feather fibers (CFF) were prepared by first heating to 225 ^oC (below the melting point)in N2 for 26 hours to crosslink and stabilize the fiber structure; then carbonization occurred by increasing the temperature to 450 ^oC for two more hours. The resulting CFF were hollow, stiff and strong and had an affine 80% weight loss, which is near the theoretical value for the C-content of keratin. Initial studies showed that a composite with the CFF and an epoxidized soybean oil (AESO) gave an improved fiber modulus ECFF of order 13.5--66.1 GPa. With continued research, the goals are to increase the stiffness of the feathers to 100 GPa, while increasing the strength in the range of 5-10 GPa.

Keratins and lipids in ethnic hair.

PubMed

Cruz, C F; Fernandes, M M; Gomes, A C; Coderch, L; Martí, M; Méndez, S; Gales, L; Azoia, N G; Shimanovich, U; Cavaco-Paulo, A

2013-06-01

Human hair has an important and undeniable relevance in society due to its important role in visual appearance and social communication. Hair is mainly composed of structural proteins, mainly keratin and keratin associated proteins and lipids. Herein, we report a comprehensive study of the content and distribution of the lipids among ethnic hair, African, Asian and Caucasian hair. More interestingly, we also report the study of the interaction between those two main components of hair, specifically, the influence of the hair internal lipids in the structure of the hair keratin. This was achieved by the use of a complete set of analytical tools, such as thin layer chromatography-flame ionization detector, X-ray analysis, molecular dynamics simulation and confocal microscopy. The experimental results indicated different amounts of lipids on ethnic hair compositions and higher percentage of hair internal lipids in African hair. In this type of hair, the axial diffraction of keratin was not observed in X-ray analysis, but after hair lipids removal, the keratin returned to its typical packing arrangement. In molecular dynamic simulation, lipids were shown to intercalate dimers of keratin, changing its structure. From those results, we assume that keratin structure may be influenced by higher concentration of lipids in African hair. © 2013 Society of Cosmetic Scientists and the Société Française de Cosmétologie.

Isolation of a new class of cysteine–glycine–proline-rich beta-proteins (beta-keratins) and their expression in snake epidermis

PubMed Central

Dalla Valle, Luisa; Nardi, Alessia; Alibardi, Lorenzo

2010-01-01

Scales of snakes contain hard proteins (beta-keratins), now referred to as keratin-associated beta-proteins. In the present study we report the isolation, sequencing, and expression of a new group of these proteins from snake epidermis, designated cysteine–glycine–proline-rich proteins. One deduced protein from expressed mRNAs contains 128 amino acids (12.5 kDa) with a theoretical pI at 7.95, containing 10.2% cysteine and 15.6% glycine. The sequences of two more snake cysteine–proline-rich proteins have been identified from genomic DNA. In situ hybridization shows that the messengers for these proteins are present in the suprabasal and early differentiating beta-cells of the renewing scale epidermis. The present study shows that snake scales, as previously seen in scales of lizards, contain cysteine-rich beta-proteins in addition to glycine-rich beta-proteins. These keratin-associated beta-proteins mix with intermediate filament keratins (alpha-keratins) to produce the resistant corneous layer of snake scales. The specific proportion of these two subfamilies of proteins in different scales can determine various degrees of hardness in scales. PMID:20070430

Keratin 17 is overexpressed and predicts poor survival in estrogen receptor-negative/human epidermal growth factor receptor-2-negative breast cancer.

PubMed

Merkin, Ross D; Vanner, Elizabeth A; Romeiser, Jamie L; Shroyer, A Laurie W; Escobar-Hoyos, Luisa F; Li, Jinyu; Powers, Robert S; Burke, Stephanie; Shroyer, Kenneth R

2017-04-01

Clinicopathological features of breast cancer have limited accuracy to predict survival. By immunohistochemistry (IHC), keratin 17 (K17) expression has been correlated with triple-negative status (estrogen receptor [ER]/progesterone receptor/human epidermal growth factor receptor-2 [HER2] negative) and decreased survival, but K17 messenger RNA (mRNA) expression has not been evaluated in breast cancer. K17 is a potential prognostic cancer biomarker, targeting p27, and driving cell cycle progression. This study compared K17 protein and mRNA expression to ER/progesterone receptor/HER2 receptor status and event-free survival. K17 IHC was performed on 164 invasive breast cancers and K17 mRNA was evaluated in 1097 breast cancers. The mRNA status of other keratins (16/14/9) was evaluated in 113 ER - /HER2 - ductal carcinomas. IHC demonstrated intense cytoplasmic and membranous K17 localization in myoepithelial cells of benign ducts and lobules and tumor cells of ductal carcinoma in situ. In ductal carcinomas, K17 protein was detected in most triple-negative tumors (28/34, 82%), some non-triple-negative tumors (52/112, 46%), but never in lobular carcinomas (0/15). In ductal carcinomas, high K17 mRNA was associated with reduced 5-year event-free survival in advanced tumor stage (n = 149, hazard ratio [HR] = 3.68, P = .018), and large (n = 73, HR = 3.95, P = .047), triple-negative (n = 103, HR = 2.73, P = .073), and ER - /HER2 - (n = 113, HR = 2.99, P = .049) tumors. There were significant correlations among keratins 17, 16, 14, and 9 mRNA levels suggesting these keratins (all encoded on chromosome 17) could be coordinately expressed in breast cancer. Thus, K17 is expressed in a subset of triple-negative breast cancers, and is a marker of poor prognosis in patients with advanced stage and ER - /HER2 - breast cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

Fluorescence detection of protein content in house dust: the possible role of keratin.

PubMed

Voloshina, O V; Shirshin, E A; Lademann, J; Fadeev, V V; Darvin, M E

2017-03-01

We propose a fluorescence method for protein content assessment in fine house dust, which can be used as an indicator of the hygienic state of occupied rooms. The results of the measurements performed with 30 house dust samples, including ultrafiltration experiments, strongly suggest that the fluorescence emission of house dust extracts excited at 350 nm is mainly due to protein fragments, which are presumably keratin hydrolysates. This suggestion is supported by several facts: (i) Spectral band shapes for all the samples under investigation are close and correspond to that of keratin; (ii) fluorescence intensity correlates with the total protein content as provided by Lowry assay; (iii) treatment of the samples with proteinase K, which induces keratin hydrolysis, results in fluorescence enhancement without changing fluorescence band shape; and (iv) Raman spectra of keratin and fine house dust samples exhibit a very similar structure. Based on the obtained results and literature data, we propose a hypothesis that keratin is a major substrate for fluorescence species in fine house dust, which are responsible for emission at 350-nm excitation. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Keratin, luminal epithelial antigen and carcinoembryonic antigen in human urinary bladder carcinomas. An immunohistochemical study.

PubMed

Nathrath, W B; Arnholdt, H; Wilson, P D

1982-01-01

14 urinary bladder carcinomas of all main types were investigated with antisera to "broad spectrum keratin" (aK), "luminal epithelial antigen" (aLEA) and carcinoembryonic antigen (aCEA), using an indirect immunoperoxidase method on formalin fixed paraffin embedded sections. Keratin and LEA were both present in normal transitional epithelium, papilloma and carcinoma in situ whereas CEA was absent. Transitional cell carcinomas reacted with both aK and aLEA whereas CEA was seen only in a few foci. In squamous metaplasia and squamous carcinoma reaction with aK was particularly strong, while LEA was almost lacking and CEA was present in necrotic centres. In adenocarcinomas aK and aLEA reacted equally while aCEA reacted only on the surface.

Raman spectroscopic study of keratin 8 knockdown oral squamous cell carcinoma derived cells

NASA Astrophysics Data System (ADS)

Singh, S. P.; Alam, Hunain; Dmello, Crismita; Vaidya, Milind M.; Krishna, C. Murali

2012-03-01

Keratins are one of most widely used markers for oral cancers. Keratin 8 and 18 are expressed in simple epithelia and perform both mechanical and regulatory functions. Their expression are not seen in normal oral tissues but are often expressed in oral squamous cell carcinoma. Aberrant expression of keratins 8 and 18 is most common change in human oral cancer. Optical-spectroscopic methods are sensitive to biochemical changes and being projected as novel diagnostic tools for cancer diagnosis. Aim of this study was to evaluate potentials of Raman spectroscopy in detecting minor changes associated with differential level of keratin expression in tongue-cancer-derived AW13516 cells. Knockdown clones for K8 were generated and synchronized by growing under serum-free conditions. Cell pellets of three independent experiments in duplicate were used for recording Raman spectra with fiberoptic-probe coupled HE-785 Raman-instrument. A total of 123 and 96 spectra from knockdown clones and vector controls respectively in 1200-1800 cm-1 region were successfully utilized for classification using LDA. Two separate clusters with classification-efficiency of ~95% were obtained. Leave-one-out cross-validation yielded ~63% efficiency. Findings of the study demonstrate the potentials of Raman spectroscopy in detecting even subtle changes such as variations in keratin expression levels. Future studies towards identifying Raman signals from keratin in oral cells can help in precise cancer diagnosis.

Epithelial Keratins Modulate cMet Expression and Signaling and Promote InlB-Mediated Listeria monocytogenes Infection of HeLa Cells.

PubMed

Cruz, Rui; Pereira-Castro, Isabel; Almeida, Maria T; Moreira, Alexandra; Cabanes, Didier; Sousa, Sandra

2018-01-01

The host cytoskeleton is a major target for bacterial pathogens during infection. In particular, pathogens usurp the actin cytoskeleton function to strongly adhere to the host cell surface, to induce plasma membrane remodeling allowing invasion and to spread from cell to cell and disseminate to the whole organism. Keratins are cytoskeletal proteins that are the major components of intermediate filaments in epithelial cells however, their role in bacterial infection has been disregarded. Here we investigate the role of the major epithelial keratins, keratins 8 and 18 (K8 and K18), in the cellular infection by Listeria monocytogenes . We found that K8 and K18 are required for successful InlB/cMet-dependent L. monocytogenes infection, but are dispensable for InlA/E-cadherin-mediated invasion. Both K8 and K18 accumulate at InlB-mediated internalization sites following actin recruitment and modulate actin dynamics at those sites. We also reveal the key role of K8 and K18 in HGF-induced signaling which occurs downstream the activation of cMet. Strikingly, we show here that K18, and at a less extent K8, controls the expression of cMet and other surface receptors such TfR and integrin β1, by promoting the stability of their corresponding transcripts. Together, our results reveal novel functions for major epithelial keratins in the modulation of actin dynamics at the bacterial entry sites and in the control of surface receptors mRNA stability and expression.

Microarray analysis identifies keratin loci as sensitive biomarkers for thyroid hormone disruption in the salamander Ambystoma mexicanum.

PubMed

Page, Robert B; Monaghan, James R; Samuels, Amy K; Smith, Jeramiah J; Beachy, Christopher K; Voss, S Randal

2007-02-01

Ambystomatid salamanders offer several advantages for endocrine disruption research, including genomic and bioinformatics resources, an accessible laboratory model (Ambystoma mexicanum), and natural lineages that are broadly distributed among North American habitats. We used microarray analysis to measure the relative abundance of transcripts isolated from A. mexicanum epidermis (skin) after exogenous application of thyroid hormone (TH). Only one gene had a >2-fold change in transcript abundance after 2 days of TH treatment. However, hundreds of genes showed significantly different transcript levels at days 12 and 28 in comparison to day 0. A list of 123 TH-responsive genes was identified using statistical, BLAST, and fold level criteria. Cluster analysis identified two groups of genes with similar transcription patterns: up-regulated versus down-regulated. Most notably, several keratins exhibited dramatic (1000 fold) increases or decreases in transcript abundance. Keratin gene expression changes coincided with morphological remodeling of epithelial tissues. This suggests that keratin loci can be developed as sensitive biomarkers to assay temporal disruptions of larval-to-adult gene expression programs. Our study has identified the first collection of loci that are regulated during TH-induced metamorphosis in a salamander, thus setting the stage for future investigations of TH disruption in the Mexican axolotl and other salamanders of the genus Ambystoma.

Degradation and regeneration of feather keratin in NMMO solution.

PubMed

Ma, Bomou; Sun, Qisong; Yang, Jing; Wizi, Jakpa; Hou, Xiuliang; Yang, Yiqi

2017-07-01

Chicken feather, a potential source of keratin, is often disposed as waste material. Although some methods, i.e., hydrolysis, reduction, and oxidation, have been developed to isolate keratin for composites, it has been limited due to the rising environmental concerns. In this work, a green solvent N-methylmorpholine N-oxide (NMMO) was used to extract keratin from chicken feather waste. Eighty-nine percent of keratin was extracted using 75% NMMO solution. However, the result from size exclusion HPLC showed that most of the keratin degraded into polypeptide with molecular weight of 2189 and only 25.3% regenerated keratin was obtained with molecular weight of 14,485. Analysis of amino acid composition showed a severe damage to the disulfide bonds in keratin during the extraction procedure. Oxidization had an important effect on the reconstitution of the disulfide bonds, which formed a stable three-dimensional net structure in the regenerated keratins. Besides, Raman spectra, NMR, FT-IR, XRD, and TGA were used to characterize the properties of regenerated keratin and raw chicken feather. In the end, a possible mechanism was proposed based on the results.

Genetic background effects of keratin 8 and 18 in a DDC-induced hepatotoxicity and Mallory-Denk body formation mouse model.

PubMed

Haybaeck, Johannes; Stumptner, Cornelia; Thueringer, Andrea; Kolbe, Thomas; Magin, Thomas M; Hesse, Michael; Fickert, Peter; Tsybrovskyy, Oleksiy; Müller, Heimo; Trauner, Michael; Zatloukal, Kurt; Denk, Helmut

2012-06-01

Keratin 8 (K8) and keratin 18 (K18) form the major hepatocyte cytoskeleton. We investigated the impact of genetic loss of either K8 or K18 on liver homeostasis under toxic stress with the hypothesis that K8 and K18 exert different functions. krt8⁻/⁻ and krt18⁻/⁻ mice crossed into the same 129-ola genetic background were treated by acute and chronic administration of 3,5-diethoxy-carbonyl-1,4-dihydrocollidine (DDC). In acutely DDC-intoxicated mice, macrovesicular steatosis was more pronounced in krt8⁻/⁻ and krt18⁻/⁻ compared with wild-type (wt) animals. Mallory-Denk bodies (MDBs) appeared in krt18⁻/⁻ mice already at an early stage of intoxication in contrast to krt8⁻/⁻ mice that did not display MDB formation when fed with DDC. Keratin-deficient mice displayed significantly lower numbers of apoptotic hepatocytes than wt animals. krt8⁻/⁻, krt18⁻/⁻ and control mice displayed comparable cell proliferation rates. Chronically DDC-intoxicated krt18⁻/⁻ and wt mice showed a similarly increased degree of steatohepatitis with hepatocyte ballooning and MDB formation. In krt8⁻/⁻ mice, steatosis was less, ballooning, and MDBs were absent. krt18⁻/⁻ mice developed MDBs whereas krt8⁻/⁻ mice on the same genetic background did not, highlighting the significance of different structural properties of keratins. They are independent of the genetic background as an intrinsic factor. By contrast, toxicity effects may depend on the genetic background. krt8⁻/⁻ and krt18⁻/⁻ mice on the same genetic background show similar sensitivity to DDC intoxication and almost resemble wt animals regarding survival, degree of porphyria, liver-to-body weight ratio, serum bilirubin and liver enzyme levels. This stands in contrast to previous work where krt8⁻/⁻ and krt18⁻/⁻ mice on different genetic backgrounds were investigated.

Keratin 8 and 18 Loss in Epithelial Cancer Cells Increases Collective Cell Migration and Cisplatin Sensitivity through Claudin1 Up-regulation*

PubMed Central

Fortier, Anne-Marie; Asselin, Eric; Cadrin, Monique

2013-01-01

Keratins 8 and 18 (K8/18) are simple epithelial cell-specific intermediate filament proteins. Keratins are essential for tissue integrity and are involved in intracellular signaling pathways that regulate cell response to injuries, cell growth, and death. K8/18 expression is maintained during tumorigenesis; hence, they are used as a diagnostic marker in tumor pathology. In recent years, studies have provided evidence that keratins should be considered not only as markers but also as regulators of cancer cell signaling. The loss of K8/18 expression during epithelial-mesenchymal transition (EMT) is associated with metastasis and chemoresistance. In the present study, we investigated whether K8/18 expression plays an active role in EMT. We show that K8/18 stable knockdown using shRNA increased collective migration and invasiveness of epithelial cancer cells without modulating EMT markers. K8/18-depleted cells showed PI3K/Akt/NF-κB hyperactivation and increased MMP2 and MMP9 expression. K8/18 deletion also increased cisplatin-induced apoptosis. Increased Fas receptor membrane targeting suggests that apoptosis is enhanced via the extrinsic pathway. Interestingly, we identified the tight junction protein claudin1 as a regulator of these processes. This is the first indication that modulation of K8/18 expression can influence the phenotype of epithelial cancer cells at a transcriptional level and supports the hypothesis that keratins play an active role in cancer progression. PMID:23449973

Biodegradable materials based on silk fibroin and keratin.

PubMed

Vasconcelos, Andreia; Freddi, Giuliano; Cavaco-Paulo, Artur

2008-04-01

Wool and silk were dissolved and used for the preparation of blended films. Two systems are proposed: (1) blend films of silk fibroin and keratin aqueous solutions and (2) silk fibroin and keratin dissolved in formic acid. The FTIR spectra of pure films cast from aqueous solutions indicated that the keratin secondary structure mainly consists of alpha-helix and random coil conformations. The IR spectrum of pure SF is characteristic of films with prevalently amorphous structure (random coil conformation). Pure keratin film cast from formic acid shows an increase in the amount of beta-sheet and disordered keratin structures. The FTIR pattern of SF dissolved in formic acid is characteristic of films with prevalently beta-sheet conformations with beta-sheet crystallites embedded in an amorphous matrix. The thermal behavior of the blends confirmed the FTIR results. DSC curve of pure SF is typical of amorphous SF and the curve of pure keratin show the characteristic melting peak of alpha-helices for the aqueous system. These patterns are no longer observed in the films cast from formic acid due to the ability of formic acid to induce crystallization of SF and to increase the amount of beta-sheet structures on keratin. The nonlinear trend of the different parameters obtained from FTIR analysis and DSC curves of both SF/keratin systems indicate that when proteins are mixed they do not follow additives rules but are able to establish intermolecular interactions. Degradable polymeric biomaterials are preferred candidates for medical applications. It was investigated the degradation behavior of both SF/keratin systems by in vitro enzymatic incubation with trypsin. The SF/keratin films cast from water underwent a slower biological degradation than the films cast from formic acid. The weight loss obtained is a function of the amount of keratin in the blend. This study encourages the further investigation of the type of matrices presented here to be applied whether in scaffolds

Keratin 1 maintains skin integrity and participates in an inflammatory network in skin through interleukin-18.

PubMed

Roth, Wera; Kumar, Vinod; Beer, Hans-Dietmar; Richter, Miriam; Wohlenberg, Claudia; Reuter, Ursula; Thiering, Sören; Staratschek-Jox, Andrea; Hofmann, Andrea; Kreusch, Fatima; Schultze, Joachim L; Vogl, Thomas; Roth, Johannes; Reichelt, Julia; Hausser, Ingrid; Magin, Thomas M

2012-11-15

Keratin 1 (KRT1) and its heterodimer partner keratin 10 (KRT10) are major constituents of the intermediate filament cytoskeleton in suprabasal epidermis. KRT1 mutations cause epidermolytic ichthyosis in humans, characterized by loss of barrier integrity and recurrent erythema. In search of the largely unknown pathomechanisms and the role of keratins in barrier formation and inflammation control, we show here that Krt1 is crucial for maintenance of skin integrity and participates in an inflammatory network in murine keratinocytes. Absence of Krt1 caused a prenatal increase in interleukin-18 (IL-18) and the S100A8 and S100A9 proteins, accompanied by a barrier defect and perinatal lethality. Depletion of IL-18 partially rescued Krt1(-/-) mice. IL-18 release was keratinocyte-autonomous, KRT1 and caspase-1 dependent, supporting an upstream role of KRT1 in the pathology. Finally, transcriptome profiling revealed a Krt1-mediated gene expression signature similar to atopic eczema and psoriasis, but different from Krt5 deficiency and epidermolysis bullosa simplex. Our data suggest a functional link between KRT1 and human inflammatory skin diseases.

Keratin 17 modulates hair follicle cycling in a TNFα-dependent fashion

PubMed Central

Tong, Xuemei; Coulombe, Pierre A.

2006-01-01

Mammalian hair follicles cycle between stages of rapid growth (anagen) and metabolic quiescence (telogen) throughout life. Transition from anagen to telogen involves an intermediate stage, catagen, consisting of a swift, apoptosis-driven involution of the lower half of the follicle. How catagen is coordinated, and spares the progenitor cells needed for anagen re-entry, is poorly understood. Keratin 17 (K17)-null mice develop alopecia in the first week post-birth, correlating with hair shaft fragility and untimely apoptosis in the hair bulb. Here we show that this abnormal apoptosis reflects premature entry into catagen. Of the proapoptotic challenges tested, K17-null skin keratinocytes in primary culture are selectively more sensitive to TNFα. K17 interacts with TNF receptor 1 (TNFR1)-associated death domain protein (TRADD), a death adaptor essential for TNFR1-dependent signal relay, suggesting a functional link between this keratin and TNFα signaling. The activity of NF-κB, a downstream target of TNFα, is increased in K17-null skin. We also find that TNFα is required for a timely anagen–catagen transition in mouse pelage follicles, and that its ablation partially rescues the hair cycling defect of K17-null mice. These findings identify K17 and TNFα as two novel and interdependent regulators of hair cycling. PMID:16702408

Progress towards genetic and pharmacological therapies for keratin genodermatoses: current perspective and future promise

PubMed Central

Chamcheu, Jean Christopher; Wood, Gary S.; Siddiqui, Imtiaz A.; Syed, Deeba N.; Adhami, Vaqar M.; Teng, Joyce M.; Mukhtar, Hasan

2012-01-01

Hereditary keratin disorders of the skin and its appendages comprise a large group of clinically heterogeneous disfiguring blistering and ichthyotic diseases, primarily characterized by the loss of tissue integrity, blistering and hyperkeratosis in severely affected tissues. Pathogenic mutations in keratins cause these afflictions. Typically, these mutations in concert with characteristic features have formed the basis for improved disease diagnosis, prognosis and most recently therapy development. Examples include epidermolysis bullosa simplex, keratinopathic ichthyosis, pachyonychia congenita and several other tissue-specific hereditary keratinopathies. Understanding the molecular and genetic events underlying skin dysfunction has initiated alternative treatment approaches that may provide novel therapeutic opportunities for affected patients. Animal and in vitro disease modelling studies have shed more light on molecular pathogenesis, further defining the role of keratins in disease processes and promoting the translational development of new gene and pharmacological therapeutic strategies. Given that the molecular basis for these monogenic disorders is well established, gene therapy and drug discovery targeting pharmacological compounds with the ability to reinforce the compromised cytoskeleton may lead to promising new therapeutic strategies for treating hereditary keratinopathies. In this review, we will summarize and discuss recent advances in the preclinical and clinical modelling and development of gene, natural product, pharmacological and protein-based therapies for these disorders, highlighting the feasibility of new approaches for translational clinical therapy. PMID:22716242

Progress towards genetic and pharmacological therapies for keratin genodermatoses: current perspective and future promise.

PubMed

Chamcheu, Jean Christopher; Wood, Gary S; Siddiqui, Imtiaz A; Syed, Deeba N; Adhami, Vaqar M; Teng, Joyce M; Mukhtar, Hasan

2012-07-01

Hereditary keratin disorders of the skin and its appendages comprise a large group of clinically heterogeneous disfiguring blistering and ichthyotic diseases, primarily characterized by the loss of tissue integrity, blistering and hyperkeratosis in severely affected tissues. Pathogenic mutations in keratins cause these afflictions. Typically, these mutations in concert with characteristic features have formed the basis for improved disease diagnosis, prognosis and most recently therapy development. Examples include epidermolysis bullosa simplex, keratinopathic ichthyosis, pachyonychia congenita and several other tissue-specific hereditary keratinopathies. Understanding the molecular and genetic events underlying skin dysfunction has initiated alternative treatment approaches that may provide novel therapeutic opportunities for affected patients. Animal and in vitro disease modelling studies have shed more light on molecular pathogenesis, further defining the role of keratins in disease processes and promoting the translational development of new gene and pharmacological therapeutic strategies. Given that the molecular basis for these monogenic disorders is well established, gene therapy and drug discovery targeting pharmacological compounds with the ability to reinforce the compromised cytoskeleton may lead to promising new therapeutic strategies for treating hereditary keratinopathies. In this review, we will summarize and discuss recent advances in the preclinical and clinical modelling and development of gene, natural product, pharmacological and protein-based therapies for these disorders, highlighting the feasibility of new approaches for translational clinical therapy. © 2012 John Wiley & Sons A/S.

The Chicken Frizzle Feather Is Due to an α-Keratin (KRT75) Mutation That Causes a Defective Rachis

PubMed Central

Foley, John; Foley, Anne; McDonald, Merry-Lynn; Juan, Wen-Tau; Huang, Chih-Jen; Lai, Yu-Ting; Lo, Wen-Sui; Chen, Chih-Feng; Leal, Suzanne M.; Zhang, Huanmin; Widelitz, Randall B.; Patel, Pragna I.; Li, Wen-Hsiung; Chuong, Cheng-Ming

2012-01-01

Feathers have complex forms and are an excellent model to study the development and evolution of morphologies. Existing chicken feather mutants are especially useful for identifying genetic determinants of feather formation. This study focused on the gene F, underlying the frizzle feather trait that has a characteristic curled feather rachis and barbs in domestic chickens. Our developmental biology studies identified defects in feather medulla formation, and physical studies revealed that the frizzle feather curls in a stepwise manner. The frizzle gene is transmitted in an autosomal incomplete dominant mode. A whole-genome linkage scan of five pedigrees with 2678 SNPs revealed association of the frizzle locus with a keratin gene-enriched region within the linkage group E22C19W28_E50C23. Sequence analyses of the keratin gene cluster identified a 69 bp in-frame deletion in a conserved region of KRT75, an α-keratin gene. Retroviral-mediated expression of the mutated F cDNA in the wild-type rectrix qualitatively changed the bending of the rachis with some features of frizzle feathers including irregular kinks, severe bending near their distal ends, and substantially higher variations among samples in comparison to normal feathers. These results confirmed KRT75 as the F gene. This study demonstrates the potential of our approach for identifying genetic determinants of feather forms. PMID:22829773

Genome-wide profiling identifies a subset of methamphetamine (METH)-induced genes associated with METH-induced increased H4K5Ac binding in the rat striatum.

PubMed

Cadet, Jean Lud; Jayanthi, Subramaniam; McCoy, Michael T; Ladenheim, Bruce; Saint-Preux, Fabienne; Lehrmann, Elin; De, Supriyo; Becker, Kevin G; Brannock, Christie

2013-08-12

METH is an illicit drug of abuse that influences gene expression in the rat striatum. Histone modifications regulate gene transcription. We therefore used microarray analysis and genome-scale approaches to examine potential relationships between the effects of METH on gene expression and on DNA binding of histone H4 acetylated at lysine 4 (H4K5Ac) in the rat dorsal striatum of METH-naïve and METH-pretreated rats. Acute and chronic METH administration caused differential changes in striatal gene expression. METH also increased H4K5Ac binding around the transcriptional start sites (TSSs) of genes in the rat striatum. In order to relate gene expression to histone acetylation, we binned genes of similar expression into groups of 100 genes and proceeded to relate gene expression to H4K5Ac binding. We found a positive correlation between gene expression and H4K5Ac binding in the striatum of control rats. Similar correlations were observed in METH-treated rats. Genes that showed acute METH-induced increased expression in saline-pretreated rats also showed METH-induced increased H4K5Ac binding. The acute METH injection caused similar increases in H4K5Ac binding in METH-pretreated rats, without affecting gene expression to the same degree. Finally, genes that showed METH-induced decreased expression exhibited either decreases or no changes in H4K5Ac binding. Acute METH injections caused increased gene expression of genes that showed increased H4K5Ac binding near their transcription start sites.

The use of isoelectric focusing to identify rhinoceros keratins.

PubMed

Butler, D J; De Forest, P R; Kobilinsky, L

1990-03-01

Keratins represent the principal structural proteins of hair. They are also found in horn, nail, claw, hoof, and feather. Hair and nail samples from human and canine sources and hair samples from mule deer, white tail deer, cat, moose, elk, antelope, caribou, raccoon, and goat were studied. Parrot and goose feathers were also analyzed. Keratins are polymorphic, and species differences are known to exist. Proteinaceous extracts of deer and antelope antlers and bovine and rhinoceros horn were prepared by solubilizing 10 mg of horn sample in 200 microL of a solution containing 12M urea, 74mM Trizma base, and 78mM dithiothreitol (DTT). Extraction took place over a 48-h period. A 25-microL aliquot of extract was removed and incubated with 5 microL of 0.1 M DTT for 10 min at 25 degrees C. Keratins were then separated by isoelectric focusing (IEF) on 5.2% polyacrylamide gels for 3 h and visualized using silver staining. At least 20 bands could be observed for each species studied. However, band patterns differed in the position of each band, in the number of bands, and in band coloration resulting from the silver staining process. Horn from two species of rhinoceros was examined. For both specimens, most bands occurred in the pH range of 4 to 5. Although similar patterns for both species were observed, they differed sufficiently to differentiate one from the other. As might be expected, the closer two species are related phylogenetically, the greater the similarity in the IEF pattern produced from their solubilized keratin.(ABSTRACT TRUNCATED AT 250 WORDS)

Skn-1a/Oct-11 and {Delta}Np63{alpha} exert antagonizing effects on human keratin expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lena, Anna Maria; Cipollone, Rita; Amelio, Ivano

2010-10-29

Research highlights: {yields} Skn-1a markedly downregulates {Delta}Np63-driven K14 expression. {yields} {Delta}Np63 inhibits Skn-1a-mediated K10 expression. {yields} {Delta}Np63, mutated in SAM domain, is less effecting in K10 downregulation. {yields} Immunolocalization in human skin of the two transcription factors is partially overlapping. {yields} The antagonistic effects of Skn-1a and p63 is through competition for overlapping responsive elements or through an indirect interaction. -- Abstract: The formation of a stratified epidermis requires a carefully controlled balance between keratinocyte proliferation and differentiation. Here, we report the reciprocal effect on keratin expression of {Delta}Np63, pivotal in normal epidermal morphogenesis and maintenance, and Skn-1a/Oct-11, a POUmore » transcription factor that triggers and regulates the differentiation of keratinocytes. The expression of Skn-1a markedly downregulated {Delta}Np63-driven K14 expression in luciferase reporter assays. The extent of downregulation was comparable to the inhibition of Skn-1a-mediated K10 expression upon expression of {Delta}Np63. {Delta}Np63, mutated in the protein-protein interaction domain (SAM domain; mutated in human ectodermal dysplasia syndrome), was significantly less effecting in downregulating K10, raising the possibility of a direct interaction among Skn-1a and {Delta}Np63. Immunolocalization in human skin biopsies revealed that the expression of the two transcription factors is partially overlapping. Co-immunoprecipitation experiments did not, however, demonstrate a direct interaction between {Delta}Np63 and Skn-1a, suggesting that the antagonistic effects of Skn-1a and p63 on keratin promoter transactivation is probably through competition for overlapping binding sites on target gene promoter or through an indirect interaction.« less

Genome-wide profiling identifies a subset of methamphetamine (METH)-induced genes associated with METH-induced increased H4K5Ac binding in the rat striatum

PubMed Central

2013-01-01

Background METH is an illicit drug of abuse that influences gene expression in the rat striatum. Histone modifications regulate gene transcription. Methods We therefore used microarray analysis and genome-scale approaches to examine potential relationships between the effects of METH on gene expression and on DNA binding of histone H4 acetylated at lysine 4 (H4K5Ac) in the rat dorsal striatum of METH-naïve and METH-pretreated rats. Results Acute and chronic METH administration caused differential changes in striatal gene expression. METH also increased H4K5Ac binding around the transcriptional start sites (TSSs) of genes in the rat striatum. In order to relate gene expression to histone acetylation, we binned genes of similar expression into groups of 100 genes and proceeded to relate gene expression to H4K5Ac binding. We found a positive correlation between gene expression and H4K5Ac binding in the striatum of control rats. Similar correlations were observed in METH-treated rats. Genes that showed acute METH-induced increased expression in saline-pretreated rats also showed METH-induced increased H4K5Ac binding. The acute METH injection caused similar increases in H4K5Ac binding in METH-pretreated rats, without affecting gene expression to the same degree. Finally, genes that showed METH-induced decreased expression exhibited either decreases or no changes in H4K5Ac binding. Conclusion Acute METH injections caused increased gene expression of genes that showed increased H4K5Ac binding near their transcription start sites. PMID:23937714

Keratin impact on PKCδ- and ASMase-mediated regulation of hepatocyte lipid raft size – implication for FasR-associated apoptosis

PubMed Central

Gilbert, Stéphane; Loranger, Anne; Omary, M. Bishr

2016-01-01

ABSTRACT Keratins are epithelial cell intermediate filament (IF) proteins that are expressed as pairs in a cell-differentiation-regulated manner. Hepatocytes express the keratin 8 and 18 pair (denoted K8/K18) of IFs, and a loss of K8 or K18, as in K8-null mice, leads to degradation of the keratin partner. We have previously reported that a K8/K18 loss in hepatocytes leads to altered cell surface lipid raft distribution and more efficient Fas receptor (FasR, also known as TNFRSF6)-mediated apoptosis. We demonstrate here that the absence of K8 or transgenic expression of the K8 G62C mutant in mouse hepatocytes reduces lipid raft size. Mechanistically, we find that the lipid raft size is dependent on acid sphingomyelinase (ASMase, also known as SMPD1) enzyme activity, which is reduced in absence of K8/K18. Notably, the reduction of ASMase activity appears to be caused by a less efficient redistribution of surface membrane PKCδ toward lysosomes. Moreover, we delineate the lipid raft volume range that is required for an optimal FasR-mediated apoptosis. Hence, K8/K18-dependent PKCδ- and ASMase-mediated modulation of lipid raft size can explain the more prominent FasR-mediated signaling resulting from K8/K18 loss. The fine-tuning of ASMase-mediated regulation of lipid rafts might provide a therapeutic target for death-receptor-related liver diseases. PMID:27422101

Molecular evolution of the keratin associated protein gene family in mammals, role in the evolution of mammalian hair.

PubMed

Wu, Dong-Dong; Irwin, David M; Zhang, Ya-Ping

2008-08-23

Hair is unique to mammals. Keratin associated proteins (KRTAPs), which contain two major groups: high/ultrahigh cysteine and high glycine-tyrosine, are one of the major components of hair and play essential roles in the formation of rigid and resistant hair shafts. The KRTAP family was identified as being unique to mammals, and near-complete KRTAP gene repertoires for eight mammalian genomes were characterized in this study. An expanded KRTAP gene repertoire was found in rodents. Surprisingly, humans have a similar number of genes as other primates despite the relative hairlessness of humans. We identified several new subfamilies not previously reported in the high/ultrahigh cysteine KRTAP genes. Genes in many subfamilies of the high/ultrahigh cysteine KRTAP genes have evolved by concerted evolution with frequent gene conversion events, yielding a higher GC base content for these gene sequences. In contrast, the high glycine-tyrosine KRTAP genes have evolved more dynamically, with fewer gene conversion events and thus have a lower GC base content, possibly due to positive selection. Most of the subfamilies emerged early in the evolution of mammals, thus we propose that the mammalian ancestor should have a diverse KRTAP gene repertoire. We propose that hair content characteristics have evolved and diverged rapidly among mammals because of rapid divergent evolution of KRTAPs between species. In contrast, subfamilies of KRTAP genes have been homogenized within each species due to concerted evolution.

Synthesis of Keratin-based Nanofiber for Biomedical Engineering.

PubMed

Thompson, Zanshe S; Rijal, Nava P; Jarvis, David; Edwards, Angela; Bhattarai, Narayan

2016-02-07

Electrospinning, due to its versatility and potential for applications in various fields, is being frequently used to fabricate nanofibers. Production of these porous nanofibers is of great interest due to their unique physiochemical properties. Here we elaborate on the fabrication of keratin containing poly (ε-caprolactone) (PCL) nanofibers (i.e., PCL/keratin composite fiber). Water soluble keratin was first extracted from human hair and mixed with PCL in different ratios. The blended solution of PCL/keratin was transformed into nanofibrous membranes using a laboratory designed electrospinning set up. Fiber morphology and mechanical properties of the obtained nanofiber were observed and measured using scanning electron microscopy and tensile tester. Furthermore, degradability and chemical properties of the nanofiber were studied by FTIR. SEM images showed uniform surface morphology for PCL/keratin fibers of different compositions. These PCL/keratin fibers also showed excellent mechanical properties such as Young's modulus and failure point. Fibroblast cells were able to attach and proliferate thus proving good cell viability. Based on the characteristics discussed above, we can strongly argue that the blended nanofibers of natural and synthetic polymers can represent an excellent development of composite materials that can be used for different biomedical applications.

Flow behavior of regenerated wool-keratin proteins in different mediums.

PubMed

Alemdar, Ayse; Iridag, Yesim; Kazanci, Murat

2005-04-01

Keratin is abundantly present in nature and the major component of hair, wool, feather, nail and horns. Dissolution of keratin is often required when non-textile applications are demanded. However, the low solubility of keratin in water is the major problem. It becomes unstable and precipitated when stored for a long time. Therefore, it is necessary to find a good solvent that provides high stability and easy processibility. In this research, we used formic acid and dimethylformamide (DMF) to dissolve regenerated keratin protein films. It is shown that formic acid is a good solvent for regenerated keratin proteins for the purpose of storage. Transparent and stable regenerated keratin solution is obtained in formic acid.

Keratin impact on PKCδ- and ASMase-mediated regulation of hepatocyte lipid raft size - implication for FasR-associated apoptosis.

PubMed

Gilbert, Stéphane; Loranger, Anne; Omary, M Bishr; Marceau, Normand

2016-09-01

Keratins are epithelial cell intermediate filament (IF) proteins that are expressed as pairs in a cell-differentiation-regulated manner. Hepatocytes express the keratin 8 and 18 pair (denoted K8/K18) of IFs, and a loss of K8 or K18, as in K8-null mice, leads to degradation of the keratin partner. We have previously reported that a K8/K18 loss in hepatocytes leads to altered cell surface lipid raft distribution and more efficient Fas receptor (FasR, also known as TNFRSF6)-mediated apoptosis. We demonstrate here that the absence of K8 or transgenic expression of the K8 G62C mutant in mouse hepatocytes reduces lipid raft size. Mechanistically, we find that the lipid raft size is dependent on acid sphingomyelinase (ASMase, also known as SMPD1) enzyme activity, which is reduced in absence of K8/K18. Notably, the reduction of ASMase activity appears to be caused by a less efficient redistribution of surface membrane PKCδ toward lysosomes. Moreover, we delineate the lipid raft volume range that is required for an optimal FasR-mediated apoptosis. Hence, K8/K18-dependent PKCδ- and ASMase-mediated modulation of lipid raft size can explain the more prominent FasR-mediated signaling resulting from K8/K18 loss. The fine-tuning of ASMase-mediated regulation of lipid rafts might provide a therapeutic target for death-receptor-related liver diseases. © 2016. Published by The Company of Biologists Ltd.

FAM83H and casein kinase I regulate the organization of the keratin cytoskeleton and formation of desmosomes

PubMed Central

Kuga, Takahisa; Sasaki, Mitsuho; Mikami, Toshinari; Miake, Yasuo; Adachi, Jun; Shimizu, Maiko; Saito, Youhei; Koura, Minako; Takeda, Yasunori; Matsuda, Junichiro; Tomonaga, Takeshi; Nakayama, Yuji

2016-01-01

FAM83H is essential for the formation of dental enamel because a mutation in the FAM83H gene causes amelogenesis imperfecta (AI). We previously reported that the overexpression of FAM83H often occurs and disorganizes the keratin cytoskeleton in colorectal cancer cells. We herein show that FAM83H regulates the organization of the keratin cytoskeleton and maintains the formation of desmosomes in ameloblastoma cells. FAM83H is expressed and localized on keratin filaments in human ameloblastoma cell lines and in mouse ameloblasts and epidermal germinative cells in vivo. FAM83H shows preferential localization to keratin filaments around the nucleus that often extend to cell-cell junctions. Alterations in the function of FAM83H by its overexpression, knockdown, or an AI-causing truncated mutant prevent the proper organization of the keratin cytoskeleton in ameloblastoma cells. Furthermore, the AI-causing mutant prevents desmosomal proteins from being localized to cell-cell junctions. The effects of the AI-causing mutant depend on its binding to and possible inhibition of casein kinase I (CK-1). The suppression of CK-1 by its inhibitor, D4476, disorganizes the keratin cytoskeleton. Our results suggest that AI caused by the FAM83H mutation is mediated by the disorganization of the keratin cytoskeleton and subsequent disruption of desmosomes in ameloblasts. PMID:27222304

FAM83H and casein kinase I regulate the organization of the keratin cytoskeleton and formation of desmosomes.

PubMed

Kuga, Takahisa; Sasaki, Mitsuho; Mikami, Toshinari; Miake, Yasuo; Adachi, Jun; Shimizu, Maiko; Saito, Youhei; Koura, Minako; Takeda, Yasunori; Matsuda, Junichiro; Tomonaga, Takeshi; Nakayama, Yuji

2016-05-25

FAM83H is essential for the formation of dental enamel because a mutation in the FAM83H gene causes amelogenesis imperfecta (AI). We previously reported that the overexpression of FAM83H often occurs and disorganizes the keratin cytoskeleton in colorectal cancer cells. We herein show that FAM83H regulates the organization of the keratin cytoskeleton and maintains the formation of desmosomes in ameloblastoma cells. FAM83H is expressed and localized on keratin filaments in human ameloblastoma cell lines and in mouse ameloblasts and epidermal germinative cells in vivo. FAM83H shows preferential localization to keratin filaments around the nucleus that often extend to cell-cell junctions. Alterations in the function of FAM83H by its overexpression, knockdown, or an AI-causing truncated mutant prevent the proper organization of the keratin cytoskeleton in ameloblastoma cells. Furthermore, the AI-causing mutant prevents desmosomal proteins from being localized to cell-cell junctions. The effects of the AI-causing mutant depend on its binding to and possible inhibition of casein kinase I (CK-1). The suppression of CK-1 by its inhibitor, D4476, disorganizes the keratin cytoskeleton. Our results suggest that AI caused by the FAM83H mutation is mediated by the disorganization of the keratin cytoskeleton and subsequent disruption of desmosomes in ameloblasts.

Proteome analysis reveals that de novo regenerated mucosa over fibula flap-reconstructed mandibles resembles mature keratinized oral mucosa.

PubMed

Kumar, Vinay V; James, Bonney L; Ruß, Manuela; Mikkat, Stefan; Suresh, Amritha; Kämmerer, Peer W; Glocker, Michael O

2018-03-01

The aim of this study was to determine whether intra-oral de novo regenerated mucosa (D) that grew over free fibula flap reconstructed-mandibles resembled the donor tissue i.e. external skin (S) of the lateral leg, or the recipient site tissue, i.e. keratinized oral mucosa (K). Differential proteome analysis was performed with ten tissue samples from each of the three groups: de novo regenerated mucosa (D), external skin (S), and keratinized oral mucosa (K). Expression differences of cornulin and involucrin were validated by Western blot analysis and their spatial distributions in the respective tissues were ascertained by immunohistochemistry. From all three investigated tissue types a total of 1188 proteins were identified, 930 of which were reproducibly and robustly quantified by proteome analysis. The best differentiating proteins were assembled in an oral mucosa proteome signature that encompasses 56 differentially expressed proteins. Principal component analysis of both, the 930 quantifiable proteins and the 56 oral mucosa signature proteins revealed that the de novo regenerated mucosa resembles keratinized oral mucosa much closer than extra-oral skin. Differentially expressed cornification-related proteins comprise proteins from all subclasses of the cornified cell envelope. Prominently expressed in intra-oral mucosa tissues were (i) cornifin-A, cornifin-B, SPRR3, and involucrin from the cornified-cell-envelope precursor group, (ii) S100A9, S100A8 and S100A2 from the S100 group, and (iii) cornulin which belongs to the fused-gene-protein group. According to its proteome signature de novo regenerated mucosa over the free fibula flap not only presents a passive structural surface layer but has adopted active tissue function. Copyright © 2018 Elsevier Ltd. All rights reserved.

Rejoining of cut wounds by engineered gelatin-keratin glue.

PubMed

Thirupathi Kumara Raja, S; Thiruselvi, T; Sailakshmi, G; Ganesh, S; Gnanamani, A

2013-08-01

Rejoining of cut tissue ends of a critical site challenges clinicians. The toxicity, antigenicity, low adhesive strength, flexibility, swelling and cost of the currently employed glue demands an alternative. Engineered gelatin-keratin glue (EGK-glue) described in the present study was found to be suitable for wet tissue approximation. EGK-glue was prepared by engineering gelatin with caffeic acid using EDC and conjugating with keratin by periodate oxidation. UV-visible, (1)H NMR and circular dichroism analyses followed by experiments on gelation time, rheology, gel adhesive strength (in vitro), wet tissue approximation (in vivo), H&E staining of tissue sections at scheduled time intervals and tensile strength of the healed skin were carried out to assess the effectiveness of the EGK-glue in comparison with fibrin glue and cyanoacrylate. Results of UV-visible, NMR and CD analyses confirmed the functionalization and secondary structural changes. Increasing concentration of keratin reduces the gelation time (<15s). Lap-shear test demonstrates the maximum adhesive strength of 16.6±1.2kPa. Results of hemocompatibility and cytocompatibility studies suggested the suitability of the glue for clinical applications. Tissue approximation property assessed using the incision wound model (Wistar strain) in comparison with cyanoacrylate and fibrin glue suggested, that EGK-glue explicitly accelerates the rejoining of tissue with a 1.86 fold increase in skin tensile strength after healing. Imparting quinone moiety to gelatin-keratin conjugates through caffeic acid and a weaker oxidizing agent provides an adhesive glue with appreciable strength, and hemocompatible, cytocompatible and biodegradable properties, which, rejoin the cut tissue ends effectively. EGK-glue obtained in the present study finds wide biomedical/clinical applications. Copyright © 2013 Elsevier B.V. All rights reserved.

Effects of Plectin Depletion on Keratin Network Dynamics and Organization

PubMed Central

Moch, Marcin; Windoffer, Reinhard; Schwarz, Nicole; Pohl, Raphaela; Omenzetter, Andreas; Schnakenberg, Uwe; Herb, Fabian; Chaisaowong, Kraisorn; Merhof, Dorit; Ramms, Lena; Fabris, Gloria; Hoffmann, Bernd; Merkel, Rudolf; Leube, Rudolf E.

2016-01-01

The keratin intermediate filament cytoskeleton protects epithelial cells against various types of stress and is involved in fundamental cellular processes such as signaling, differentiation and organelle trafficking. These functions rely on the cell type-specific arrangement and plasticity of the keratin system. It has been suggested that these properties are regulated by a complex cycle of assembly and disassembly. The exact mechanisms responsible for the underlying molecular processes, however, have not been clarified. Accumulating evidence implicates the cytolinker plectin in various aspects of the keratin cycle, i.e., by acting as a stabilizing anchor at hemidesmosomal adhesion sites and the nucleus, by affecting keratin bundling and branching and by linkage of keratins to actin filament and microtubule dynamics. In the present study we tested these hypotheses. To this end, plectin was downregulated by shRNA in vulvar carcinoma-derived A431 cells. As expected, integrin β4- and BPAG-1-positive hemidesmosomal structures were strongly reduced and cytosolic actin stress fibers were increased. In addition, integrins α3 and β1 were reduced. The experiments furthermore showed that loss of plectin led to a reduction in keratin filament branch length but did not alter overall mechanical properties as assessed by indentation analyses using atomic force microscopy and by displacement analyses of cytoplasmic superparamagnetic beads using magnetic tweezers. An increase in keratin movement was observed in plectin-depleted cells as was the case in control cells lacking hemidesmosome-like structures. Yet, keratin turnover was not significantly affected. We conclude that plectin alone is not needed for keratin assembly and disassembly and that other mechanisms exist to guarantee proper keratin cycling under steady state conditions in cultured single cells. PMID:27007410

Isolation and Analysis of Keratins and Keratin-Associated Proteins from Hair and Wool.

PubMed

Deb-Choudhury, Santanu; Plowman, Jeffrey E; Harland, Duane P

2016-01-01

The presence of highly cross-linked protein networks in hair and wool makes them very difficult substrates for protein extraction, a prerequisite for further protein analysis and characterization. It is therefore imperative that these cross-links formed by disulfide bridges are first disrupted for the efficient extraction of proteins. Chaotropes such as urea are commonly used as efficient extractants. However, a combination of urea and thiourea not only improves recovery of proteins but also results in improved resolution of the keratins in 2DE gels. Reductants also play an important role in protein dissolution. Dithiothreitol effectively removes keratinous material from the cortex, whereas phosphines, like Tris(2-carboxyethyl)phosphine, remove material from the exocuticle. The relative extractability of the keratins and keratin-associated proteins is also dependent on the concentration of chaotropes, reductants, and pH, thus providing a means to preferentially extract these proteins. Ionic liquids such as 1-butyl-3-methylimidazolium chloride (BMIM(+)[Cl](-)) are known to solubilize wool by disrupting noncovalent interactions, specifically intermolecular hydrogen bonds. BMIM(+)[Cl](-) proved to be an effective extractant of wool proteins and complementary in nature to chaotropes such as urea and thiourea for identifying unique peptides of wool proteins using mass spectrometry (MS). Successful identification of proteins resolved by one- or two-dimensional electrophoresis and MS is highly dependent on the optimal recovery of its protease-digested peptides with an efficient removal of interfering substances. The detergent sodium deoxycholate used in conjunction with Empore™ disks improved identification of proteins by mass spectrometry leading to higher percentage sequence coverage, identification of unique peptides and higher score. Copyright © 2016 Elsevier Inc. All rights reserved.

Novel bilayer wound dressing based on electrospun gelatin/keratin nanofibrous mats for skin wound repair.

PubMed

Yao, Chun-Hsu; Lee, Chia-Yu; Huang, Chiung-Hua; Chen, Yueh-Sheng; Chen, Kuo-Yu

2017-10-01

A bilayer membrane (GKU) with a commercial polyurethane wound dressing as an outer layer and electrospun gelatin/keratin nanofibrous mat as an inner layer was fabricated as a novel wound dressing. Scanning electron micrographs showed that gelatin/keratin nanofibers had a uniform morphology and bead-free structure with average fiber diameter of 160.4nm. 3-(4,5-Dimethylthiazolyl)-2,5-diphenyltetrazolium bromide assay using L929 fibroblast cells indicated that the residues released from the gelatin/keratin composite nanofibrous mat accelerated cell proliferation. Cell attachment experiments revealed that adhered cells spread better and migrated deeper into the gelatin/keratin nanofibrous mat than that into the gelatin nanofibrous mat. In animal studies, compared with the bilayer membrane without keratin, gauze and commercial wound dressing, Comfeel®, GKU membrane gave much more number of blood vessels and a greater reduction in wound area at 4days, and better wound repair at 14days with a thicker epidermis and larger number of newly formed hair follicles. GKU membrane, thus, could be a good candidate for wound dressing applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Multicolor immunofluorescence reveals that p63- and/or K5-positive progenitor cells contribute to normal breast epithelium and usual ductal hyperplasia but not to low-grade intraepithelial neoplasia of the breast.

PubMed

Boecker, Werner; Stenman, Göran; Schroeder, Tina; Schumacher, Udo; Loening, Thomas; Stahnke, Lisa; Löhnert, Catharina; Siering, Robert Michael; Kuper, Arthur; Samoilova, Vera; Tiemann, Markus; Korsching, Eberhard; Buchwalow, Igor

2017-05-01

We contend that knowledge about the cellular composition of normal breast epithelium is a prerequisite for understanding proliferative breast disease. Against this background, we used multicolor immunofluorescence to study normal breast epithelium and two types of intraepithelial proliferative breast lesion for expression of the p63, basal keratin K5, glandular keratin K8/18, SMA, ER-alpha, and Ki67. We studied eight normal breast epithelium samples, 12 cases of usual ductal hyperplasia, and 33 cases of low-grade intraepithelial neoplasia (9 flat epithelial atypia, 14 low-grade ductal carcinoma in situ and 10 cases of lobular neoplasia). Usual ductal hyperplasia showed striking similarity to normal luminal breast epithelium including p63+ and/or K5+ luminal progenitor cells and the full spectrum of luminal progeny cells. In normal breast epithelium and usual ductal hyperplasia, expression of ER-alpha was associated with lack of expression of the proliferation antigen Ki67. In contrast, we found in both types of low-grade intraepithelial neoplasia robust expression of keratin K8/18 and a positive association between ER-alpha and Ki67 expression. However, these lesions were consistently negative for p63 and/or K5. Our observational study supports the view that usual ductal hyperplasia and low-grade intraepithelial neoplasia are different entities rather than part of a spectrum of the same disease. We propose a new operational model of cell differentiation that may serve to better understand correlations between normal breast epithelium and proliferative breast diseases. From our data we conclude that p63+ and/or K5+ progenitor cells contribute to maintenance of normal epithelium and usual ductal hyperplasia, but not to low-grade intraepithelial neoplasia of the breast.

Gene Expression Profiling in Pachyonychia Congenita Skin

PubMed Central

Cao, Yu-An; Hickerson, Robyn P.; Seegmiller, Brandon L.; Grapov, Dmitry; Gross, Maren M.; Bessette, Marc R.; Phinney, Brett S.; Flores, Manuel A.; Speaker, Tycho J.; Vermeulen, Annaleen; Bravo, Albert A.; Bruckner, Anna L.; Milstone, Leonard M.; Schwartz, Mary E.; Rice, Robert H.; Kaspar, Roger L.

2015-01-01

Background Pachyonychia congenita (PC) is a skin disorder resulting from mutations in keratin (K) proteins including K6a, K6b, K16, and K17. One of the major symptoms is painful plantar keratoderma. The pathogenic sequelae resulting from the keratin mutations remain unclear. Objective To better understand PC pathogenesis. Methods RNA profiling was performed on biopsies taken from PC-involved and uninvolved plantar skin of seven genotyped PC patients (two K6a, one K6b, three K16, and one K17) as well as from control volunteers. Protein profiling was generated from tape-stripping samples. Results A comparison of PC-involved skin biopsies to adjacent uninvolved plantar skin identified 112 differentially-expressed mRNAs common to patient groups harboring K6 (i.e., both K6a and K6b) and K16 mutations. Among these mRNAs, 25 encode structural proteins including keratins, small proline-rich and late cornified envelope proteins, 20 are related to metabolism and 16 encode proteases, peptidases, and their inhibitors including kallikrein-related peptidases (KLKs), and serine protease inhibitors (SERPINs). mRNAs were also identified to be differentially expressed only in K6 (81) or K16 (141) patient samples. Furthermore, 13 mRNAs were identified that may be involved in pain including nociception and neuropathy. Protein profiling, comparing three K6a plantar tape-stripping samples to non-PC controls, showed changes in the PC corneocytes similar, but not identical, to the mRNA analysis. Conclusion Many differentially-expressed genes identified in PC-involved skin encode components critical for skin barrier homeostasis including keratinocyte proliferation, differentiation, cornification, and desquamation. The profiling data provide a foundation for unraveling the pathogenesis of PC and identifying targets for developing effective PC therapeutics. PMID:25656049

New keratin isolates: actives for natural hair protection.

PubMed

Roddick-Lanzilotta, Alisa; Kelly, Rob; Scott, Sonya; Chahal, Surinder

2007-01-01

Hair is primarily composed of keratin proteins and it is well established that peptides and proteins bestow desirable effects on the hair, for example improving moisturization and softness. In the present work we describe how keratin actives with unique properties convey a range of beneficial properties to a variety of hair types. It has been observed that these functional keratins protect hair from damage associated with chemical treatments such as perming and relaxation, help to restore the mechanical strength of damaged fibers and decrease fading of colored hair.

Pangolin armor: Overlapping, structure, and mechanical properties of the keratinous scales.

PubMed

Wang, Bin; Yang, Wen; Sherman, Vincent R; Meyers, Marc A

2016-09-01

The pangolin has a flexible dermal armor consisting of overlapping keratinous scales. Although they show potential for bioinspired flexible armor, the design principles of pangolin armor are barely known. Here we report on the overlapping organization, hierarchical structure (from the nano to the mesolevel), and mechanical response of scales from ground (Chinese) and arboreal (African tree) pangolins. Both scales exhibit the same overlapping organization, with each scale at the center of neighboring scales arranged in a hexagonal pattern. The scales have a cuticle of several layers of loosely attached flattened keratinized cells, while the interior structure exhibits three regions distinguished by the geometry and orientations of the keratinized cells, which form densely packed lamellae; each one corresponds to one layer of cells. Unlike most other keratinous materials, the scales show a crossed-lamellar structure (∼5μm) and crossed fibers (∼50μm). A nano-scale suture structure, observed for the first time, outlines cell membranes and leads to an interlocking interface between lamellae, thus enhancing the bonding and shear resistance. The tensile response of the scales shows an elastic limit followed by a short plateau prior to failure, with Young's modulus ∼1 GPa and tensile strength 60-100MPa. The mechanical response is transversely isotropic, a result of the cross lamellar structure. The strain rate sensitivity in the range of 10(-5)-10(-1)s(-1) region is found to be equal to 0.07-0.08, typical of other keratins and polymers. The mechanical response is highly dependent on the degree of hydration, a characteristic of keratins. Although many fish and reptiles have protective scales and carapaces, mammals are characteristically fast and light. The pangolin is one of the few mammal possessing a flexible dermal armor for protection from predators, such as lions. Here we study the arrangement of the scales as well as their hierarchical structure from the nano

UV irradiation-induced methionine oxidation in human skin keratins: Mass spectrometry-based non-invasive proteomic analysis.

PubMed

Lee, Seon Hwa; Matsushima, Keita; Miyamoto, Kohei; Oe, Tomoyuki

2016-02-05

Ultraviolet (UV) radiation is the major environmental factor that causes oxidative skin damage. Keratins are the main constituents of human skin and have been identified as oxidative target proteins. We have recently developed a mass spectrometry (MS)-based non-invasive proteomic methodology to screen oxidative modifications in human skin keratins. Using this methodology, UV effects on methionine (Met) oxidation in human skin keratins were investigated. The initial screening revealed that Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UVA (or UVB) irradiation of human tape-stripped skin. Subsequent liquid chromatography/electrospray ionization-MS and tandem MS analyses confirmed amino acid sequences and oxidation sites of tryptic peptides D(290)VDGAYMTK(298) (P1) and N(258)MQDMVEDYR(267) (P2). The relative oxidation levels of P1 and P2 increased in a time-dependent manner upon UVA irradiation. Butylated hydroxytoluene was the most effective antioxidant for artifactual oxidation of Met residues. The relative oxidation levels of P1 and P2 after UVA irradiation for 48 h corresponded to treatment with 100mM hydrogen peroxide for 15 min. In addition, Met(259) was oxidized by only UVA irradiation. The Met sites identified in conjunction with the current proteomic methodology can be used to evaluate skin damage under various conditions of oxidative stress. We demonstrated that the relative Met oxidation levels in keratins directly reflected UV-induced damages to human tape-stripped skin. Human skin proteins isolated by tape stripping were analyzed by MS-based non-invasive proteomic methodology. Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UV irradiation. Met(259) was oxidized by only UVA irradiation. Quantitative LC/ESI-SRM/MS analyses confirmed a time-dependent increase in the relative oxidation of target peptides (P1 and P2) containing these Met residues, upon UVA irradiation

Effects of grain processing methods on the expression of genes involved in volatile fatty acid transport and pH regulation, and keratinization in rumen epithelium of beef cattle

PubMed Central

Del Bianco Benedeti, Pedro; Silva, Breno de Castro; Pacheco, Marcos Vinícius Carneiro; Carvalho Filho, Ivan; Lopes, Mariana Mescouto; Marcondes, Marcos Inácio; Mantovani, Hilário Cuquetto; Valadares Filho, Sebastião de Campos; Detmann, Edenio

2018-01-01

Two experiments were carried out to evaluate the effects of corn and sorghum with different processing methods on the expression of genes involved in volatile fatty acids transport and pH regulation, and ruminal keratinization in rumen epithelium of finishing bulls. For Exp. 1, five rumen cannulated Nellore bulls were used in a 5x5 Latin square arrangement, with 14 d for adaptation and 9 d for sample collection. Treatments were: dry ground corn, dry ground sorghum, reconstituted corn, reconstituted sorghum, and control (forage-based diet). Samples of rumen epithelium from ventral sac were excised, rinsed, snap-frozen and stored at -80°C until total RNA isolation and quantitative real-time PCR analysis. In the Exp. 2, 24 Nellore bulls were assigned to a completely randomized design lasting 168 d. Experimental treatments were similar to those at Exp. 1, but without the control treatment. After the experimental period, bulls were slaughtered and rumen epithelium samples were rapidly excised for further histological analysis. Rumen epithelial tissue from animals fed reconstituted corn had lower expression of downregulated-in-adenoma (P = 0.03) and Na+/H+ exchanger 2 (trend; P = 0.09). The expression of Na+/ H+ exchanger 1 (P = 0.10) and putative anion transporter (P = 0.06) tended to be lower in rumen epithelium of bulls fed reconstituted grains. Ruminal concentration of valerate was greater for animals fed reconstituted grain (P = 0.01). Likewise, animals fed reconstituted corn tended to have greater butyrate ruminal concentration (P = 0.08). Keratinized layer thickness did not differ among treatments (P > 0.10). Therefore, reconstituted grains (especially corn) decrease the mRNA expression of genes involved in volatile fatty acids transport and pH control in the rumen epithelium. PMID:29902237

"Panta rhei": Perpetual cycling of the keratin cytoskeleton.

PubMed

Leube, Rudolf E; Moch, Marcin; Kölsch, Anne; Windoffer, Reinhard

2011-01-01

The filamentous cytoskeletal systems fulfil seemingly incompatible functions by maintaining a stable scaffolding to ensure tissue integrity and simultaneously facilitating rapid adaptation to intracellular processes and environmental stimuli. This paradox is particularly obvious for the abundant keratin intermediate filaments in epithelial tissues. The epidermal keratin cytoskeleton, for example, supports the protective and selective barrier function of the skin while enabling rapid growth and remodelling in response to physical, chemical and microbial challenges. We propose that these dynamic properties are linked to the perpetual re-cycling of keratin intermediate filaments that we observe in cultured cells. This cycle of assembly and disassembly is independent of protein biosynthesis and consists of distinct, temporally and spatially defined steps. In this way, the keratin cytoskeleton remains in constant motion but stays intact and is also able to restructure rapidly in response to specific regulatory cues as is needed, e.g., during division, differentiation and wound healing.

Single-Nucleotide Polymorphisms Associated with Skin Naphthyl–Keratin Adduct Levels in Workers Exposed to Naphthalene

PubMed Central

Jiang, Rong; French, John E.; Stober, Vandy P.; Kang-Sickel, Juei-Chuan C.; Zou, Fei

2012-01-01

Background: Individual genetic variation that results in differences in systemic response to xenobiotic exposure is not accounted for as a predictor of outcome in current exposure assessment models. Objective: We developed a strategy to investigate individual differences in single-nucleotide polymorphisms (SNPs) as genetic markers associated with naphthyl–keratin adduct (NKA) levels measured in the skin of workers exposed to naphthalene. Methods: The SNP-association analysis was conducted in PLINK using candidate-gene analysis and genome-wide analysis. We identified significant SNP–NKA associations and investigated the potential impact of these SNPs along with personal and workplace factors on NKA levels using a multiple linear regression model and the Pratt index. Results: In candidate-gene analysis, a SNP (rs4852279) located near the CYP26B1 gene contributed to the 2-naphthyl–keratin adduct (2NKA) level. In the multiple linear regression model, the SNP rs4852279, dermal exposure, exposure time, task replacing foam, age, and ethnicity all were significant predictors of 2NKA level. In genome-wide analysis, no single SNP reached genome-wide significance for NKA levels (all p ≥ 1.05 × 10–5). Pathway and network analyses of SNPs associated with NKA levels were predicted to be involved in the regulation of cellular processes and homeostasis. Conclusions: These results provide evidence that a quantitative biomarker can be used as an intermediate phenotype when investigating the association between genetic markers and exposure–dose relationship in a small, well-characterized exposed worker population. PMID:22391508

Feather keratin hydrolysates obtained from microbial keratinases: effect on hair fiber

PubMed Central

2013-01-01

Background Hair is composed mainly of keratin protein and a small amount of lipid. Protein hydrolysates, in particular those with low molecular weight distribution have been known to protect hair against chemical and environmental damage. Many types of protein hydrolysates from plants and animals have been used in hair and personal care such as keratin hydrolysates obtained from nails, horns and wool. Most of these hydrolysates are obtained by chemical hydrolysis and hydrothermal methods, but recently hydrolyzed hair keratin, feather keratin peptides, and feather meal peptides have been obtained by enzymatic hydrolysis using Bacillus spp in submerged fermentation. Results Keratin peptides were obtained by enzymatic hydrolysis of keratinases using Bacillus subtilis AMR. The microorganism was grown on a feather medium, pH 8.0 (1% feathers) and supplemented with 0.01% of yeast extract, for 5 days, at 28°C with agitation. The supernatant containing the hydrolysates was colleted by centrifugation and ultra filtered in an AMICON system using nano–membranes (Millipore – YC05). The Proteins and peptides were analyzed using HPTLC and MALDI-TOF-MS. Commercial preparations of keratin hydrolysates were used as a comparative standard. After five days the feather had been degraded (90-95%) by the peptidases and keratinases of the microorganism. MALDI-TOF mass spectrometry showed multiple peaks that correspond to peptides in the range of 800 to 1079 Daltons and the commercial hydrolysate was in the range of 900 to 1400 Da. HPTLC showed lower molecular mass peptides and amino acids in the enzymatic hydrolysate when compared with the commercial hydrolysate . A mild shampoo and a rinse off conditioner were formulated with the enzymatic hydrolysate and applied to hair fibers to evaluate the hydration, with and without heat, using a Corneometer® CM 825. The hydration was more efficient with heat, suggesting a more complete incorporation of hydrolysates into the fibers

Keratin-6 driven ODC expression to hair follicle keratinocytes enhances stemness and tumorigenesis by negatively regulating Notch

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arumugam, Aadithya; Weng, Zhiping; Chaudhary, Sandeep C.

Highlights: • Targeting ODC to hair follicle augments skin carcinogenesis and invasive SCCs. • Hair follicle ODC expands stem cell compartment carrying CD34{sup +}/K15{sup +}/p63{sup +} keratinocytes. • Negatively regulated Notch1 is associated with expansion of stem cell compartment. - Abstract: Over-expression of ornithine decarboxylase (ODC) is known to be involved in the epidermal carcinogenesis. However, the mechanism by which it enhances skin carcinogenesis remains undefined. Recently, role of stem cells localized in various epidermal compartments has been shown in the pathogenesis of skin cancer. To direct ODC expression in distinct epidermal compartments, we have developed keratin 6 (K6)-ODC/SKH-1 andmore » keratin 14 (K14)-ODC/SKH-1 mice and employed them to investigate the role of ODC directed to these epidermal compartments on UVB-induced carcinogenesis. K6-driven ODC over-expression directed to outer root sheath (ORS) of hair follicle was more effective in augmenting tumorigenesis as compared to mice where K14-driven ODC expression was directed to inter-follicular epidermal keratinocytes. Chronically UVB-irradiated K6-ODC/SKH-1 developed 15 ± 2.5 tumors/mouse whereas K14-ODC/SKH-1 developed only 6.8 ± 1.5 tumors/mouse. K6-ODC/SKH-1 showed augmented UVB-induced proliferation and much higher pro-inflammatory responses than K14-ODC/SKH-1 mice. Tumors induced in K6-ODC/SKH-1 were rapidly growing, invasive and ulcerative squamous cell carcinoma (SCC) showing decreased expression of epidermal polarity marker E-cadherin and enhanced mesenchymal marker, fibronectin. Interestingly, the number of CD34/CK15/p63 positive stem-like cells was significantly higher in chronically UVB-irradiated K6-ODC/SKH-1 as compared to K14-ODC/SKH-1 mice. Reduced Notch1 expression was correlated with the expansion of stem cell compartment in these animals. However, other signaling pathways such as DNA damage response or mTOR signaling pathways were not significantly

[Immunohistochemical observation on keratin filaments of cultured tumor cells by ABC staining].

PubMed

Wang, J; Yang, F

1991-06-01

Avidin-Biotin Peroxidase complex technique, ABV staining, was employed by using monoclonal anti-keratin antibody HK2 in this study. The organization and dynamics of keratins in both interphase and mitotic T56 and HeLa cells were analysed. We also observed the effects of microtubule (MT) and microfilament (MF) inhibitors, colchicine and cytochalasin B, on the organization of keratin filaments in T56 and HeLa cells. The results showed that a significant alteration in the structural organization and distribution of keratin filaments occurred during mitosis, and an extensive rearrangement of keratin networks of the two cell lines was induced in interphase after the MT and MF were disrupted by combined treatment with the two drugs, colchicine and cytochalasin B; the keratin networks turned into a star-like lattice rapidly within 1-2h. Neither colchicine nor cytochalasin B alone elicited significant organizational change in the keratin networks of the two cell lines.

Wear resistance of Polymethyl Methacrylate (PMMA) with the Addition of Bone Ash, Hydroxylapatite and Keratin

NASA Astrophysics Data System (ADS)

Emre, G.; Akkus, A.; Karamış, M. B.

2018-01-01

In this study mechanichal and tribological properties of keratin, bone ash and hydroxylapatite by adding to PMMA ( known as the main prosthesis material) were investigated. Hydroxylapatite, bone ash, and keratin materials were added as PMMA in to the content of PMMA, in the proportions of %1, %3 and %5, respectively. The resulting mixtures were put into the molds and solidified in order to obtain samples to be used in the wear experiments. Each experiment was conducted by preparing three experimental samples. The wear data were compared according to the average values of the experimental samples. In the wear test, the results were also evaluated according to the average values obtained from each group and the results of the control group. It was observed that, the wear resistance of the PMMA including 3%, 5% bone ash and PMMA including 5% keratin flour were higher than the values of the control group.

Microbial decomposition of keratin in nature-a new hypothesis of industrial relevance.

PubMed

Lange, Lene; Huang, Yuhong; Busk, Peter Kamp

2016-03-01

Discovery of keratin-degrading enzymes from fungi and bacteria has primarily focused on finding one protease with efficient keratinase activity. Recently, an investigation was conducted of all keratinases secreted from a fungus known to grow on keratinaceous materials, such as feather, horn, and hooves. The study demonstrated that a minimum of three keratinases is needed to break down keratin, an endo-acting, an exo-acting, and an oligopeptide-acting keratinase. Further, several studies have documented that disruption of sulfur bridges of the keratin structure acts synergistically with the keratinases to loosen the molecular structure, thus giving the enzymes access to their substrate, the protein structure. With such complexity, it is relevant to compare microbial keratin decomposition with the microbial decomposition of well-studied polymers such as cellulose and chitin. Interestingly, it was recently shown that the specialized enzymes, lytic polysaccharide monoxygenases (LPMOs), shown to be important for breaking the recalcitrance of cellulose and chitin, are also found in keratin-degrading fungi. A holistic view of the complex molecular self-assembling structure of keratin and knowledge about enzymatic and boosting factors needed for keratin breakdown have been used to formulate a hypothesis for mode of action of the LPMOs in keratin decomposition and for a model for degradation of keratin in nature. Testing such hypotheses and models still needs to be done. Even now, the hypothesis can serve as an inspiration for designing industrial processes for keratin decomposition for conversion of unexploited waste streams, chicken feather, and pig bristles into bioaccessible animal feed.

The role of allogenic keratin-derived dressing in wound healing in a mouse model.

PubMed

Konop, Marek; Sulejczak, Dorota; Czuwara, Joanna; Kosson, Piotr; Misicka, Aleksandra; Lipkowski, Andrzej W; Rudnicka, Lidia

2017-01-01

Keratin is an interesting protein needed for wound healing and tissue recovery. We have recently proposed a new, simple and inexpensive method to obtain fur and hair keratin-derived biomaterials suitable for medical application. The aim of the study was to evaluate the role of the fur keratin-derived protein (FKDP) dressing in the allogenic full-thickness surgical skin wound model. The data obtained using scanning electron microscopy showed that employed processed biomaterial had higher surface porosity compared with control raw material. From the MTS test, it was found keratin biomaterial is not only toxic to the NIH/3T3 cell line (p < 0.05), but also enhances cell proliferation compared with the control. In vivo studies have shown keratin dressings are tissue biocompatible, accelerate wound closure and epithelialization to the statistically significant differences on day 5 (p < 0.05) in comparison to control wounds. Histological examination revealed, that in FKDP-treated wounds the inflammatory response contained predominantly macrophages whilst their morphological untreated variants showed mixed cell infiltrates rich in neutrophils. Predominant macrophages based response creates more favorable environment for healing. In FKDP-dressed wounds the number of microhemorrhages was also significantly decreased (p < 0.05) as compared with undressed wounds. Applied keratin dressing favors reconstruction of a more regular skin structure and assures better cosmetic effect in terms of scar formation and appearance. In conclusion, fur keratin-derived protein dressings significantly accelerated wound healing in the mouse model. Further studies are needed to determine the molecular mechanisms involved in the multilayer wound healing process and to assess the possible use of these dressings for medical purposes. © 2016 by the Wound Healing Society.

Brazilian keratin hair treatment: a review.

PubMed

Weathersby, Courtney; McMichael, Amy

2013-06-01

Brazilian keratin treatments are widely available products that are used by women all over the world to straighten hair. Marketers of these products claim that the keratin treatments render naturally curly hair more manageable and frizz-free while enhancing color and shine, giving the hair a healthier appearance. Although widely used, there have been virtually no reports of adverse side effects. Unfortunately, many of the products that are applied by salon professionals contain formaldehyde or its derivatives and are being marketed as safe. © 2013 Wiley Periodicals, Inc.

Keratin gel in the management of Epidermolysis bullosa.

PubMed

Denyer, J; Marsh, C; Kirsner, R S

2015-10-01

Epidermolysis bullosa (EB) describes a number of genetically inherited conditions which cause skin fragility and minor trauma leading to skin damage, skin loss and wounding. Owing to the fragility of the skin and requirement for frequent dressing changes, at present, the optimal dressing(s) is not clear. Our objective was to assess the use of a keratin gel in the management of wounds in patients with different forms of EB. We treated patients with different types of EB and a range of wounds with a novel keratin gel. In a convenience sample of consecutive patients, we introduced the keratin gel into their treatment regimen maintaining other aspects of their care. Patients reported faster healing and more resilient healed skin. Of the ten patients treated in this pilot study, six found the gel effective; two found it ineffective; and in two patients, it caused itching leading to discontinuation of the treatment. The results of this case study series suggest that keratin gel can be useful in the management of EB and are consistent with previous published experiences.

The structure of the "amorphous" matrix of keratins.

PubMed

Kadir, Murat; Wang, Xinwei; Zhu, Bowen; Liu, Jing; Harland, Duane; Popescu, Crisan

2017-05-01

Various keratin fibers, particularly human hairs, were investigated by transmission electron microscopy, TEM, solid-state 1 H NMR and Transient Electro-Thermal Technique, TET. The results converge to suggest that the matrix of keratin fiber cortex, far from being amorphous, has a well-defined nano-scale grainy structure, the size of these grains being around 2-4nm. The size of the grains appears to strongly depend on the chemical treatment of the fiber, on the temperature and on the relative humidity of the environment, as well as on the physiological factors at the level of fiber production in follicle. By suggesting an organization at the nano-scale of the protein chains in these grains, likely to be Keratin Associated Proteins, the results challenge the view of matrix as a homogeneous glassy material. Moreover, they indicate the potential of further investigating the purpose of this structure that appears to reflect not only chemical treatments of keratins but also biological processes at the level of the follicle. Copyright © 2017 Elsevier Inc. All rights reserved.

Normal keratinized mucosa transplants in nude mice.

PubMed

Holmstrup, P; Dabelsteen, E; Reibel, J; Harder, F

1981-01-01

Two types of normal keratinized mucosa were transplanted to subcutaneous sites of nude mice of two different strains. 24 intact specimens of clinically normal human palatal mucosa were transplanted to nude mice of the strain nu/nu NC. The transplants were recovered after 42 d with a recovery rate of 96%. Moreover, 22 intact specimens of normal rat forestomach mucosa were transplanted to nude mice of the strain nu/nu BALB/c/BOM. These transplants were recovered after 21 d with a recovery rate of 63%. The histologic features of the transplants were essentially the same as those of the original tissues. However, epithelial outgrowths from the transplants differed with respect to the pattern of keratinization. The outgrowths of human palatal mucosa transplants were essentially unkeratinized, while the outgrowths of the rat forestomach transplants showed continued keratinization.

Epigenetic control of skin differentiation genes by phytocannabinoids

PubMed Central

Pucci, Mariangela; Rapino, Cinzia; Di Francesco, Andrea; Dainese, Enrico; D'Addario, Claudio; Maccarrone, Mauro

2013-01-01

BACKGROUND AND PURPOSE Endocannabinoid signalling has been shown to have a role in the control of epidermal physiology, whereby anandamide is able to regulate the expression of skin differentiation genes through DNA methylation. Here, we investigated the possible epigenetic regulation of these genes by several phytocannabinoids, plant-derived cannabinoids that have the potential to be novel therapeutics for various human diseases. EXPERIMENTAL APPROACH The effects of cannabidiol, cannabigerol and cannabidivarin on the expression of skin differentiation genes keratins 1 and 10, involucrin and transglutaminase 5, as well as on DNA methylation of keratin 10 gene, were investigated in human keratinocytes (HaCaT cells). The effects of these phytocannabinoids on global DNA methylation and the activity and expression of four major DNA methyltransferases (DNMT1, 3a, 3b and 3L) were also examined. KEY RESULTS Cannabidiol and cannabigerol significantly reduced the expression of all the genes tested in differentiated HaCaT cells, by increasing DNA methylation of keratin 10 gene, but cannabidivarin was ineffective. Remarkably, cannabidiol reduced keratin 10 mRNA through a type-1 cannabinoid (CB1) receptor-dependent mechanism, whereas cannabigerol did not affect either CB1 or CB2 receptors of HaCaT cells. In addition, cannabidiol, but not cannabigerol, increased global DNA methylation levels by selectively enhancing DNMT1 expression, without affecting DNMT 3a, 3b or 3L. CONCLUSIONS AND IMPLICATIONS These findings show that the phytocannabinoids cannabidiol and cannabigerol are transcriptional repressors that can control cell proliferation and differentiation. This indicates that they (especially cannabidiol) have the potential to be lead compounds for the development of novel therapeutics for skin diseases. PMID:23869687

Development and Characterization of a 3D Printed, Keratin-Based Hydrogel.

PubMed

Placone, Jesse K; Navarro, Javier; Laslo, Gregory W; Lerman, Max J; Gabard, Alexis R; Herendeen, Gregory J; Falco, Erin E; Tomblyn, Seth; Burnett, Luke; Fisher, John P

2017-01-01

Keratin, a naturally-derived polymer derived from human hair, is physiologically biodegradable, provides adequate cell support, and can self-assemble or be crosslinked to form hydrogels. Nevertheless, it has had limited use in tissue engineering and has been mainly used as casted scaffolds for drug or growth factor delivery applications. Here, we present and assess a novel method for the printed, sequential production of 3D keratin scaffolds. Using a riboflavin-SPS-hydroquinone (initiator-catalyst-inhibitor) photosensitive solution we produced 3D keratin constructs via UV crosslinking in a lithography-based 3D printer. The hydrogels obtained have adequate printing resolution and result in compressive and dynamic mechanical properties, uptake and swelling capacities, cytotoxicity, and microstructural characteristics that are comparable or superior to those of casted keratin scaffolds previously reported. The novel keratin-based printing resin and printing methodology presented have the potential to impact future research by providing an avenue to rapidly and reproducibly manufacture patient-specific hydrogels for tissue engineering and regenerative medicine applications.

K-sam, an amplified gene in stomach cancer, is a member of the heparin-binding growth factor receptor genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hattori, Yutaka; Odagiri, Hiroki; Nakatani, Hiroshi

1990-08-01

DNA fragments amplified in a stomach cancer-derived cell line, KATO-III, were previously identified by the in-gel DNA renaturation method, and a 0.2-kilobase-pair fragment of the amplified sequence was subsequently cloned. By genomic walking, a portion of the exon of the gene flanking this 0.2-kilobase-pair fragment was cloned, and the gene was designated as K-sam ({und K}ATO-III cell-derived {und s}tomach cancer {und am}plified gene). The K-sam cDNAs, corresponding to the 3.5-kilobase K-sam mRNA, were cloned from the KATO-III cells. Sequence analysis revealed that this gene coded for 682 amino acid residues that satisfied the characteristics of the receptor tyrosine kinase. Themore » K-sam gene had significant homologies with bek, FLG, and chicken basic fibroblast growth factor receptor gene. The K-sam gene was amplified in KATO-III cells with the major transcript of 3.5-kilobases in size. This gene was also expressed in some other stomach cancer cells, a small cell lung cancer, and germ cell tumors.« less

Linking the molecular evolution of avian beta (β) keratins to the evolution of feathers.

PubMed

Greenwold, Matthew J; Sawyer, Roger H

2011-12-15

Feathers of today's birds are constructed of beta (β)-keratins, structural proteins of the epidermis that are found solely in reptiles and birds. Discoveries of "feathered dinosaurs" continue to stimulate interest in the evolutionary origin of feathers, but few studies have attempted to link the molecular evolution of their major structural proteins (β-keratins) to the appearance of feathers in the fossil record. Using molecular dating methods, we show that before the appearance of Anchiornis (∼155 Million years ago (Ma)) the basal β-keratins of birds began diverging from their archosaurian ancestor ∼216 Ma. However, the subfamily of feather β-keratins, as found in living birds, did not begin diverging until ∼143 Ma. Thus, the pennaceous feathers on Anchiornis, while being constructed of avian β-keratins, most likely did not contain the feather β-keratins found in the feathers of modern birds. Our results demonstrate that the evolutionary origin of feathers does not coincide with the molecular evolution of the feather β-keratins found in modern birds. More likely, during the Late Jurassic, the epidermal structures that appeared on organisms in the lineage leading to birds, including early forms of feathers, were constructed of avian β-keratins other than those found in the feathers of modern birds. Recent biophysical studies of the β-keratins in feathers support the view that the appearance of the subfamily of feather β-keratins altered the biophysical nature of the feather establishing its role in powered flight. Copyright © 2011 Wiley Periodicals, Inc., A Wiley Company.

Keratinous inclusion cyst of oesophagus: unusual finding

PubMed Central

Wan Abdul Rahman, Wan Faiziah; Mutum, Samarendra Singh; Fauzi, Mohd Hashairi

2013-01-01

Cysts of the oesophagus are unusual findings and they are classified according to the embryological site of origin. It may represent inclusion cysts, retention cysts and developmental cysts. We present a case of keratinous inclusion cyst of the lower oesophagus in a 71-year-old Malay woman who presented with dyspepsia and severe epigastric pain. An oesophago-gastro-duodenoscopy demonstrated a sliding hiatus hernia with whitish ulcer-like lesion at the lower oesophagus. Biopsy from the lesion revealed a keratinous inclusion cyst. The patient was given pantoprazole and put on regular follow-up for monitoring any other development. PMID:23878290

Keratin 17 Mutations in Four Families from India with Pachyonychia Congenita

PubMed Central

Agarwala, Manoj; Salphale, Pankaj; Peter, Dincy; Wilson, Neil J; Pulimood, Susanne; Schwartz, Mary E; Smith, Frances J D

2017-01-01

Pachyonychia congenita (PC) is a rare autosomal dominant genetic skin disorder due to a mutation in any one of the five keratin genes, KRT6A, KRT6B, KRT6C, KRT16, or KRT17. The main features are palmoplantar keratoderma, plantar pain, and nail dystrophy. Cysts of various types, follicular hyperkeratosis, oral leukokeratosis, hyperhidrosis, and natal teeth may also be present. Four unrelated Indian families presented with a clinical diagnosis of PC. This was confirmed by genetic testing; mutations in KRT17 were identified in all affected individuals. PMID:28794556

Click chemistry modification of natural keratin fibers for sustained shrink-resist performance.

PubMed

Yu, Dan; Cai, Jackie Y; Church, Jeffrey S; Wang, Lijing

2015-01-01

This paper introduces a novel chemical treatment for achieving sustained shrink-resist performance on natural keratin fibers. The new treatment involves the controlled reduction of keratin in the cuticle region of the fiber, and the application of a water soluble diacrylate, namely glycerol 1,3-diglycerolate diacrylate (GDA), on the reduced keratin substrate. The acrylate groups of the GDA react with cysteine residues in the reduced keratin through thiol-ene click reactions at room temperature, leading to GDA grafting and the formation of GDA crosslinks in the keratin structure. The modified substrates were characterized by infrared spectroscopy and scanning electron microscopy, and assessed for its shrink-resistance and wet burst strength. This chemical modification has shown to alter the fiber surface morphology and hydrophilicity, resulting in substantially improved shrink-resistance with good fiber strength retention. Possible shrink-resistance mechanisms were also discussed. Crown Copyright © 2015. Published by Elsevier B.V. All rights reserved.

Keratin-lipid structural organization in the corneous layer of snake.

PubMed

Ripamonti, Alberto; Alibardi, Lorenzo; Falini, Giuseppe; Fermani, Simona; Gazzano, Massimo

2009-12-01

The shed epidermis (molt) of snakes comprises four distinct layers. The upper two layers, here considered as beta-layer, contain essentially beta-keratin. The following layer, known as mesos-layer, is similar to the human stratum corneum, and is formed by thin cells surrounded by intercellular lipids. The latter layer mainly contains alpha-keratin. In this study, the molecular assemblies of proteins and lipids contained in these layers have been analyzed in the scale of two species of snakes, the elapid Tiger snake (TS, Notechis scutatus) and the viperid Gabon viper (GV, Bitis gabonica). Scanning X-ray micro-diffraction, FTIR and Raman spectroscopies, thermal analysis, and scanning electron microscopy experiments confirm the presence of the three layers in the GV skin scale. Conversely, in the TS molt a typical alpha-keratin layer appears to be absent. In the latter, experimental data suggest the presence of two domains similar to those found in the lipid intercellular matrix of stratum corneum. X-ray diffraction data also allow to determine the relative orientation of keratins and lipids. The keratin fibrils are randomly oriented inside the layers parallel to the surface of scales while the lipids are organized in lamellar structures having aliphatic chains normal to the scale surface. The high ordered lipid organization in the mature mesos layer probably increases its effectiveness in limiting water-loss.

Cytoskeleton in motion: the dynamics of keratin intermediate filaments in epithelia.

PubMed

Windoffer, Reinhard; Beil, Michael; Magin, Thomas M; Leube, Rudolf E

2011-09-05

Epithelia are exposed to multiple forms of stress. Keratin intermediate filaments are abundant in epithelia and form cytoskeletal networks that contribute to cell type-specific functions, such as adhesion, migration, and metabolism. A perpetual keratin filament turnover cycle supports these functions. This multistep process keeps the cytoskeleton in motion, facilitating rapid and protein biosynthesis-independent network remodeling while maintaining an intact network. The current challenge is to unravel the molecular mechanisms underlying the regulation of the keratin cycle in relation to actin and microtubule networks and in the context of epithelial tissue function.

Cytoskeleton in motion: the dynamics of keratin intermediate filaments in epithelia

PubMed Central

Windoffer, Reinhard; Beil, Michael; Magin, Thomas M.

2011-01-01

Epithelia are exposed to multiple forms of stress. Keratin intermediate filaments are abundant in epithelia and form cytoskeletal networks that contribute to cell type–specific functions, such as adhesion, migration, and metabolism. A perpetual keratin filament turnover cycle supports these functions. This multistep process keeps the cytoskeleton in motion, facilitating rapid and protein biosynthesis–independent network remodeling while maintaining an intact network. The current challenge is to unravel the molecular mechanisms underlying the regulation of the keratin cycle in relation to actin and microtubule networks and in the context of epithelial tissue function. PMID:21893596

Glutamic acid promotes monacolin K production and monacolin K biosynthetic gene cluster expression in Monascus.

PubMed

Zhang, Chan; Liang, Jian; Yang, Le; Chai, Shiyuan; Zhang, Chenxi; Sun, Baoguo; Wang, Chengtao

2017-12-01

This study investigated the effects of glutamic acid on production of monacolin K and expression of the monacolin K biosynthetic gene cluster. When Monascus M1 was grown in glutamic medium instead of in the original medium, monacolin K production increased from 48.4 to 215.4 mg l -1 , monacolin K production increased by 3.5 times. Glutamic acid enhanced monacolin K production by upregulating the expression of mokB-mokI; on day 8, the expression level of mokA tended to decrease by Reverse Transcription-polymerase Chain Reaction. Our findings demonstrated that mokA was not a key gene responsible for the quantity of monacolin K production in the presence of glutamic acid. Observation of Monascus mycelium morphology using Scanning Electron Microscope showed glutamic acid significantly increased the content of Monascus mycelium, altered the permeability of Monascus mycelium, enhanced secretion of monacolin K from the cell, and reduced the monacolin K content in Monascus mycelium, thereby enhancing monacolin K production.

Cultivation of human dermal fibroblasts and epidermal keratinocytes on keratin-coated silica bead substrates.

PubMed

Tan, Bee Yi; Nguyen, Luong T H; Kim, Hyo-Sop; Kim, Jae-Ho; Ng, Kee Woei

2017-10-01

Human hair keratin is promising as a bioactive material platform for various biomedical applications. To explore its versatility further, human hair keratin was coated onto monolayers of silica beads to produce film-like substrates. This combination was hypothesized to provide a synergistic effect in improving the biochemical properties of the resultant composite. Atomic force microscopy analysis showed uniform coatings of keratin on the silica beads with a slight increase in the resulting surface roughness. Keratin-coated silica beads had higher surface energy and relatively lower negative charge than those of bare silica beads. To investigate cell response, human dermal fibroblasts (HDFs), and human epidermal keratinocytes (HEKs) were cultured on the substrates over 4 days. Results showed that keratin coatings significantly enhanced the metabolic activity of HDFs and encouraged cell spreading but did not exert any significant effects on HEKs. HDF expression of collagen I was significantly more intense on the keratin-coated compared to the bare silica substrates. Furthermore, HDF secretion of various cytokines suggested that keratin coatings triggered active cell responses related to wound healing. Collectively, our study demonstrated that human hair keratin-coated silica bead monolayers have the potential to modulate HDF behavior in culture and may be exploited further. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2789-2798, 2017. © 2017 Wiley Periodicals, Inc.

A Cancer Gene Selection Algorithm Based on the K-S Test and CFS.

PubMed

Su, Qiang; Wang, Yina; Jiang, Xiaobing; Chen, Fuxue; Lu, Wen-Cong

2017-01-01

To address the challenging problem of selecting distinguished genes from cancer gene expression datasets, this paper presents a gene subset selection algorithm based on the Kolmogorov-Smirnov (K-S) test and correlation-based feature selection (CFS) principles. The algorithm selects distinguished genes first using the K-S test, and then, it uses CFS to select genes from those selected by the K-S test. We adopted support vector machines (SVM) as the classification tool and used the criteria of accuracy to evaluate the performance of the classifiers on the selected gene subsets. This approach compared the proposed gene subset selection algorithm with the K-S test, CFS, minimum-redundancy maximum-relevancy (mRMR), and ReliefF algorithms. The average experimental results of the aforementioned gene selection algorithms for 5 gene expression datasets demonstrate that, based on accuracy, the performance of the new K-S and CFS-based algorithm is better than those of the K-S test, CFS, mRMR, and ReliefF algorithms. The experimental results show that the K-S test-CFS gene selection algorithm is a very effective and promising approach compared to the K-S test, CFS, mRMR, and ReliefF algorithms.

Immunocytochemical and autoradiographic studies on the process of keratinization in avian epidermis suggests absence of keratohyalin.

PubMed

Alibardi, Lorenzo

2004-02-01

The process of keratinization in apteric avian epidermis and in scutate scales of some avian species has been studied by autoradiography for histidine and immunohistochemistry for keratins and other epidermal proteins. Acidic or basic alpha-keratins are present in basal, spinosus, and transitional layers, but are not seen in the corneous layer. Keratinization-specific alpha-keratins (AE2-positive) are observed in the corneous layer of apteric epidermis but not in that of scutate scales, which contain mainly beta-keratin. Alpha-keratin bundles accumulate along the plasma membrane of transitional cells of apteric epidermis. In contrast to the situation in scutate scales, in the transitional layer and in the lowermost part of the corneous layer of apteric epidermis, filaggrin-like, loricrin-like, and transglutaminase immunoreactivities are present. The lack of isopeptide bond immunoreactivity suggests that undetectable isopeptide bonds are present in avian keratinocytes. Using immunogold ultrastructural immunocytochemistry a low but localized loricrin-like and, less, filaggrin-like labeling is seen over round-oval granules or vesicles among keratin bundles of upper spinosus and transitional keratinocytes of apteric epidermis. Filaggrin-and loricrin-labeling are absent in alpha-keratin bundles localized along the plasma membrane and in the corneous layer, formerly considered keratohyalin. Using ultrastructural autoradiography for tritiated histidine, occasional trace grains are seen among these alpha-keratin bundles. A different mechanism of redistribution of matrix and corneous cell envelope proteins probably operates in avian keratinocytes as compared to that of mammals. Keratin bundles are compacted around the lipid-core of apteric epidermis keratinocytes, which do not form complex chemico/mechanical-resistant corneous cell envelopes as in mammalian keratinocytes. These observations suggest that low amounts of matrix proteins are present among keratin bundles of

Immunohistochemical demonstration of keratins in the epidermal layers of the Malayan pangolin (Manis javanica), with remarks on the evolution of the integumental scale armour.

PubMed

Meyer, W; Liumsiricharoen, M; Suprasert, A; Fleischer, L G; Hewicker-Trautwein, M

2013-09-16

Using immunohistochemistry, the study demonstrates the distribution of keratins (pan-keratin with CK1-8, 10, 14-16, 19; keratins CK1, 5, 6, 9, 10; hair keratins AE13, AE14) in the epidermis of the Malayan pangolin (Manis javanica). A varying reaction spectrum was observed for pan-keratin, with body region-dependent negative to very strong reaction intensities. The dorsolateral epidermis exhibited positive reactions only in its vital layers, whereas the abdominal epidermis showed strong positive reactions in the soft two outer strata. The single acidic and basic-to-neutral (cyto)keratins produced clear variations compared to the pan-keratin tinging. E.g., CK1 appeared in all epidermal layers of both body regions, except for the ventral stratum corneum, whereas CK5, 6, 9, 10 were restricted to the soft ventral epidermis. Here, distinctly positive reactions were confined to the stratum granulosum, except for CK6 that appeared in the soft stratum corneum. A different staining pattern was obvious for the hair keratins, i.e., positive reactions of AE13 concentrated only in the granular layer of the dorsal epidermis. In the abdominal epidermis, remarkable tinging for AE14 was visible in the stratum basale, decreasing toward the corneal layer, but was also found in the outer root sheath cells of the hair follicles in the ventral body part. Our findings are discussed related to the evolution of the horny dorsal scales of the pangolin, which may have started from the tail root, projecting forward to the head.

Modified approach for keratinized tissue augmentation in multiple teeth

PubMed Central

Terenzi, Mayara; Pigossi, Suzane Cristina; Pires, Luana Carla; Cirelli, Joni Augusto; Sampaio, José Eduardo

2017-01-01

This case report demonstrated a modified technique of free gingival graft (FGG) aiming to increase keratinized attached tissue in large recipient areas. A FGG to increase the amount of attached gingival tissue, facilitate oral hygiene, and prevent further clinical attachment loss was realized in two patients. Because the extensive recipient area, a modified technique was performed to obtain a smaller graft of the donor area. A template of the graft was made about 25%–30% smaller than the total recipient area. After graft removal, interspersed incisions were made in the upper and lower edges of it. After 9–24 months of follow-up, the final width of the keratinized tissue was 4.0–4.4 times larger in comparison to initial clinical condition. In conclusion, this FGG technique can be considered an alternative to gain sufficient amount of keratinized gingival tissue using a smaller graft. PMID:29551874

Keratinization-associated miR-7 and miR-21 Regulate Tumor Suppressor Reversion-inducing Cysteine-rich Protein with Kazal Motifs (RECK) in Oral Cancer*

PubMed Central

Jung, Hyun Min; Phillips, Brittany L.; Patel, Rushi S.; Cohen, Donald M.; Jakymiw, Andrew; Kong, William W.; Cheng, Jin Q.; Chan, Edward K. L.

2012-01-01

MicroRNAs (miRNAs) are small non-coding RNAs that posttranscriptionally regulate gene expression during many biological processes. Recently, the aberrant expressions of miRNAs have become a major focus in cancer research. The purpose of this study was to identify deregulated miRNAs in oral cancer and further focus on specific miRNAs that were related to patient survival. Here, we report that miRNA expression profiling provided more precise information when oral squamous cell carcinomas were subcategorized on the basis of clinicopathological parameters (tumor primary site, histological subtype, tumor stage, and HPV16 status). An innovative radar chart analysis method was developed to depict subcategories of cancers taking into consideration the expression patterns of multiple miRNAs combined with the clinicopathological parameters. Keratinization of tumors and the high expression of miR-21 were the major factors related to the poor prognosis of patients. Interestingly, a majority of the keratinized tumors expressed high levels of miR-21. Further investigations demonstrated the regulation of the tumor suppressor gene reversion-inducing cysteine-rich protein with kazal motifs (RECK) by two keratinization-associated miRNAs, miR-7 and miR-21. Transfection of miR-7 and miR-21-mimics reduced the expression of RECK through direct miRNA-mediated regulation, and these miRNAs were inversely correlated with RECK in CAL 27 orthotopic xenograft tumors. Furthermore, a similar inverse correlation was demonstrated in CAL 27 cells treated in vitro by different external stimuli such as trypsinization, cell density, and serum concentration. Taken together, our data show that keratinization is associated with poor prognosis of oral cancer patients and keratinization-associated miRNAs mediate deregulation of RECK which may contribute to the aggressiveness of tumors. PMID:22761427

Prevalence of genes encoding virulence factors among Escherichia coli with K1 antigen and non-K1 E. coli strains.

PubMed

Kaczmarek, Agnieszka; Budzynska, Anna; Gospodarek, Eugenia

2012-10-01

Multiplex PCR was used to detect genes encoding selected virulence determinants associated with strains of Escherichia coli with K1 antigen (K1(+)) and non-K1 E. coli (K1(-)). The prevalence of the fimA, fimH, sfa/foc, ibeA, iutA and hlyF genes was studied for 134 (67 K1(+) and 67 K1(-)) E. coli strains isolated from pregnant women and neonates. The fimA gene was present in 83.6 % of E. coli K1(+) and in 86.6 % of E. coli K1(-) strains. The fimH gene was present in all tested E. coli K1(+) strains and in 97.0 % of non-K1 strains. E. coli K1(+) strains were significantly more likely to possess the following genes than E. coli K1(-) strains: sfa/foc (37.3 vs 16.4 %, P = 0.006), ibeA (35.8 vs 4.5 %, P<0.001), iutA (82.1 vs 35.8 %, P<0.001) and hlyF (28.4 vs 6.0 %, P<0.001). In conclusion, E. coli K1(+) seems to be more virulent than E. coli K1(-) strains in developing severe infections, thereby increasing possible sepsis or neonatal bacterial meningitis.

Self-assembly of keratin peptides: Its implication on the performance of electrospun PVA nanofibers

PubMed Central

Kadirvelu, Kavitha; Fathima, Nishter Nishad

2016-01-01

Drawing inspiration from the field of designer self-assembling materials, this work is aimed to focus on the self-assembling nature of extracted peptides. Hair keratin, a proteinacious reject in tanning industry has been chosen since they have been extracted and used for wide range of applications. Keratin source was subjected to five hydrolysis treatments (viz., sulphitolysis, β-mercaptoethanol, ionic liquid, thioglycolic acid and alkali) and assayed for functional groups. This was followed by the prediction of secondary structure using circular dichroism, determining the microstructural level to which the extracted peptide has self-assembled. Sulphitolysis and thioglycolic acid based hydrolysates exist in monomeric conformation, whereas β-mercaptoethanol based hydrolysate exhibited dimeric conformation. The subsequent part of the study is to incorporate these peptides into the nanofibers to study the structural implication of keratin peptides on its characteristics. Accordingly, the peptides were electrospun with PVA and subjected to morphological, mechanical, thermal and biological characterizations. Monomeric nanofiber mat has high tensile strength of around 5.5 MPa and offered lower mass transport resistance, whereas dimeric mat has high Tm of around 290 °C and was more biocompatible. These results help in understanding the extraction-structure-function aspect of the hydrolysates stressing the role of extraction methods on the choice of application. PMID:27812004

Keratin based bioplastic film from chicken feathers and its characterization.

PubMed

Ramakrishnan, Navina; Sharma, Swati; Gupta, Arun; Alashwal, Basma Yahya

2018-05-01

Plastics have been one of the highly valued materials and it plays an significant role in human's life such as in food packaging and biomedical applications. Bioplastic materials can gradually work as a substitute for various materials based on fossil oil. The issue like sustainability and environmental challenges which occur due to manufacturing and disposal of synthetic plastics can be conquering by bio-based plastics. Feathers are among the most inexpensive abundant, and renewable protein sources. Feathers disposal to the landfills leads to environmental pollutions and it results into wastage of 90% of protein raw material. Keratin is non-burning hydrophilic, and biodegradable due to which it can be applicable in various ways via chemical processing. Main objective of this research is to synthesis bioplastic using keratin from chicken feathers. Extracted keratin solution mixed with different concentration of glycerol (2 to 10%) to produce plastic films. The mixture was stirred under constant magnetic stirring at 60 °C for 5 h. The mixtures are then poured into aluminum weighing boat and dried in an oven at 60 °C for 24 h. The mechanical properties of the samples were tested and the physic-chemical properties of the bioplastic were studied. According to the results, Scanning Electron Microscopy test showed good compatible morphologies without holes, cavity and edge. The difference in chemical composition was analyzed using Fourier transform infrared spectroscopy (FTIR). The samples were also characterized by thermo gravimetric analysis (TGA), differential scanning calorimetry (DSC), X-Ray diffraction (XRD) to check the thermal and crystallinity properties. Other than that, bioplastic made up from keratin with 2% of glycerol has the best mechanical and thermal properties. According to biodegradability test, all bioplastic produced are proven biodegradable. Therefore, the results showed possible application of the film as an alternative to fossil oil

Clinical efficacy of a xenogeneic collagen matrix in augmenting keratinized mucosa around implants: a randomized controlled prospective clinical trial.

PubMed

Lorenzo, Ramón; García, Virginia; Orsini, Marco; Martin, Conchita; Sanz, Mariano

2012-03-01

The aim of this controlled randomized clinical trial was to evaluate the efficacy of a xenogeneic collagen matrix (CM) to augment the keratinized tissue around implants supporting prosthetic restorations at 6 months when compared with the standard treatment, the connective tissue autograft, CTG). This randomized longitudinal parallel controlled clinical trial studied 24 patients with at least one location with minimal keratinized tissue (≤1 mm). The 6-month width of keratinized tissue. As secondary outcomes the esthetic outlook, the maintenance of peri-implant mucosal health and the patient morbidity were assessed pre-operatively and 1, 3, and 6 months post-operatively. At 6 months, Group CTG attained a mean width of keratinized tissue of 2.75 (1.5) mm, while the corresponding figure in Group CM was 2.8 (0.4) mm, the inter-group differences not being statistically significant. The surgical procedure in both groups did not alter significantly the mucosal health in the affected abutments. There was a similar esthetic result and significant increase in the vestibular depth in both groups as a result of the surgery. In the CM group it changed from 2.2 (3.3) to 5.1 (2.5) mm at 6 months. The patients treated with the CM referred less pain, needed less pain medication, and the surgical time was shorter, although these differences were not statistically significant when compared with the CTG group. These results prove that this new CM was as effective and predictable as the CTG for attaining a band of keratinized tissue. © 2011 John Wiley & Sons A/S.

Investigation of preparation techniques for δ2H analysis of keratin materials and a proposed analytical protocol

USGS Publications Warehouse

Qi, H.; Coplen, T.B.

2011-01-01

Accurate hydrogen isotopic measurements of keratin materials have been a challenge due to exchangeable hydrogen in the sample matrix and the paucity of appropriate isotopic reference materials for calibration. We found that the most reproducible δ2HVSMOW-SLAP and mole fraction of exchangeable hydrogen, x(H)ex, of keratin materials were measured with equilibration at ambient temperature using two desiccators and two different equilibration waters with two sets of the keratin materials for 6 days. Following equilibration, drying the keratin materials in a vacuum oven for 4 days at 60 °C was most critical. The δ2H analysis protocol also includes interspersing isotopic reference waters in silver tubes among samples in the carousel of a thermal conversion elemental analyzer (TC/EA) reduction unit. Using this analytical protocol, δ2HVSMOW-SLAP values of the non-exchangeable fractions of USGS42 and USGS43 human-hair isotopic reference materials were determined to be –78.5 ± 2.3 ‰ and –50.3 ± 2.8 ‰, respectively. The measured x(H)ex values of keratin materials analyzed with steam equilibration and N2 drying were substantially higher than those previously published, and dry N2 purging was unable to remove absorbed moisture completely, even with overnight purging. The δ2H values of keratin materials measured with steam equilibration were about 10 ‰ lower than values determined with equilibration in desiccators at ambient temperatures when on-line evacuation was used to dry samples. With steam equilibrations the x(H)ex of commercial keratin powder was as high as 28 %. Using human-hair isotopic reference materials to calibrate other keratin materials, such as hoof or horn, can introduce bias in δ2H measurements because the amount of absorbed water and the x(H)ex values may differ from those of unknown samples. Correct δ2HVSMOW-SLAP values of the non-exchangeable fractions of unknown human-hair samples can be determined with atmospheric moisture

Drug-induced keratin 9 interaction with Hsp70 in bladder cancer cells.

PubMed

Andolino, C; Hess, C; Prince, T; Williams, H; Chernin, M

2018-05-25

A pull-down experiment (co-immunoprecipitation) was performed on a T24 human bladder cancer cell lysate treated with the Hsp inhibitor VER155008 using an Hsp70 antibody attached to Dynabeads. Keratin 9, a cytoskeleton intermediate filament protein, was identified by LC MS/MS analysis. This novel finding was confirmed by Western blotting, RT-PCR, and immunocytochemistry. Other members of the keratin family of proteins have been shown to be involved in cancer progression, most recently identified to be associated with cell invasion and metastasis. The specific role of keratin 9 expression in these cells is yet to be determined.

Novel immobilization of a quaternary ammonium moiety on keratin fibers for medical applications.

PubMed

Yu, Dan; Cai, Jackie Y; Liu, Xin; Church, Jeffrey S; Wang, Lijing

2014-09-01

This paper introduces a new approach for immobilizing a quaternary ammonium moiety on a keratinous substrate for enhanced medical applications. The method involves the generation of thiols by controlled reduction of cystine disulfide bonds in the keratin, followed by reaction with [2-(acryloyloxy)ethyl]trimethylammonium chloride through thiol-ene click chemistry. The modified substrate was characterized with Raman and infrared spectroscopy, and assessed for its antibacterial efficacy and other performance changes. The results have demonstrated that the quaternary ammonium moiety has been effectively attached onto the keratin structure, and the resultant keratin substrate exhibits a multifunctional effect including antibacterial and antistatic properties, improved liquid moisture management property, improved dyeability and a non-leaching characteristic of the treated substrate. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.

Clinical evaluation of a collagen matrix to enhance the width of keratinized gingiva around dental implants

PubMed Central

Lee, Kang-Ho; Kim, Byung-Ock

2010-01-01

Purpose The purpose of this study was to evaluate the effect of collagen matrix with apically positioned flap (APF) on the width of keratinized gingiva, comparing to the results of APF only and APF combined with free gingival graft (FGG) at the second implant surgery. Methods Nine patients were selected from those who had received treatments at the Department of Periodontics, Chosun University Dental Hospital, Gwangju, Korea. We performed APF, APF combined with FGG, and APF combined with collagen matrix coverage respectively. Clinical evaluation of keratinized gingival was performed by measuring the distance from the gingival crest to the mucogingival junction at the mid-buccal point, using a periodontal probe before and after the surgery. Results The ratio of an increase was 0.3, 0.6, and 0.6 for the three subjects in the APF cases, 3, 5, and 7 for the three in the APF combined with FGG case, and 1.5, 0.5, and 3 for the three in the APF combined with collagen matrix coverage case. Conclusions This study suggests that the collagen matrix when used as a soft tissue substitute with the aim of increasing the width of keratinized tissue or mucosa, was as effective and predictable as the FGG. PMID:20498767

Recombinant Escherichia coli K5 strain with the deletion of waaR gene decreases the molecular weight of the heparosan capsular polysaccharide.

PubMed

Huang, Haichan; Liu, Xiaobo; Lv, Shencong; Zhong, Weihong; Zhang, Fuming; Linhardt, Robert J

2016-09-01

Heparosan, the capsular polysaccharide of Escherichia coli K5 having a carbohydrate backbone similar to that of heparin, has become a potential precursor for bioengineering heparin. In the heparosan biosynthesis pathway, the gene waaR encoding α-1-, 2- glycosyltransferase catalyze s the third glucosyl residues linking to the oligosaccharide chain. In the present study, a waaR deletion mutant of E. coli K5 was constructed. The mutant showed improvement of capsule polysaccharide yield. It is interesting that the heparosan molecular weight of the mutant is reduced and may become more suitable as a precursor for the production of low molecular weight heparin derived from the wild-type K5 capsular polysaccharide.

Inclusion bodies as potential vehicles for recombinant protein delivery into epithelial cells

PubMed Central

2012-01-01

Background We present the potential of inclusion bodies (IBs) as a protein delivery method for polymeric filamentous proteins. We used as cell factory a strain of E. coli, a conventional host organism, and keratin 14 (K14) as an example of a complex protein. Keratins build the intermediate filament cytoskeleton of all epithelial cells. In order to build filaments, monomeric K14 needs first to dimerize with its binding partner (keratin 5, K5), which is then followed by heterodimer assembly into filaments. Results K14 IBs were electroporated into SW13 cells grown in culture together with a “reporter” plasmid containing EYFP labeled keratin 5 (K5) cDNA. As SW13 cells do not normally express keratins, and keratin filaments are built exclusively of keratin heterodimers (i.e. K5/K14), the short filamentous structures we obtained in this study can only be the result of: a) if both IBs and plasmid DNA are transfected simultaneously into the cell(s); b) once inside the cells, K14 protein is being released from IBs; c) released K14 is functional, able to form heterodimers with EYFP-K5. Conclusions Soluble IBs may be also developed for complex cytoskeletal proteins and used as nanoparticles for their delivery into epithelial cells. PMID:22624805

A low K+ signal is required for functional high-affinity K+ uptake through HAK5 transporters.

PubMed

Rubio, Francisco; Fon, Mario; Ródenas, Reyes; Nieves-Cordones, Manuel; Alemán, Fernando; Rivero, Rosa M; Martínez, Vicente

2014-11-01

The high-affinity K(+) transporter HAK5 is a key system for root K(+) uptake and, under very low external K(+), the only one capable of supplying K(+) to the plant. Functional HAK5-mediated K(+) uptake should be tightly regulated for plant adaptation to different environmental conditions. Thus, it has been described that the gene encoding the transporter is transcriptionally regulated, being highly induced under K(+) limitation. Here we show that environmental conditions, such as the lack of K(+), NO(3)(-) or P, that induced a hyperpolarization of the plasma membrane of root cells, induce HAK5 transcription. However, only the deprivation of K(+) produces functional HAK5-mediated K(+) uptake in the root. These results suggest on the one hand the existence of a posttranscriptional regulation of HAK5 elicited by the low K(+) signal and on the other that HAK5 may be involved in yet-unknown functions related to NO(3)(-) and P deficiencies. These results have been obtained here with Solanum lycopersicum (cv. Micro-Tom) as well as Arabidopsis thaliana plants, suggesting that the posttranscriptional regulation of high-affinity HAK transporters take place in all plant species. © 2014 Scandinavian Plant Physiology Society.

α-keratin/Alginate Biosorbent for Removal of Methylene Blue on Aqueous Solution in a Batch System

NASA Astrophysics Data System (ADS)

Fadillah, G.; Putri, E. N. K.; Febrianastuti, S.; Munawaroh, H.; Purnawan, C.; Wahyuningsih, S.

2018-03-01

Methylene Blue (MB) is a cationic dyes which is commonly used in textile industries for coloring agent. The precence of MB in water caused some negative effect on the environment and human health. Many common technologies such as membrane filtration, electrophoresis and adsorption have been widely empolyed for removal of MB in water, but the adsorption technique still has advantages than the others. In this study, removal of MB used a biosorbent α-keratin/alginate (KA). The biosorbent KA was prepared by using the encapsulation technique in CaCl2 2 % (w/v) solution. The biosorbent was characterized by Fourier Transform Infrared (FTIR) and Scanning Electron Microscope (SEM). The effect of composition of α-keratin and alginate, the pH of solution and contact time on the adsorption were investigated. The optimum adsorption of MB in aqueous solution was found at the composition of α-keratin and alginate of 1:2 (w/w), the pH at 5.0 and contact time at 4 hours. The adsorption of MB on KA biosorbent was comparatively higher than α-keratin and alginate only. Adsorption of MB dyes in aqueous solution followed the Langmuir adsorption isotherm, and the dynamic adsorption model could be described through a pseudo-second order kinetics.

Various Techniques to Increase Keratinized Tissue for Implant Supported Overdentures: Retrospective Case Series

PubMed Central

Cayarga, Rodrigo; Suzuki, Takanori; Kaufman, Zev

2015-01-01

Purpose. The purpose of this retrospective case series is to describe and compare different surgical techniques that can be utilized to augment the keratinized soft tissue around implant-supported overdentures. Materials and Methods. The data set was extracted as deidentified information from the routine treatment of patients at the Ashman Department of Periodontology and Implant Dentistry at New York University College of Dentistry. Eight edentulous patients were selected to be included in this study. Patients were treated for lack of keratinized tissue prior to implant placement, during the second stage surgery, and after delivery of the final prosthesis. Results. All 8 patients in this study were wearing a complete maxillary and/or mandibular denture for at least a year before the time of the surgery. One of the following surgical techniques was utilized to increase the amount of keratinized tissue: apically positioned flap (APF), pedicle graft (PG), connective tissue graft (CTG), or free gingival graft (FGG). Conclusions. The amount of keratinized tissue should be taken into consideration when planning for implant-supported overdentures. The apical repositioning flap is an effective approach to increase the width of keratinized tissue prior to the implant placement. PMID:26124833

Preparation and study on the structure of keratin/PVA membrane containing wool fibers

NASA Astrophysics Data System (ADS)

Wu, Min; Shen, Shuming; Yang, Xuhong; Tang, Rencheng

2017-10-01

The urea / sodium sulfide / sodium dodecyl sulfate (SDS) method was used to dissolve the wool in this study. Then the Wool fiber/keratin/PVA composites with different proportions were prepared, and the surface morphology, molecular structure, mechanical property of the composite films and the influence of the proportions on their structure and properties were studied. The results showed that, there are α-helix structure, β-sheet and random coil conformations in the pure keratin film, as well as in the wool fiber. Compared with wool fiber, the crystallinity of keratin decreased. PVA can obviously improve the mechanical property of the blended film. When the blended ratio of keratin/PVA is 20/80, the mechanical property of the blended film is greatly improved. The composite films with 8%-16% of wool fibers have better flexibility than those without wool fibers.

Calcium phosphate coated Keratin-PCL scaffolds for potential bone tissue regeneration.

PubMed

Zhao, Xinxin; Lui, Yuan Siang; Choo, Caleb Kai Chuen; Sow, Wan Ting; Huang, Charlotte Liwen; Ng, Kee Woei; Tan, Lay Poh; Loo, Joachim Say Chye

2015-04-01

The incorporation of hydroxyapatite (HA) nanoparticles within or on the surface of electrospun polymeric scaffolds is a popular approach for bone tissue engineering. However, the fabrication of osteoconductive composite scaffolds via benign processing conditions still remains a major challenge to date. In this work, a new method was developed to achieve a uniform coating of calcium phosphate (CaP) onto electrospun keratin-polycaprolactone composites (Keratin-PCL). Keratin within PCL was crosslinked to decrease its solubility, before coating of CaP. A homogeneous coating was achieved within a short time frame (~10min) by immersing the scaffolds into Ca(2+) and (PO4)(3-) solutions separately. Results showed that the incorporation of keratin into PCL scaffolds not only provided nucleation sites for Ca(2+) adsorption and subsequent homogeneous CaP surface deposition, but also facilitated cell-matrix interactions. An improvement in the mechanical strength of the resultant composite scaffold, as compared to other conventional coating methods, was also observed. This approach of developing a biocompatible bone tissue engineering scaffold would be adopted for further in vitro osteogenic differentiation studies in the future. Copyright © 2015 Elsevier B.V. All rights reserved.

Preparation of keratin-based microcapsules for encapsulation of hydrophilic molecules.

PubMed

Rajabinejad, Hossein; Patrucco, Alessia; Caringella, Rosalinda; Montarsolo, Alessio; Zoccola, Marina; Pozzo, Pier Davide

2018-01-01

The interest towards microcapsules based on non-toxic, biodegradable and biocompatible polymers, such as proteins, is increasing considerably. In this work, microcapsules were prepared using water soluble keratin, known as keratoses, with the aim of encapsulating hydrophilic molecules. Keratoses were obtained via oxidizing extraction of pristine wool, previously degreased by Soxhlet. In order to better understand the shell part of microcapsules, pristine wool and obtained keratoses were investigated by FT-IR, gel-electrophoresis and HPLC. Production of the microcapsules was carried out by a sonication method. Thermal properties of microcapsules were investigated by DSC. Microencapsulation and dye encapsulation yields were obtained by UV-spectroscopy. Morphological structure of microcapsules was studied by light microscopy, SEM, and AFM. The molecular weights of proteins analyzed using gel-electrophoresis resulted in the range of 38-62kDa. The results confirmed that the hydrophilic dye (Telon Blue) was introduced inside the keratoses shells by sonication and the final microcapsules diameter ranged from 0.5 to 4µm. Light microscope investigation evidenced the presence of the dye inside the keratoses vesicles, confirming their capability of encapsulating hydrophilic molecules. The microcapsule yield and dye encapsulation yield were found to be 28.87±3% and 83.62±5% respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

Developing Breast Cancer Program at Xavier; Genomic and Proteomic Analysis of Signaling Pathways Involved in Xenohormone and MEK5 Regulation of Breast Cancer

DTIC Science & Technology

2010-05-01

Heat shock 70 kDa protein 4 (HSPA4), Glutathione S-transferase P ( GSTP1 ), and Creatine kinase B-type (CKB), and downregulations of Keratin 8 (KRT8...VIM), glutathione-S-transferase P ( GSTP1 ), and creatine kinase B-type (CKB) were upregulated, and keratin 8 (KRT8), keratin 19 (KRT19) and...upregulation of VIM/vim, GSTP1 / gstp1 , and CKB/ckb; and downregulation of KRT8/krt8, KRT19/krt19, and GSTM3/gstm3). We further demonstrate that MEK5

Epithelial estrogen receptor 1 intrinsically mediates squamous differentiation in the mouse vagina.

PubMed

Miyagawa, Shinichi; Iguchi, Taisen

2015-10-20

Estrogen-mediated actions in female reproductive organs are tightly regulated, mainly through estrogen receptor 1 (ESR1). The mouse vaginal epithelium cyclically exhibits cell proliferation and differentiation in response to estrogen and provides a unique model for analyzing the homeostasis of stratified squamous epithelia. To address the role of ESR1-mediated tissue events during homeostasis, we analyzed mice with a vaginal epithelium-specific knockout of Esr1 driven by keratin 5-Cre (K5-Esr1KO). We show here that loss of epithelial ESR1 in the vagina resulted in aberrant epithelial cell proliferation in the suprabasal cell layers and led to failure of keratinized differentiation. Gene expression analysis showed that several known estrogen target genes, including erbB growth factor ligands, were not induced by estrogen in the K5-Esr1KO mouse vagina. Organ culture experiments revealed that the addition of erbB growth factor ligands, such as amphiregulin, could activate keratinized differentiation in the absence of epithelial ESR1. Thus, epithelial ESR1 integrates estrogen and growth factor signaling to mediate regulation of cell proliferation in squamous differentiation, and our results provide new insights into estrogen-mediated homeostasis in female reproductive organs.

Keratin film ablation for the fabrication of brick and mortar skin structure using femtosecond laser pulses

NASA Astrophysics Data System (ADS)

Haq, Bibi Safia; Khan, Hidayat Ullah; Dou, Yuehua; Alam, Khan; Attaullah, Shehnaz; Zari, Islam

2015-09-01

The patterning of thin keratin films has been explored to manufacture model skin surfaces based on the "bricks and mortar" view of the relationship between keratin and lipids. It has been demonstrated that laser light is capable of preparing keratin-based "bricks and mortar" wall structure as in epidermis, the outermost layer of the human skin. "Bricks and mortar" pattern in keratin films has been fabricated using an ArF excimer laser (193 nm wavelength) and femtosecond laser (800 and 400 nm wavelength). Due to the very low ablation threshold of keratin, femtosecond laser systems are practical for laser processing of proteins. These model skin structures are fabricated for the first time that will help to produce potentially effective moisturizing products for the protection of skin from dryness, diseases and wrinkles.

Investigating the microstructure of keratin extracted from wool: peptide sequence (MALDI-TOF/TOF) and protein conformation (FTIR)

USDA-ARS?s Scientific Manuscript database

Keratin was extracted from wool by reduction with 2-mercaptoethanol. It was isolated as intact keratin and characterized by its similar molecular weight, protein composition, and secondary structure to native keratin. Gel electrophoresis patterns and MALDI-TOF/TOF peptide sequences provided the ide...

A Sporadic Neonatal Case of Epidermolysis Bullosa Simplex Generalized Intermediate with KRT5 and KRT14 Gene Mutations

PubMed Central

Wakiguchi, Hiroyuki; Hasegawa, Shunji; Maeba, Shinji; Kimura, Sasagu; Ito, Satoko; Tateishi, Hiroshi; Ueda, Kazuhiro; Ohga, Shouichi

2016-01-01

Background Epidermolysis bullosa simplex (EBS) is a rare genodermatosis resulting from multiple gene mutations, including KRT5 and KRT14. The clinical expression of the mechanobullous skin fragility disease has not been fully explained by the genotype. Case Description An 11-day-old Japanese newborn infant was hospitalized because of herpetiform skin blistering on the feet, which expanded systemically after birth. There was no evidence of virus infection. The biopsied skin lesion showed a blister on the lamina densa without keratin clumps, indicating a diagnosis of EBS-generalized intermediate. We punctured the blisters to remove the contents daily, which led to no exacerbation or infection. The genetic study determined that the patient carried double substitutions of KRT5 c.1424A > G (p.E475G) and KRT14 c.1237G > A (p.A413T). The asymptomatic mother and sister carried the KRT14 substitution, but the healthy father had no substitution of the KRT gene. Conclusion This is the first report of EBS-generalized intermediate in a newborn with de novo KRT5 gene mutation and KRT14 gene polymorphism, and no familial history of epidermolysis. Neonatal blistering due to EBS requires optimal skin management after excluding infectious and immunobullous diseases. PMID:26929861

Localization of Alpha-Keratin and Beta-Keratin (Corneous Beta Protein) in the Epithelium on the Ventral Surface of the Lingual Apex and Its Lingual Nail in the Domestic Goose (Anser Anser f. domestica) by Using Immunohistochemistry and Raman Microspectroscopy Analysis.

PubMed

Skieresz-Szewczyk, Kinga; Jackowiak, Hanna; Buchwald, Tomasz; Szybowicz, Mirosław

2017-08-01

The epithelium of the ventral surface of the apex of the tongue in most birds is specified by the presence of the special superficial layer called lingual nail. The aim of the present study is to determine the localization of the alpha-keratin and beta-keratin (corneous beta protein) in this special epithelium in the domestic goose by using immunohistochemistry staining and the Raman spectroscopy analysis. Due to lack of commercially available antibodies to detect beta-keratin (corneous beta protein), the Raman spectroscopy was used as a specific tool to detect and describe the secondary structure of proteins. The immunohistochemical (IHC) detections reveal the presence of alpha-keratin in all layers of the epithelium, but significant differences in the distribution of the alpha-keratin in the epithelial layers appear. The staining reaction is stronger from the basal layer to the upper zone of the intermediate layer. The unique result is weak staining for the alpha-keratin in the lingual nail. Applications of the Raman spectroscopy as a complementary method not only confirmed results of IHC staining for alpha-keratin, but showed that this technique could be used to demonstrate the presence of beta-keratin (corneous beta protein). Functionally, the localization of alpha-keratin in the epithelium of the ventral surface of the lingual apex provides a proper scaffold for epithelial cells and promotes structural integrity, whereas the presence of beta-keratin (corneous beta protein) in the lingual nail, described also as exoskeleton of the ventral surface of the apex, endures mechanical stress. Anat Rec, 300:1361-1368, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Intratarsal keratinous eyelid cysts in Gorlin syndrome: A review and reappraisal.

PubMed

Wolkow, Natalie; Jakobiec, Frederick A; Yoon, Michael K

2017-12-27

A 38-year-old woman presented with multiple bilateral recurrent eyelid cysts. Her medical history was notable for Gorlin (nevoid basal cell carcinoma) syndrome. Histopathologic and immunohistochemical examinations revealed that the lesions were intratarsal keratinous cysts. They were similar in appearance to sporadic intratarsal keratinous cysts and closely resembled odontogenic keratocysts of the jaw. Eyelid cysts occur in up to 40% of patients with Gorlin syndrome; however, their description has been cursory and, for the most part, outside of the ophthalmic literature. Although ophthalmologists are familiar with the periocular basal cell carcinomas that occur in patients with Gorlin syndrome, up to 10% of patients never develop a basal cell carcinoma, but they may manifest other ophthalmic findings. Awareness of these other features may contribute to the earlier diagnosis of the syndrome. We discuss the clinical and histopathologic features of intratarsal keratinous cysts in Gorlin syndrome, comparing them to sporadic intratarsal keratinous cysts, other eyelid cysts, and jaw cysts that also characterize this syndrome. We briefly review the ocular and systemic manifestations of Gorlin syndrome and recent genetic and therapeutic developments so that the eyelid cysts may be appreciated within the appropriate context of Gorlin syndrome as a whole. Copyright © 2017 Elsevier Inc. All rights reserved.

Keratin 8/18 regulation of glucose metabolism in normal versus cancerous hepatic cells through differential modulation of hexokinase status and insulin signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mathew, Jasmin; Loranger, Anne; Gilbert, Stéphane

2013-02-15

As differentiated cells, hepatocytes primarily metabolize glucose for ATP production through oxidative phosphorylation of glycolytic pyruvate, whereas proliferative hepatocellular carcinoma (HCC) cells undergo a metabolic shift to aerobic glycolysis despite oxygen availability. Keratins, the intermediate filament (IF) proteins of epithelial cells, are expressed as pairs in a lineage/differentiation manner. Hepatocyte and HCC (hepatoma) cell IFs are made solely of keratins 8/18 (K8/K18), thus providing models of choice to address K8/K18 IF functions in normal and cancerous epithelial cells. Here, we demonstrate distinctive increases in glucose uptake, glucose-6-phosphate formation, lactate release, and glycogen formation in K8/K18 IF-lacking hepatocytes and/or hepatoma cellsmore » versus their respective IF-containing counterparts. We also show that the K8/K18-dependent glucose uptake/G6P formation is linked to alterations in hexokinase I/II/IV content and localization at mitochondria, with little effect on GLUT1 status. In addition, we find that the insulin-stimulated glycogen formation in normal hepatocytes involves the main PI-3 kinase-dependent signaling pathway and that the K8/K18 IF loss makes them more efficient glycogen producers. In comparison, the higher insulin-dependent glycogen formation in K8/K18 IF-lacking hepatoma cells is associated with a signaling occurring through a mTOR-dependent pathway, along with an augmentation in cell proliferative activity. Together, the results uncover a key K8/K18 regulation of glucose metabolism in normal and cancerous hepatic cells through differential modulations of mitochondrial HK status and insulin-mediated signaling.« less

The role of the ubiquitin proteasome pathway in keratin intermediate filament protein degradation.

PubMed

Rogel, Micah R; Jaitovich, Ariel; Ridge, Karen M

2010-02-01

Lung injury, whether caused by hypoxic or mechanical stresses, elicits a variety of responses at the cellular level. Alveolar epithelial cells respond and adapt to such injurious stimuli by reorganizing the cellular cytoskeleton, mainly accomplished through modification of the intermediate filament (IF) network. The structural and mechanical integrity in epithelial cells is maintained through this adaptive reorganization response. Keratin, the predominant IF expressed in epithelial cells, displays highly dynamic properties in response to injury, sometimes in the form of degradation of the keratin IF network. Post-translational modification, such as phosphorylation, targets keratin proteins for degradation in these circumstances. As with other structural and regulatory proteins, turnover of keratin is regulated by the ubiquitin (Ub)-proteasome pathway. The degradation process begins with activation of Ub by the Ub-activating enzyme (E1), followed by the exchange of Ub to the Ub-conjugating enzyme (E2). E2 shuttles the Ub molecule to the substrate-specific Ub ligase (E3), which then delivers the Ub to the substrate protein, thereby targeting it for degradation. In some cases of injury and IF-related disease, aggresomes form in epithelial cells. The mechanisms that regulate aggresome formation are currently unknown, although proteasome overload may play a role. Therefore, a more complete understanding of keratin degradation--causes, mechanisms, and consequences--will allow for a greater understanding of epithelial cell biology and lung pathology alike.

Keratin sponge/hydrogel II, active agent delivery

USDA-ARS?s Scientific Manuscript database

Keratin sponge/hydrogels from oxidation and reduction hydrolysis of fine and coarse wool fibers were formed to behave as cationic hydrogels to swell and release active agents in the specific region of the gastro-intestinal (GI) tract. Their porous, interpenetrating networks (IPN) were effective for...

Immunohistochemical localization of thrombomodulin in the stratified epithelium of the rat is restricted to the keratinizing epidermis.

PubMed

Daimon, T; Nakano, M

1999-12-01

The expression and function of thrombomodulin (TM), an endothelial cofactor protein for thrombin-mediated protein C activation, in the epithelium are not fully characterized. This report describes the distribution and localization of TM in the various types of epithelia in the rat by light and electron microscopic immunocytochemistry. TM showed a limited distribution and was expressed by the keratinizing stratified epithelia of the skin, tongue, and esophagus, but was not present on the non-keratinizing epithelia of the vagina, ureter, trachea, stomach, or gut. An identical pattern of TM expression was seen in mucocutaneous junctions, transitional zones from a non-keratinizing stratified epithelium to a keratinizing epithelium at the edge of the eyelid and in the anal canal. As the keratinization of the stratified epithelia proceeded, the staining intensity increased in the transitional zones. Within the keratinizing stratified epithelia, TM staining was limited to the keratinocytes of the spinous layer, the spinous cells. The subcellular localization of TM on the spinous cells was restricted to the plasma membrane facing the intercellular spaces. TM was not detectable on the desmosomes or the two membranes making up the junction, presumably the nexus. The functional significance of TM in keratinizing epithelia is discussed.

Viscoelastic properties of α-keratin fibers in hair.

PubMed

Yu, Yang; Yang, Wen; André Meyers, Marc

2017-12-01

Considerable viscoelasticity and strain-rate sensitivity are a characteristic of α-keratin fibers, which can be considered a biopolymer. The understanding of viscoelasticity is an important part of the knowledge of the overall mechanical properties of these biological materials. Here, horse and human hairs are examined to analyze the sources of this response. The dynamic mechanical response of α-keratin fibers over a range of frequencies and temperatures is analyzed using a dynamic mechanical analyzer. The α-keratin fibers behave more elastically at higher frequencies while they become more viscous at higher temperatures. A glass transition temperature of ∼55°C is identified. The stress relaxation behavior of α-keratin fibers at two strains, 0.02 and 0.25, is established and fit to a constitutive equation based on the Maxwell-Wiechert model. The constitutive equation is further compared to the experimental results within the elastic region and a good agreement is obtained. The two relaxation constants, 14s and 359s for horse hair and 11s and 207s for human hair, are related to two hierarchical levels of relaxation: the amorphous matrix-intermediate filament interfaces, for the short term, and the cellular components for the long term. Results of the creep test also provide important knowledge on the uncoiling and phase transformation of the α-helical structure as hair is uniaxially stretched. SEM results show that horse hair has a rougher surface morphology and damaged cuticles. It also exhibits a lower strain-rate sensitivity of 0.05 compared to that of 0.11 for human hair. After the horse and human hairs are chemically treated and the disulfide bonds are cleaved, they exhibit a similar strain-rate sensitivity of ∼0.05. FTIR results confirms that the human hair is more sensitive to the -S-S- cleavage, resulting in an increase of cysteic acid content. Therefore, the disulfide bonds in the matrix are experimentally identified as one source of the strain

Structure and expression of genes for a class of cysteine-rich proteins of the cuticle layers of differentiating wool and hair follicles

PubMed Central

1990-01-01

The major histological components of the hair follicle are the hair cortex and cuticle. The hair cuticle cells encase and protect the cortex and undergo a different developmental program to that of the cortex. We report the molecular characterization of a set of evolutionarily conserved hair genes which are transcribed in the hair cuticle late in follicle development. Two genes were isolated and characterized, one expressed in the human follicle and one in the sheep follicle. Each gene encodes a small protein of 16 kD, containing greater than 50 cysteine residues, ranging from 31 to 36 mol% cysteine. Their high cysteine content and in vitro expression data identify them as ultra-high-sulfur (UHS) keratin proteins. The predicted proteins are composed almost entirely of cysteine-rich and glycine-rich repeats. Genomic blots reveal that the UHS keratin proteins are encoded by related multigene families in both the human and sheep genomes. Tissue in situ hybridization demonstrates that the expression of both genes is localized to the hair fiber cuticle and occurs at a late stage in fiber morphogenesis. PMID:1703541

Novel keratin modified bacterial cellulose nanocomposite production and characterization for skin tissue engineering.

PubMed

Keskin, Zalike; Sendemir Urkmez, Aylin; Hames, E Esin

2017-06-01

As it is known that bacterial cellulose (BC) is a biocompatible and natural biopolymer due to which it has a large set of biomedical applications. But still it lacks some desired properties, which limits its uses in many other applications. Therefore, the properties of BC need to be boosted up to an acceptable level. Here in this study for the first time, a new natural nanocomposite was produced by the incorporating keratin (isolated from human hair) to the BC (produced by Acetobacter xylinum) to enhance dermal fibroblast cells' attachment. Two different approaches were used in BC based nanocomposite production: in situ and post modifications. BC/keratin nanocomposites were characterized using SEM, FTIR, EDX, XRD, DSC and XPS analyses. Both production methods have yielded successful results for production of BC based nanocomposite-containing keratin. In vitro cell culture experiments performed with human skin keratinocytes and human skin fibroblast cells indicate the potential of the novel BC/keratin nanocomposites for use in skin tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

5S ribosomal RNA database Y2K

PubMed Central

Szymanski, Maciej; Barciszewska, Miroslawa Z.; Barciszewski, Jan; Erdmann, Volker A.

2000-01-01

This paper presents the updated version (Y2K) of the database of ribosomal 5S ribonucleic acids (5S rRNA) and their genes (5S rDNA), http://rose.man/poznan. pl/5SData/index.html . This edition of the database contains 1985 primary structures of 5S rRNA and 5S rDNA. They include 60 archaebacterial, 470 eubacterial, 63 plastid, nine mitochondrial and 1383 eukaryotic sequences. The nucleotide sequences of the 5S rRNAs or 5S rDNAs are divided according to the taxonomic position of the source organisms. PMID:10592212

5S ribosomal RNA database Y2K.

PubMed

Szymanski, M; Barciszewska, M Z; Barciszewski, J; Erdmann, V A

2000-01-01

This paper presents the updated version (Y2K) of the database of ribosomal 5S ribonucleic acids (5S rRNA) and their genes (5S rDNA), http://rose.man/poznan.pl/5SData/index.html. This edition of the database contains 1985primary structures of 5S rRNA and 5S rDNA. They include 60 archaebacterial, 470 eubacterial, 63 plastid, nine mitochondrial and 1383 eukaryotic sequences. The nucleotide sequences of the 5S rRNAs or 5S rDNAs are divided according to the taxonomic position of the source organisms.

Molecular evidence of keratin and melanosomes in feathers of the Early Cretaceous bird Eoconfuciusornis.

PubMed

Pan, Yanhong; Zheng, Wenxia; Moyer, Alison E; O'Connor, Jingmai K; Wang, Min; Zheng, Xiaoting; Wang, Xiaoli; Schroeter, Elena R; Zhou, Zhonghe; Schweitzer, Mary H

2016-12-06

Microbodies associated with feathers of both nonavian dinosaurs and early birds were first identified as bacteria but have been reinterpreted as melanosomes. Whereas melanosomes in modern feathers are always surrounded by and embedded in keratin, melanosomes embedded in keratin in fossils has not been demonstrated. Here we provide multiple independent molecular analyses of both microbodies and the associated matrix recovered from feathers of a new specimen of the basal bird Eoconfuciusornis from the Early Cretaceous Jehol Biota of China. Our work represents the oldest ultrastructural and immunological recognition of avian beta-keratin from an Early Cretaceous (∼130-Ma) bird. We apply immunogold to identify protein epitopes at high resolution, by localizing antibody-antigen complexes to specific fossil ultrastructures. Retention of original keratinous proteins in the matrix surrounding electron-opaque microbodies supports their assignment as melanosomes and adds to the criteria employable to distinguish melanosomes from microbial bodies. Our work sheds new light on molecular preservation within normally labile tissues preserved in fossils.

Molecular evidence of keratin and melanosomes in feathers of the Early Cretaceous bird Eoconfuciusornis

PubMed Central

Pan, Yanhong; Zheng, Wenxia; Moyer, Alison E.; O’Connor, Jingmai K.; Zheng, Xiaoting; Wang, Xiaoli; Schroeter, Elena R.; Zhou, Zhonghe; Schweitzer, Mary H.

2016-01-01

Microbodies associated with feathers of both nonavian dinosaurs and early birds were first identified as bacteria but have been reinterpreted as melanosomes. Whereas melanosomes in modern feathers are always surrounded by and embedded in keratin, melanosomes embedded in keratin in fossils has not been demonstrated. Here we provide multiple independent molecular analyses of both microbodies and the associated matrix recovered from feathers of a new specimen of the basal bird Eoconfuciusornis from the Early Cretaceous Jehol Biota of China. Our work represents the oldest ultrastructural and immunological recognition of avian beta-keratin from an Early Cretaceous (∼130-Ma) bird. We apply immunogold to identify protein epitopes at high resolution, by localizing antibody–antigen complexes to specific fossil ultrastructures. Retention of original keratinous proteins in the matrix surrounding electron-opaque microbodies supports their assignment as melanosomes and adds to the criteria employable to distinguish melanosomes from microbial bodies. Our work sheds new light on molecular preservation within normally labile tissues preserved in fossils. PMID:27872291

Pachyonychia Congenita (K16) with Unusual Features and Good Response to Acitretin

PubMed Central

Almutawa, Fahad; Thusaringam, Thusanth; Watters, Kevin; Gayden, Tenzin; Jabado, Nada; Sasseville, Denis

2015-01-01

Background Pachyonychia congenita (PC) is a rare autosomal dominant disease whose main clinical features include hypertrophic onychodystrophy and palmoplantar keratoderma. The new classification is based on genetic variants with mutations in keratin KRT6A, KRT6B, KRT6C, KRT16, KRT17, and an unknown mutation. Here, we present a case of PC with unusual clinical and histological features and a favorable response to oral acitretin. Case A 49-year-old male presented with diffuse and striate palmoplantar keratoderma, thickened nails, knuckle pads, and pseudoainhum. Histology showed compact hyperkeratosis, prominent irregular acanthosis, and extensive epidermolytic hyperkeratosis, suggestive of Vörner's palmoplantar keratoderma. However, keratin 9 and 1 were not mutated, and full exome sequencing showed heterozygous missense mutation in type I keratin K16. Conclusion To our knowledge, epidermolytic hyperkeratosis has not been previously described with PC. Our patient had an excellent response, maintained over the last 5 years, to a low dose of acitretin. We wish to emphasize the crucial role of whole exome sequencing in establishing the correct diagnosis. PMID:26464567

GCN5 regulates the activation of PI3K/Akt survival pathway in B cells exposed to oxidative stress via controlling gene expressions of Syk and Btk.

PubMed

Kikuchi, Hidehiko; Kuribayashi, Futoshi; Takami, Yasunari; Imajoh-Ohmi, Shinobu; Nakayama, Tatsuo

2011-02-25

Histone acetyltransferase(s) (HATs) are involved in the acetylation of core histones, which is an important event for transcription regulation through alterations in the chromatin structure in eukaryotes. General control non-depressible 5 (GCN5) was first identified as a global coactivator and transcription-related HAT. Here we report that GCN5 regulates the activation of phosphatidylinositol 3-kinase (PI3K)/acutely transforming retrovirus AKT8 in rodent T cell lymphoma (Akt) survival pathway in B cells exposed to oxidative stress via controlling gene expressions of spleen tyrosine kinase (Syk) and Bruton's tyrosine kinase (Btk). The GCN5-deficiency remarkably caused apoptotic cell death by treatment with exogenous hydrogen peroxide (H(2)O(2)) in chicken DT40 cells. In GCN5-deficient DT40 cells, gene expressions of Syk and Btk, which are involved in activation of PI3K/Akt survival pathway in DT40 cells exposed to exogenous H(2)O(2), were remarkably decreased compared with those in wild type DT40 cells. In addition, phosphorylation of Akt in H(2)O(2)-treated GCN5-deficient cells was remarkably suppressed as compared to that of DT40. Chromatin immunoprecipitation assay revealed that GCN5 binds to proximal 5'-upstream regions of Syk and Btk genes in vivo. These results suggest that GCN5 takes part in transcriptional regulations of the Syk and Btk genes, and plays a key role in epigenetic regulation of PI3K/Akt survival pathway in B cells exposed to reactive oxygen species such as H(2)O(2). Copyright © 2011 Elsevier Inc. All rights reserved.

Myelin management by the 18.5–kDa and 21.5–kDa classic myelin basic protein isoforms

PubMed Central

Harauz, George; Boggs, Joan M.

2013-01-01

The classic myelin basic protein (MBP) splice isoforms range in nominal molecular mass from 14 to 21.5 kDa, and arise from the gene in the oligodendrocyte lineage (Golli) in maturing oligodendrocytes. The 18.5-kDa isoform that predominates in adult myelin adheres the cytosolic surfaces of oligodendrocyte membranes together, and forms a two-dimensional molecular sieve restricting protein diffusion into compact myelin. However, this protein has additional roles including cytoskeletal assembly and membrane extension, binding to SH3-domains, participation in Fyn-mediated signaling pathways, sequestration of phosphoinositides, and maintenance of calcium homeostasis. Of the diverse post-translational modifications of this isoform, phosphorylation is the most dynamic, and modulates 18.5-kDa MBP’s protein-membrane and protein-protein interactions, indicative of a rich repertoire of functions. In developing and mature myelin, phosphorylation can result in microdomain or even nuclear targeting of the protein, supporting the conclusion that 18.5-kDa MBP has significant roles beyond membrane adhesion. The full-length, early-developmental 21.5-kDa splice isoform is predominantly karyophilic due to a non-traditional P-Y nuclear localization signal, with effects such as promotion of oligodendrocyte proliferation. We discuss in vitro and recent in vivo evidence for multifunctionality of these classic basic proteins of myelin, and argue for a systematic evaluation of the temporal and spatial distributions of these protein isoforms, and their modified variants, during oligodendrocyte differentiation. PMID:23398367

A functional promoter shift of a chloroplast gene: a transcriptional fusion between a novel psbA gene copy and the trnK (UUU) gene in Pinus contorta.

PubMed

Lidholm, J; Gustafsson, P

1992-11-01

A comparative transcription analysis of the chloroplast trnK-psbA-trnH region of the two pine species Pinus contorta and Pinus sylvestris is reported. The chloroplast genome of P. contorta has previously been shown to contain a duplicated psbA gene copy integrated closely upstream of the split trnK gene. This rearrangement has resulted in the gene order psbAI-trnK-psbAII-trnH, where psbAII is the ancestral psbA gene copy. In P. sylvestris, a species which lacks the psbA duplication, transcription of the trnK gene originates from a position 291 bp upstream of the trnK 5' exon, adjacent to a canonical promoter structure. In P. contorta, the corresponding promoter structure has been separated from the trnK gene by the insertion of psbAI, and has, in addition, been partially deleted. Analysis of the transcriptional organization of the trnK-psbA-trnH region of the two pine species revealed that the trnK gene in P. contorta is transcriptionally fused to the inserted psbAI gene copy. As a result, trnK is under the control of the psbA promoter in this species and has therefore acquired psbA-like expression characteristics. In P. sylvestris, accumulation of trnK transcripts is not significantly higher in light-grown than in dark-grown seedlings. In contrast, the level of trnK transcripts in P. contorta is approximately 12-fold higher in the light than in the dark. When light-grown seedlings of the two pine species were compared, an approximately 20-fold higher level of trnK RNAs was found in P. contorta. In both pine species, evidence was obtained for trnK-psbA and psbA-trnH co-transcription.

Epithelial estrogen receptor 1 intrinsically mediates squamous differentiation in the mouse vagina

PubMed Central

Miyagawa, Shinichi; Iguchi, Taisen

2015-01-01

Estrogen-mediated actions in female reproductive organs are tightly regulated, mainly through estrogen receptor 1 (ESR1). The mouse vaginal epithelium cyclically exhibits cell proliferation and differentiation in response to estrogen and provides a unique model for analyzing the homeostasis of stratified squamous epithelia. To address the role of ESR1-mediated tissue events during homeostasis, we analyzed mice with a vaginal epithelium-specific knockout of Esr1 driven by keratin 5-Cre (K5-Esr1KO). We show here that loss of epithelial ESR1 in the vagina resulted in aberrant epithelial cell proliferation in the suprabasal cell layers and led to failure of keratinized differentiation. Gene expression analysis showed that several known estrogen target genes, including erbB growth factor ligands, were not induced by estrogen in the K5-Esr1KO mouse vagina. Organ culture experiments revealed that the addition of erbB growth factor ligands, such as amphiregulin, could activate keratinized differentiation in the absence of epithelial ESR1. Thus, epithelial ESR1 integrates estrogen and growth factor signaling to mediate regulation of cell proliferation in squamous differentiation, and our results provide new insights into estrogen-mediated homeostasis in female reproductive organs. PMID:26438838

Keratin decomposition by trogid beetles: evidence from a feeding experiment and stable isotope analysis

NASA Astrophysics Data System (ADS)

Sugiura, Shinji; Ikeda, Hiroshi

2014-03-01

The decomposition of vertebrate carcasses is an important ecosystem function. Soft tissues of dead vertebrates are rapidly decomposed by diverse animals. However, decomposition of hard tissues such as hairs and feathers is much slower because only a few animals can digest keratin, a protein that is concentrated in hairs and feathers. Although beetles of the family Trogidae are considered keratin feeders, their ecological function has rarely been explored. Here, we investigated the keratin-decomposition function of trogid beetles in heron-breeding colonies where keratin was frequently supplied as feathers. Three trogid species were collected from the colonies and observed feeding on heron feathers under laboratory conditions. We also measured the nitrogen (δ15N) and carbon (δ13C) stable isotope ratios of two trogid species that were maintained on a constant diet (feathers from one heron individual) during 70 days under laboratory conditions. We compared the isotopic signatures of the trogids with the feathers to investigate isotopic shifts from the feathers to the consumers for δ15N and δ13C. We used mixing models (MixSIR and SIAR) to estimate the main diets of individual field-collected trogid beetles. The analysis indicated that heron feathers were more important as food for trogid beetles than were soft tissues under field conditions. Together, the feeding experiment and stable isotope analysis provided strong evidence of keratin decomposition by trogid beetles.

Cellulose nanocrystal-reinforced keratin bioadsorbent for effective removal of dyes from aqueous solution.

PubMed

Song, Kaili; Xu, Helan; Xu, Lan; Xie, Kongliang; Yang, Yiqi

2017-05-01

High-efficiency and recyclable three-dimensional bioadsorbents were prepared by incorporating cellulose nanocrystal (CNC) as reinforcements in keratin sponge matrix to remove dyes from aqueous solution. Adsorption performance of dyes by CNC-reinforced keratin bioadsorbent was improved significantly as a result of adding CNC as filler. Batch adsorption results showed that the adsorption capacities for Reactive Black 5 and Direct Red 80 by the bioadsorbent were 1201 and 1070mgg -1 , respectively. The isotherms and kinetics for adsorption of both dyes on bioadsorbent followed the Langmuir isotherm model and pseudo-second order model, respectively. Desorption and regeneration experiments showed that the removal efficiencies of the bioadsorbent for both dyes could remain above 80% at the fifth recycling cycles. Moreover, the bioadsorbent possessed excellent packed-bed column operation performance. Those results suggested that the adsorbent could be considered as a high-performance and promising candidate for dye wastewater treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

Evaluation of the efficacy of keratinized mucosa augmentation techniques around dental implants: a systematic review.

PubMed

Wu, Qingqing; Qu, Yili; Gong, Ping; Wang, Tianlu; Gong, Ting; Man, Yi

2015-05-01

The absence of periimplant keratinized mucosa is considered risky in patients with a predisposition to periodontitis or recession. Although various soft tissue augmentation techniques exist, dentists are seeking for more efficient approaches to augment periimplant keratinized mucosa. The purpose of this systematic review was to evaluate the efficacy of the various techniques and biomaterials adopted in periimplant keratinized mucosa augmentation and whether one technique or biomaterial is superior. A search in Medline-PubMed and the Cochrane Central Register of controlled trials was conducted. Randomized clinical trials, prospective cohort studies, clinical control studies, and case series from January 1, 1980, to December 31, 2013, with a follow-up of at least 6 months reporting changes on keratinized mucosa width were included. Several journals were hand-searched for related articles. The bibliographies of all publications selected for inclusion were also scanned. The screening of titles and abstracts resulted in 60 relevant publications. Six of them were finally included. Free gingival graft, connective tissue graft, acellular dermal matrix, and collagen matrix were used for keratinized mucosa augmentation. Because of heterogeneity of the studies, only descriptive analysis was performed. Improvements in keratinized mucosa width were reported in all studies. A definitive conclusion could not be achieved owing to the lack of well-designed studies and appropriate methods of studying soft tissue. The establishment of universal surgical guidelines and measurement systems is imperative in the future. Copyright © 2015 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.

Modifying surface resistivity and liquid moisture management property of keratin fibers through thiol-ene click reactions.

PubMed

Yu, Dan; Cai, Jackie Y; Church, Jeffrey S; Wang, Lijing

2014-01-22

This paper reports on a new method for improving the antistatic and liquid moisture management properties of keratinous materials. The method involves the generation of thiols by controlled reduction of cystine disulfide bonds in keratin with tris(2-carboxyethyl) phosphine hydrochloride and subsequent grafting of hydrophilic groups onto the reduced keratin by reaction with an acrylate sulfonate or acrylamide sulfonate through thiol-ene click chemistry. The modified substrates were characterized with Raman spectroscopy and scanning electron microscopy and evaluated for their performance changes in liquid moisture management, surface resistivity, and wet burst strength. The results have revealed that the thiol-acrylate reaction is more efficient than the thiol-acrylamide reaction, and the keratinous substrate modified with an acrylate sulfonate salt exhibits significantly improved antistatic and liquid moisture management properties.

Free Gingival Graft to Increase Keratinized Mucosa after Placing of Mandibular Fixed Implant-Supported Prosthesis

PubMed Central

Marcantonio, Elcio

2017-01-01

Insufficiently keratinized tissue can be increased surgically by free gingival grafting. The presence or reconstruction of keratinized mucosa around the implant can facilitate restorative procedure and allow the maintenance of an oral hygiene routine without irritation or discomfort to the patient. The aim of this clinical case report is to describe an oral rehabilitation procedure of an edentulous patient with absence of keratinized mucosa in the interforaminal area, using a free gingival graft associated with a mandibular fixed implant-supported prosthesis. The treatment included the manufacturing of a maxillary complete denture and a mandibular fixed implant-supported prosthesis followed by a free gingival graft to increase the width of the mandibular keratinized mucosa. Free gingival graft was obtained from the palate and grafted on the buccal side of interforaminal area. The follow-up of 02 and 12 months after mucogingival surgery showed that the free gingival graft promoted peri-implant health, hygiene, and patient comfort. Clinical Significance. The free gingival graft is an effective treatment in increasing the width of mandibular keratinized mucosa on the buccal side of the interforaminal area and provided an improvement in maintaining the health of peri-implant tissues which allows for better oral hygiene. PMID:28293441

A phylogeny of cycads (Cycadales) inferred from chloroplast matK gene, trnK intron, and nuclear rDNA ITS region.

PubMed

Chaw, Shu-Miaw; Walters, Terrence W; Chang, Chien-Chang; Hu, Shu-Hsuan; Chen, Shin-Hsiao

2005-10-01

Phylogenetic relationships among the three families and 12 living genera of cycads were reconstructed by distance and parsimony criteria using three markers: the chloroplast matK gene, the chloroplast trnK intron and the nuclear ITS/5.8S rDNA sequence. All datasets indicate that Cycadaceae (including only the genus Cycas) is remotely related to other cycads, in which Dioon was resolved as the basal-most clade, followed by Bowenia and a clade containing the remaining nine genera. Encephalartos and Lepidozamia are closer to each other than to Macrozamia. The African genus Stangeria is embedded within the New World subfamily Zamiodeae. Therefore, Bowenia is an unlikely sister to Stangeria, contrary to the view that they form the Stangeriaceae. The generic status of Dyerocycas and Chigua is unsupportable as they are paraphyletic with Cycas and the Zamia, respectively. Nonsense mutations in the matK gene and indels in the other two datasets lend evidence to reinforce the above conclusions. According to the phylogenies, the past geography of the genera of cycads and the evolution of character states are hypothesized and discussed. Within the suborder Zamiieae, Stangeria, and the tribe Zamieae evolved significantly faster than other genera. The matK gene and ITS/5.8S region contain more useful information than the trnK intron in addressing phylogeny. Redelimitations of Zamiaceae, Stangeriaceae, subfamily Encephalartoideae and subtribe Macrozamiineae are necessary.

Identification of essential genes and synthetic lethal gene combinations in Escherichia coli K-12.

PubMed

Mori, Hirotada; Baba, Tomoya; Yokoyama, Katsushi; Takeuchi, Rikiya; Nomura, Wataru; Makishi, Kazuichi; Otsuka, Yuta; Dose, Hitomi; Wanner, Barry L

2015-01-01

Here we describe the systematic identification of single genes and gene pairs, whose knockout causes lethality in Escherichia coli K-12. During construction of precise single-gene knockout library of E. coli K-12, we identified 328 essential gene candidates for growth in complex (LB) medium. Upon establishment of the Keio single-gene deletion library, we undertook the development of the ASKA single-gene deletion library carrying a different antibiotic resistance. In addition, we developed tools for identification of synthetic lethal gene combinations by systematic construction of double-gene knockout mutants. We introduce these methods herein.

Gene profiling of cathepsin K deficiency in atherogenesis: profibrotic but lipogenic.

PubMed

Lutgens, S P M; Kisters, N; Lutgens, E; van Haaften, R I M; Evelo, C T A; de Winther, M P J; Saftig, P; Daemen, M J A P; Heeneman, S; Cleutjens, K B J M

2006-11-01

Recently, we showed that cathepsin K deficiency reduces atherosclerotic plaque progression, induces plaque fibrosis, but aggravates macrophage foam cell formation in the ApoE -/- mouse. To obtain more insight into the molecular mechanisms by which cathepsin K disruption evokes the observed phenotypic changes, we used microarray analysis for gene expression profiling of aortic arches of CatK -/-/ApoE -/- and ApoE -/- mice on a mouse oligo microarray. Out of 20 280 reporters, 444 were significantly differentially expressed (p-value of < 0.05, fold change of > or = 1.4 or < or = - 1.4, and intensity value of > 2.5 times background in at least one channel). Ingenuity Pathway Analysis and GenMAPP revealed upregulation of genes involved in lipid uptake, trafficking, and intracellular storage, including caveolin - 1, - 2, - 3 and CD36, and profibrotic genes involved in transforming growth factor beta (TGFbeta) signalling, including TGFbeta2, latent TGFbeta binding protein-1 (LTBP1), and secreted protein, acidic and rich in cysteine (SPARC), in CatK -/-/ApoE -/- mice. Differential gene expression was confirmed at the mRNA and protein levels. In vitro modified low density lipoprotein (LDL) uptake assays, using bone marrow derived macrophages preincubated with caveolae and scavenger receptor inhibitors, confirmed the importance of caveolins and CD36 in increasing modified LDL uptake in the absence of cathepsin K. In conclusion, we suggest that cathepsin K deficiency alters plaque phenotype not only by decreasing proteolytic activity, but also by stimulating TGFbeta signalling. Besides this profibrotic effect, cathepsin K deficiency has a lipogenic effect owing to increased lipid uptake mediated by CD36 and caveolins. Copyright 2006 Pathological Society of Great Britain and Ireland.

The amelioration of cardiac dysfunction after myocardial infarction by the injection of keratin biomaterials derived from human hair.

PubMed

Shen, Deliang; Wang, Xiaofang; Zhang, Li; Zhao, Xiaoyan; Li, Jingyi; Cheng, Ke; Zhang, Jinying

2011-12-01

Cardiac dysfunction following acute myocardial infarction is a major cause of advanced cardiomyopathy. Conventional pharmacological therapies rely on prompt reperfusion and prevention of repetitive maladaptive pathways. Keratin biomaterials can be manufactured in an autologous fashion and are effective in various models of tissue regeneration. However, its potential application in cardiac regeneration has not been tested. Keratin biomaterials were derived from human hair and its structure morphology, carryover of beneficial factors, biocompatibility with cardiomyocytes, and in vivo degradation profile were characterized. After delivery into infarcted rat hearts, the keratin scaffolds were efficiently infiltrated by cardiomyocytes and endothelial cells. Injection of keratin biomaterials promotes angiogenesis but does not exacerbate inflammation in the post-MI hearts. Compared to control-injected animals, keratin biomaterials-injected animals exhibited preservation of cardiac function and attenuation of adverse ventricular remodeling over the 8 week following time course. Tissue western blot analysis revealed up-regulation of beneficial factors (BMP4, NGF, TGF-beta) in the keratin-injected hearts. The salient functional benefits, the simplicity of manufacturing and the potentially autologous nature of this biomaterial provide impetus for further translation to the clinic. Copyright © 2011 Elsevier Ltd. All rights reserved.

Keratin Durability Has Implications for the Fossil Record: Results from a 10 Year Feather Degradation Experiment

PubMed Central

Moyer, Alison E.; Zheng, Wenxia; Schweitzer, Mary H.

2016-01-01

Keratinous ‘soft tissue’ structures (i.e. epidermally derived and originally non-biomineralized), include feathers, skin, claws, beaks, and hair. Despite their relatively common occurrence in the fossil record (second only to bone and teeth), few studies have addressed natural degradation processes that must occur in all organic material, including those keratinous structures that are incorporated into the rock record as fossils. Because feathers have high preservation potential and strong phylogenetic signal, in the current study we examine feathers subjected to different burial environments for a duration of ~10 years, using transmission electron microscopy (TEM) and in situ immunofluorescence (IF). We use morphology and persistence of specific immunoreactivity as indicators of preservation at the molecular and microstructural levels. We show that feather keratin is durable, demonstrates structural and microstructural integrity, and retains epitopes suitable for specific antibody recognition in even the harshest conditions. These data support the hypothesis that keratin antibody reactivity can be used to identify the nature and composition of epidermal structures in the rock record, and to address evolutionary questions by distinguishing between alpha- (widely distributed) and beta- (limited to sauropsids) keratin. PMID:27384819

Effect of clay content on morphology and processability of electrospun keratin/poly(lactic acid) nanofiber.

PubMed

Isarankura Na Ayutthaya, Siriorn; Tanpichai, Supachok; Sangkhun, Weradesh; Wootthikanokkhan, Jatuphorn

2016-04-01

This research work has concerned the development of volatile organic compounds (VOCs) removal filters from biomaterials, based on keratin extracted from chicken feather waste and poly(lactic acid) (PLA) (50/50%w/w) blend. Clay (Na-montmorillonite) was also added to the blend solution prior to carrying out an electro-spinning process. The aim of this study was to investigate the effect of clay content on viscosity, conductivity, and morphology of the electrospun fibers. Scanning electron micrographs showed that smooth and bead-free fibers were obtained when clay content used was below 2 pph. XRD patterns of the electrospun fibers indicated that the clay was intercalated and exfoliated within the polymers matrix. Percentage crystallinity of keratin in the blend increased after adding the clay, as evidenced from FTIR spectra and DSC thermograms. Transmission electron micrographs revealed a kind of core-shell structure with clay being predominately resided within the keratin rich shell and at the interfacial region. Filtration performance of the electrospun keratin/PLA fibers, described in terms of pressure drop and its capability of removing methylene blue, were also explored. Overall, our results demonstrated that it was possible to improve process-ability, morphology and filtration efficiency of the electrospun keratin fibers by adding a suitable amount of clay. Copyright © 2016 Elsevier B.V. All rights reserved.

Moisture, anisotropy, stress state, and strain rate effects on bighorn sheep horn keratin mechanical properties

DOE PAGES

Johnson, K. L.; Trim, M. W.; Francis, D. K.; ...

2016-10-01

Our paper investigates the effects of moisture, anisotropy, stress state, and strain rate on the mechanical properties of the bighorn sheep (Ovis Canadensis) horn keratin. The horns consist of fibrous keratin tubules extending along the length of the horn and are contained within an amorphous keratin matrix. We tested samples in the rehydrated (35 wt.% water) and ambient dry (10 wt.% water) conditions along the longitudinal and radial directions under tension and compression. Increased moisture content was found to increase ductility and decrease strength, as well as alter the stress state dependent nature of the material. Furthermore, the horn keratinmore » demonstrates a significant strain rate dependence in both tension and compression, and also showed increased energy absorption in the hydrated condition at high strain rates when compared to quasi-static data, with increases of 114% in tension and 192% in compression. Compressive failure occurred by lamellar buckling in the longitudinal orientation followed by shear delamination. Tensile failure in the longitudinal orientation occurred by lamellar delamination combined with tubule pullout and fracture. Finally, the structure-property relationships quantified here for bighorn sheep horn keratin can be used to help validate finite element simulations of ram’s impacting each other as well as being useful for other analysis regarding horn keratin on other animals.« less

Moisture, anisotropy, stress state, and strain rate effects on bighorn sheep horn keratin mechanical properties

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, K. L.; Trim, M. W.; Francis, D. K.

Our paper investigates the effects of moisture, anisotropy, stress state, and strain rate on the mechanical properties of the bighorn sheep (Ovis Canadensis) horn keratin. The horns consist of fibrous keratin tubules extending along the length of the horn and are contained within an amorphous keratin matrix. We tested samples in the rehydrated (35 wt.% water) and ambient dry (10 wt.% water) conditions along the longitudinal and radial directions under tension and compression. Increased moisture content was found to increase ductility and decrease strength, as well as alter the stress state dependent nature of the material. Furthermore, the horn keratinmore » demonstrates a significant strain rate dependence in both tension and compression, and also showed increased energy absorption in the hydrated condition at high strain rates when compared to quasi-static data, with increases of 114% in tension and 192% in compression. Compressive failure occurred by lamellar buckling in the longitudinal orientation followed by shear delamination. Tensile failure in the longitudinal orientation occurred by lamellar delamination combined with tubule pullout and fracture. Finally, the structure-property relationships quantified here for bighorn sheep horn keratin can be used to help validate finite element simulations of ram’s impacting each other as well as being useful for other analysis regarding horn keratin on other animals.« less

First detection of the staphylococcal trimethoprim resistance gene dfrK and the dfrK-carrying transposon Tn559 in enterococci.

PubMed

López, María; Kadlec, Kristina; Schwarz, Stefan; Torres, Carmen

2012-02-01

The trimethoprim resistance gene dfrK has been recently described in Staphylococcus aureus, but so far has not been found in other bacteria. A total of 166 enterococci of different species (E. faecium, E. faecalis, E. hirae, E. durans, E. gallinarum, and E. casseliflavus) and origins (food, clinical diseases in humans, healthy humans or animals, and sewage) were studied for their susceptibility to trimethoprim as determined by agar dilution (European Committee on Antimicrobial Susceptibility Testing) and the presence of (a) the dfrK gene and its genetic environment and (b) other dfr genes. The dfrK gene was detected in 49% of the enterococci (64% and 42% of isolates with minimum inhibitory concentrations of ≥2 mg/L or ≤1 mg/L, respectively). The tet(L)-dfrK linkage was detected in 21% of dfrK-positive enterococci. The chromosomal location of the dfrK gene was identified in one E. faecium isolate in which the dfrK was not linked to tet(L) gene but was part of a Tn559 element, which was integrated in the chromosomal radC gene. This Tn559 element was also found in 14 additional isolates. All combinations of dfr genes were detected among the isolates tested (dfrK, dfrG, dfrF, dfrK+dfrG, dfrK+dfrF, dfrF+dfrG, and dfrF+dfrG+dfrK). The gene dfrK gene was found together with other dfr genes in 58% of the tested enterococci. This study suggested an exchange of the trimethoprim resistance gene dfrK between enterococci and staphylococci, as previously observed for the trimethoprim resistance gene dfrG.

Keratin pattern of acanthosis nigricans in syndromelike association with polythelia, polycystic kidneys, and syndactyly.

PubMed

Bonnekoh, B; Wevers, A; Spangenberger, H; Mahrle, G; Krieg, T

1993-09-01

Acanthosis nigricans (AN) comprises a broad spectrum of etiologic subtypes. The underlying pathomechanisms have not yet been completely clarified. We present a patient affected with a syndromelike AN subtype including disturbed epidermopoiesis as evidenced by immunohistologic findings and in situ hybridization. A 54-year-old white man contracted AN during childhood. There were connate malformations consisting of webbed toes II/III on the right side and a supernumerary left mammilla. As an adult he developed psoriasis vulgaris, obesity, and latent diabetes mellitus, polycystic kidney and liver disease. With regard to keratin 6 mRNA, and the protein expression of keratin 6/16, KI-67, and proliferating cell nuclear antigen, the AN lesion showed moderate hyperproliferation. A much higher degree of hyperproliferation was evident in psoriatic areas of the patient's skin. In contrast to psoriatic tissue, basal keratinocytes of the AN showed an unusually high expression of keratin 18 and 19 protein. The observation thus deals with a unique, syndromelike constellation of AN characterized by a particular epidermal pattern of moderate hyperproliferation. A further dysregulation of protein expression in the epidermis is indicated by the demonstration of the rare keratins 18 and 19 in basal keratinocytes of the AN lesion.

Enucleation of facial sebaceous cyst by creating a minimal elliptical incision through a keratin-filled orifice.

PubMed

Chen, Wei-Liang

2016-12-01

A facial sebaceous cyst is a common benign epithelial tumor and surgical excision is frequently performed but may cause obvious scarring and may be esthetically troubling. This study evaluated the clinical outcomes of the patients with facial sebaceous cyst enucleated by creating minimal elliptical incisions through a keratin-filled orifice. Eleven patients with facial sebaceous cyst enucleated by creating minimal elliptical incisions through a keratin-filled orifice. We treated nine male and two female patients aged 25-52 years. The mean cyst size was 1.85 × 1.56 cm. All cysts were successfully enucleated. The mean wound length was 0.93 cm. The mean operative time was 15.2 min. The mean follow-up duration was 41.5 months. No recurrence was noted, and all patients were very satisfied with their esthetic outcomes. All cysts were successfully enucleated. The mean elliptical wound length was 0.93 cm (range, 0.8-1.1 cm). The mean operative time was 15.2 min. We found no evidence of wound infection, or nerve or vascular injury. Enucleation of facial sebaceous cyst via a minimal elliptical incision through the keratin-filled orifice was associated with high-level patient satisfaction, and the method is safe and useful for treating facial epidermoid cysts. © 2016 Wiley Periodicals, Inc.

Evidence of accelerated beak growth associated with avian keratin disorder in black-capped chickadees (Poecile atricapillus)

USGS Publications Warehouse

Van Hemert, Caroline R.; Handel, Colleen M.; O'Hara, Todd M.

2012-01-01

We recently documented an epizootic of beak deformities in more than 2,000 Blackcapped Chickadees (Poecile atricapillus) and other wild bird species in North America. This emerging avian disease, which has been termed avian keratin disorder, results in gross overgrowth of the rhamphotheca, the outer, keratinized layer of the beak. To test the hypothesis that the beak deformities characteristic of this disorder are associated with accelerated keratin production, we measured rates of beak growth and wear in affected Black-capped Chickadees (n=16) and a control sample of unaffected chickadees (n=14) collected from south-central (61°09'-61°38'N, 149°11' -149°48'W) and interior Alaska (64°51' -64°53'N, 147°49' -147°59'W). Rates of absolute growth were 50-100% higher in affected birds than they were in control birds and exceeded records from other passerine species. These results suggest that abnormally rapid epidermal growth is the primary physical mechanism by which beak deformities develop and are maintained in affected chickadees. Although beak overgrowth typically worsened over time, differential patterns of wear influenced the severity and morphology of deformities. In some cases, the effects of accelerated keratin growth were partially mitigated by frequent breakage of rhamphothecal tips. However, mortalities occurred in 9 of 16 birds (56%) with beak deformities during the study, suggesting that avian keratin disorder results in severe health consequences for affected birds. Additional study of factors that control beak keratin production is needed to understand the pathogenesis of this debilitating disease in wild birds.

Development of chitosan/gelatin/keratin composite containing hydrocortisone sodium succinate as a buccal mucoadhesive patch to treat desquamative gingivitis.

PubMed

Davoudi, Zahra; Rabiee, Mohammad; Houshmand, Behzad; Eslahi, Niloofar; Khoshroo, Kimia; Rasoulianboroujeni, Morteza; Tahriri, Mohammadreza; Tayebi, Lobat

2018-01-01

The aim of this research was to develop chitosan/gelatin/keratin composite containing hydrocortisone sodium succinate as a buccal mucoadhesive patch to treat desquamative gingivitis, which was fabricated through an environmental friendly process. Mucoadhesive films increase the advantage of higher efficiency and drug localization in the affected region. In this research, mucoadhesive films, for the release of hydrocortisone sodium succinate, were prepared using different ratios of chitosan, gelatin and keratin. In the first step, chitosan and gelatin proportions were optimized after evaluating the mechanical properties, swelling capacity, water uptake, stability, and biodegradation of the films. Then, keratin was added at different percentages to the optimum composite of chitosan and gelatin together with the drug. The results of surface pH showed that none of the samples were harmful to the buccal cavity. FTIR analysis confirmed the influence of keratin on the structure of the composite. The presence of a higher amount of keratin in the composite films resulted in high mechanical, mucoadhesive properties and stability, low water uptake and biodegradation in phosphate buffer saline (pH = 7.4) containing 10 4  U/ml lysozyme. The release profile of the films ascertained that keratin is a rate controller in the release of the hydrocortisone sodium succinate. Finally, chitosan/gelatin/keratin composite containing hydrocortisone sodium succinate can be employed in dental applications.

Differential expression of Cyclin D1 in keratin-producing odontogenic cysts

PubMed Central

Vera-Sirera, Beatriz; Forner-Navarro, Leopoldo

2015-01-01

Objetives: The aim of the present study was to analyze the expression levels of Cyclin D1 (CCD1), a nuclear protein that plays a crucial role in cell cycle progression, in a series of keratin-producing odontogenic cysts. Study Design: A total of 58 keratin-producing odontogenic cysts, diagnosed over ten years and classified according to the WHO 2005 criteria, were immunohistochemically analyzed in terms of CCD1 expression, which was quantified in the basal, suprabasal and intermediate/superficial epithelial compartments. The extent of immunostaining was measured as a proportion of total epithelial thickness. Quantified immunohistochemical data were correlated with clinicopathological features and clinical recurrence. Results: Keratin-producing odontogenic cysts were classified as 6 syndromic keratocystic odontogenic tumors (S-KCOT), 40 sporadic or non-syndromic KCOT (NS-KCOT) and 12 orthokeratinized odontogenic cysts (OOC). Immunohistochemically, CCD1 staining was evident predominantly in the parabasal region of all cystic lesions, but among-lesion differences were apparent, showing a clear expansion of parabasal compartment especially in the S-KCOT, followed to a lesser extent in the NS-KCOT, and being much more reduced in the OOC, which had the greatest average epithelial thickness. Conclusions: The differential expression of CCD1 noted in the present study suggests that dysregulation of cell cycle progression from G1 to the S phase contributes to the different aggressiveness of these lesions. However, CCD1 expression levels did not predict NS-KCOT recurrence, which is likely influenced by factors unrelated to lesion biology. Key words:Keratin-producing odontogenic cyst, keratocyst, keratocystic odontogenic tumor, nevoid basal cell carcinoma syndrome, orthokeratinized odontogenic cyst, cyclin D1, immunohistochemistry. PMID:25475773

Presence of keratin-specific antibody-forming cells in palatine tonsils of patients with pustulosis palmaris et plantaris (PPP) and its correlation with prognosis after tonsillectomy.

PubMed

Tanimoto, Yoichiro; Fukuyama, Satoshi; Tanaka, Norimitsu; Ohori, Jun-Ichiro; Tanimoto, Yukari; Kurono, Yuichi

2014-01-01

Keratin-specific immune responses in tonsils may be associated with the pathogenesis of pustulosis palmaris et plantaris (PPP). Evaluation of keratin-specific immune responses in tonsils might be useful to predict the effectiveness of tonsillectomy for patients with PPP. The aim of the present study was to clarify the role of keratin-specific immune responses in the pathogenesis of PPP in tonsils. It has been reported that anti-keratin antibodies in serum were higher in patients with PPP and decreased after tonsillectomy, indicating that anti-keratin antibodies might be generated in tonsils. In order to demonstrate the presence of keratin-specific immune responses in tonsils, the numbers of keratin-specific antibody-forming cells (AFCs) in tonsillar and peripheral blood lymphocytes were examined by enzyme-linked immunospot assay. The prognosis of PPP was compared after tonsillectomy. The numbers of keratin-specific IgM and IgG AFCs in tonsils and of IgG AFCs in peripheral blood were significantly increased in patients with PPP. The numbers of keratin-specific IgG AFCs in peripheral blood correlated positively with tonsil and serum IgG antibodies specific to keratin. Our data show that a good prognosis in patients with PPP depended on the numbers of keratin-specific IgG and IgM AFCs in peripheral blood and the levels of keratin-specific IgG antibodies in serum being significantly decreased 6 months after tonsillectomy.

Giant axonal neuropathy alters the structure of keratin intermediate filaments in human hair

PubMed Central

Soomro, Asfia; Alsop, Richard J.; Negishi, Atsuko; Kreplak, Laurent; Fudge, Douglas; Kuczmarski, Edward R.; Goldman, Robert D.

2017-01-01

Giant axonal neuropathy (GAN) follows an autosomal recessive genetic inheritance and impedes the peripheral and central nervous system due to axonal swellings that are packed with neurofilaments. The patients display a number of phenotypes, including hypotonia, muscle weakness, decreased reflexes, ataxia, seizures, intellectual disability, pale skin and often curled hair. We used X-ray diffraction and tensile testing to determine potential changes to the structure of keratin intermediate filaments (IFs) in the hair of patients with GAN. A statistically significant decrease in the 47 and the 27 Å diffraction signals were observed. Tensile tests determined that the hair was slightly stiffer, stronger and more extensible in GAN patients. These results suggest that the structure of keratin IFs in hair is altered in GAN, and the findings are compatible with an increased positional disorder of the keratin tetramers within the hair fibres. PMID:28424304

Giant axonal neuropathy alters the structure of keratin intermediate filaments in human hair.

PubMed

Soomro, Asfia; Alsop, Richard J; Negishi, Atsuko; Kreplak, Laurent; Fudge, Douglas; Kuczmarski, Edward R; Goldman, Robert D; Rheinstädter, Maikel C

2017-04-01

Giant axonal neuropathy (GAN) follows an autosomal recessive genetic inheritance and impedes the peripheral and central nervous system due to axonal swellings that are packed with neurofilaments. The patients display a number of phenotypes, including hypotonia, muscle weakness, decreased reflexes, ataxia, seizures, intellectual disability, pale skin and often curled hair. We used X-ray diffraction and tensile testing to determine potential changes to the structure of keratin intermediate filaments (IFs) in the hair of patients with GAN. A statistically significant decrease in the 47 and the 27 Å diffraction signals were observed. Tensile tests determined that the hair was slightly stiffer, stronger and more extensible in GAN patients. These results suggest that the structure of keratin IFs in hair is altered in GAN, and the findings are compatible with an increased positional disorder of the keratin tetramers within the hair fibres. © 2017 The Author(s).

Regenerated keratin membrane to match the in vitro drug diffusion through human epidermis

PubMed Central

Selmin, Francesca; Cilurzo, Francesco; Aluigi, Annalisa; Franzè, Silvia; Minghetti, Paola

2012-01-01

This work aimed to develop membranes made of regenerated keratin and ceramides (CERs) to match the barrier property of the human stratum corneum in in vitro percutaneous absorption studies. The membrane composition was optimized on the basis of the in vitro drug diffusion profiles of ibuprofen, propranolol and testosterone chosen as model drugs on the basis of their different diffusion and solubility properties. The data were compared to those obtained using human epidermis. The ATR-FTIR and SEM analyses revealed that CERs were suspended into the regenerated keratin matrix, even if a partial solubilization occurred. It resulted in the membranes being physically stable after exposure to aqueous buffer and/or mineral oil and the fluxes of ibuprofen and propranolol from these vehicles through membranes and human skin were of the same order of magnitude. The best relationship with human epidermis data was obtained with 180 μm-thick membrane containing 1% ceramide III and 1% ceramide VI. The data on the testosterone diffusion were affected by the exposure of the membrane to a water/ethanol solution over a prolonged period of time, indicating that such an organic solvent was able to modify the supermolecular organization of keratin and CERs. The keratin/CER membranes can represent a simplified model to assay the in vitro skin permeability study of small molecules. PMID:25755997

In situ identification of keratin-hydrolyzing organisms in swine manure inoculated anaerobic digesters.

PubMed

Xia, Yun; Massé, Daniel I; McAllister, Tim A; Beaulieu, Carole; Talbot, Guylaine; Kong, Yunhong; Seviour, Robert

2011-12-01

Feathers, a poultry byproduct, are composed of > 90% keratin which is resistant to degradation during anaerobic digestion. In this study, four 42-L anaerobic digesters inoculated with adapted swine manure were used to investigate feather digestion. Ground feathers were added into two anaerobic digesters for biogas production, whereas another two without feathers were used as negative control. Feather degradation and enhanced methane production were recorded. Keratin-hydrolyzing organisms (KHOs) were visualized in the feather bag fluids after boron-dipyrromethene (BODIPY) fluorescence casein staining. Their abundances correlated (R(2)  = 0.96) to feather digestion rates. A 16S rRNA clone library was constructed for the bacterial populations attached to the feather particles. Ninety-three clones (> 1300 bp) were retrieved and 57 (61%) belonged to class Clostridia in the phylum Firmicutes, while 34 (37%) belonged to class Bacteroidia in the phylum Bacteroidetes. Four oligonucleotide FISH probes were designed for the major Clostridia clusters and used with other FISH probes to identify the KHOs. Probe FIMs1029 hybridized with most (> 80%) of the KHOs. Its targeted sequence perfectly matches that possessed by 10 Clostridia 16S rRNA gene clones belonging to a previously uncharacterized new genus closely related to Alkaliphilus in the subfamily Clostridiaceae 2 of family Clostridiaceae. © 2011 Her Majesty the Queen in Right of Canada, as represented by the Minister of Agriculture and Agri-Food Canada. Published by Blackwell Publishing Ltd.

Evidence of accelerated beak growth associated with avian keratin disorder in Black-capped Chickadees (Poecile atricapillus)

USGS Publications Warehouse

Van Hemert, Caroline; Handel, Colleen M.; O'Hara, Todd M.

2012-01-01

We recently documented an epizootic of beak deformities in more than 2,000 Blackcapped Chickadees (Poecile atricapillus) and other wild bird species in North America. This emerging avian disease, which has been termed avian keratin disorder, results in gross overgrowth of the rhamphotheca, the outer, keratinized layer of the beak. To test the hypothesis that the beak deformities characteristic of this disorder are associated with accelerated keratin production, we measured rates of beak growth and wear in affected Black-capped Chickadees (n=16) and a control sample of unaffected chickadees (n=14) collected from south-central (61°09′−61°38′N, 149°11′ −149°48′W) and interior Alaska (64°51′ −64°53′N, 147°49′ −147°59′W). Rates of absolute growth were 50–100% higher in affected birds than they were in control birds and exceeded records from other passerine species. These results suggest that abnormally rapid epidermal growth is the primary physical mechanism by which beak deformities develop and are maintained in affected chickadees. Although beak overgrowth typically worsened over time, differential patterns of wear influenced the severity and morphology of deformities. In some cases, the effects of accelerated keratin growth were partially mitigated by frequent breakage of rhamphothecal tips. However, mortalities occurred in 9 of 16 birds (56%) with beak deformities during the study, suggesting that avian keratin disorder results in severe health consequences for affected birds. Additional study of factors that control beak keratin production is needed to understand the pathogenesis of this debilitating disease in wild birds.

Evidence of accelerated beak growth associated with avian keratin disorder in black-capped chickadees (Poecile atricapillus)

USGS Publications Warehouse

Van Hemert, Caroline R.; Handel, Colleen M.; O'Hara, Todd M.

2012-01-01

We recently documented an epizootic of beak deformities in more than 2,000 Black-capped Chickadees (Poecile atricapillus) and other wild bird species in North America. This emerging avian disease, which has been termed avian keratin disorder, results in gross overgrowth of the rhamphotheca, the outer, keratinized layer of the beak. To test the hypothesis that the beak deformities characteristic of this disorder are associated with accelerated keratin production, we measured rates of beak growth and wear in affected Black-capped Chickadees (n=16) and a control sample of unaffected chickadees (n=14) collected from south-central (61°09′–61°38′N, 149°11′–149°48′W) and interior Alaska (64°51′–64°53′N, 147°49′–147°59′W). Rates of absolute growth were 50–100% higher in affected birds than they were in control birds and exceeded records from other passerine species. These results suggest that abnormally rapid epidermal growth is the primary physical mechanism by which beak deformities develop and are maintained in affected chickadees. Although beak overgrowth typically worsened over time, differential patterns of wear influenced the severity and morphology of deformities. In some cases, the effects of accelerated keratin growth were partially mitigated by frequent breakage of rhamphothecal tips. However, mortalities occurred in 9 of 16 birds (56%) with beak deformities during the study, suggesting that avian keratin disorder results in severe health consequences for affected birds. Additional study of factors that control beak keratin production is needed to understand the pathogenesis of this debilitating disease in wild birds.

Clinical evaluation of a new collagen matrix (Mucograft prototype) to enhance the width of keratinized tissue in patients with fixed prosthetic restorations: a randomized prospective clinical trial.

PubMed

Sanz, Mariano; Lorenzo, Ramón; Aranda, Juan J; Martin, Conchita; Orsini, Marco

2009-10-01

The aim of this study was to test a new collagen matrix (CM) aimed to increase keratinized gingiva/mucosa when compared with the free connective tissue graft (CTG). This randomized longitudinal parallel controlled clinical trial studied 20 patients with at least one location with minimal keratinized tissue (keratinized tissue. As secondary outcomes, the aesthetic outlook, the maintenance of periodontal health and the patient morbidity were assessed pre-operatively at 1, 3 and 6 months. At 6 months, the CTG attained a mean width of keratinized tissue of 2.6 (0.9) mm, while the CM was 2.5 (0.9) mm, these differences being insignificant. In both groups, there was a marked contraction (60% and 67%, respectively) although the periodontal parameters were not affected. The CM group had a significantly lower patient morbidity (pain and medication intake) as well as reduced surgery time. These results prove that this new CM was as effective and predictable as the CTG for attaining a band of keratinized tissue, but its use was associated with a significantly lower patient morbidity.

Characterization of an AtCCX5 gene from Arabidopsis thaliana that involves in high-affinity K{sup +} uptake and Na{sup +} transport in yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Xinxin; Zhang, Min; Takano, Tetsuo

Highlights: {yields} The AtCCX5 protein coding a putative cation calcium exchanger was characterized. {yields} AtCCX5 expressed in yeast was localized in the plasma membrane and nuclear periphery. {yields} AtCCX5 protein did not show the same transport properties as the CAXs. {yields} AtCCX5 protein involves in mediating high-affinity K{sup +} uptake in yeast. {yields} AtCCX5 protein also involves in Na{sup +} transport in yeast. -- Abstract: The gene for a putative cation calcium exchanger (CCX) from Arabidopsis thaliana, AtCCX5, was cloned and its function was analyzed in yeast. Green fluorescent protein-tagged AtCCX5 expressed in yeast was localized in the plasma membranemore » and nuclear periphery. The yeast transformants expressing AtCCX5 were created and their growth in the presence of various cations (K{sup +}, Na{sup +}, Ca{sup 2+}, Mg{sup 2+}, Fe{sup 2+}, Cu{sup 2+}, Co{sup 2+}, Cd{sup 2+}, Mn{sup 2+}, Ba{sup 2+}, Ni{sup 2+}, Zn{sup 2+}, and Li{sup +}) were analyzed. AtCCX5 expression was found to affect the response to K{sup +} and Na{sup +} in yeast. The AtCCX5 transformant also showed a little better growth to Zn{sup 2+}. The yeast mutant 9.3 expressing AtCCX5 restored growth of the mutant on medium with low K{sup +} (0.5 mM), and also suppressed its Na{sup +} sensitivity. Ion uptake experiments showed that AtCCX5 mediated relatively high-affinity K{sup +} uptake and was also involved in Na{sup +} transport in yeast. Taken together, these findings suggest that the AtCCX5 is a novel transport protein involves in mediating high-affinity K{sup +} uptake and Na{sup +} transport in yeast.« less

Effects of Nickel Treatment on H3K4 Trimethylation and Gene Expression

PubMed Central

Tchou-Wong, Kam-Meng; Kluz, Thomas; Arita, Adriana; Smith, Phillip R.; Brown, Stuart; Costa, Max

2011-01-01

Occupational exposure to nickel compounds has been associated with lung and nasal cancers. We have previously shown that exposure of the human lung adenocarcinoma A549 cells to NiCl2 for 24 hr significantly increased global levels of trimethylated H3K4 (H3K4me3), a transcriptional activating mark that maps to the promoters of transcribed genes. To further understand the potential epigenetic mechanism(s) underlying nickel carcinogenesis, we performed genome-wide mapping of H3K4me3 by chromatin immunoprecipitation and direct genome sequencing (ChIP-seq) and correlated with transcriptome genome-wide mapping of RNA transcripts by massive parallel sequencing of cDNA (RNA-seq). The effect of NiCl2 treatment on H3K4me3 peaks within 5,000 bp of transcription start sites (TSSs) on a set of genes highly induced by nickel in both A549 cells and human peripheral blood mononuclear cells were analyzed. Nickel exposure increased the level of H3K4 trimethylation in both the promoters and coding regions of several genes including CA9 and NDRG1 that were increased in expression in A549 cells. We have also compared the extent of the H3K4 trimethylation in the absence and presence of formaldehyde crosslinking and observed that crosslinking of chromatin was required to observe H3K4 trimethylation in the coding regions immediately downstream of TSSs of some nickel-induced genes including ADM and IGFBP3. This is the first genome-wide mapping of trimethylated H3K4 in the promoter and coding regions of genes induced after exposure to NiCl2. This study may provide insights into the epigenetic mechanism(s) underlying the carcinogenicity of nickel compounds. PMID:21455298

The small heat shock protein Hsp27 affects assembly dynamics and structure of keratin intermediate filament networks.

PubMed

Kayser, Jona; Haslbeck, Martin; Dempfle, Lisa; Krause, Maike; Grashoff, Carsten; Buchner, Johannes; Herrmann, Harald; Bausch, Andreas R

2013-10-15

The mechanical properties of living cells are essential for many processes. They are defined by the cytoskeleton, a composite network of protein fibers. Thus, the precise control of its architecture is of paramount importance. Our knowledge about the molecular and physical mechanisms defining the network structure remains scarce, especially for the intermediate filament cytoskeleton. Here, we investigate the effect of small heat shock proteins on the keratin 8/18 intermediate filament cytoskeleton using a well-controlled model system of reconstituted keratin networks. We demonstrate that Hsp27 severely alters the structure of such networks by changing their assembly dynamics. Furthermore, the C-terminal tail domain of keratin 8 is shown to be essential for this effect. Combining results from fluorescence and electron microscopy with data from analytical ultracentrifugation reveals the crucial role of kinetic trapping in keratin network formation. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Keratinizing odontogenic cysts with a spectrum of verrucoid morphology: investigation of a potential role of human papillomavirus.

PubMed

Lalla, Kalpesh; Mahomed, Farzana; Meer, Shabnum

2016-11-01

The role of human papillomavirus (HPV) in keratinizing odontogenic cysts (OC) has only rarely been studied. We describe the clinicopathologic findings in a series of OCs that had unusual keratinization patterns and were investigated for a possible HPV etiology. Tissue samples from 29 patients with keratinizing OCs were studied for light microscopic features suggestive of HPV infection and by an HPV DNA polymerase chain reaction assay. The mean age at presentation was 31.1 years; 79.3% of the OCs occurred in the mandible and 46.4% were associated with an impacted tooth. The phenotypic characteristics koilocytes, hypergranulosis, and a verrucous pattern of the cyst-lining epithelium were observed in 69%, 62.1%, and 17.2% of cases, respectively. These histomorphologic features did not, however, correlate with HPV infection. HPV does not appear to play a role in keratinizing OCs and is not responsible for the wart-like histomorphologic features that may be seen in these lesions. Copyright © 2016 Elsevier Inc. All rights reserved.

The use of mucograft collagen matrix to augment the zone of keratinized tissue around teeth: a pilot study.

PubMed

Nevins, Myron; Nevins, Marc L; Kim, Soo-Woo; Schupbach, Peter; Kim, David M

2011-01-01

This prospective split-mouth pilot case series compared the use of a bilayer collagen matrix (CM) to an autogenous gingival graft (AGG) in the ability to increase the zone of keratinized attached gingiva. Five patients with inadequate amounts of keratinized attached gingiva bilaterally in the posterior mandible were enrolled using a split-mouth design. There were statistically significant increases in attached gingiva at all test (CM) and control (AGG) sites. The CM sites at 12 months blended well with surrounding tissues, while the AGG sites were morphologically dissimilar to the adjacent areas. Biopsy results showed intrapatient histologic similarity between CM and AGG treatments, with all sites exhibiting mature connective tissue covered by keratinized epithelium. Thus, the obtained data support further investigations in evaluating the role of CM as a viable alternative to AGG in augmenting areas deficient in keratinized gingiva.

Pharmacological and gene regulation properties point to the SlHAK5 K+ transporter as a system for high-affinity Cs+ uptake in tomato plants.

PubMed

Ródenas, Reyes; Nieves-Cordones, Manuel; Rivero, Rosa M; Martinez, Vicente; Rubio, Francisco

2018-04-01

Potassium (K + ) and cesium (Cs + ) are chemically similar but while K + is an essential nutrient, Cs + can be toxic for living organisms, plants included. Two different situations could lead to problems derived from the presence of Cs + in agricultural systems: (1) presence of Cs + at high concentrations that could produce toxic effects on plants, (2) presence of micromolar concentrations of radiocesium, which can be accumulated in the plant and affect animal and human health through the food chain. While K + uptake has been well described in tomato plants, information on molecular mechanisms involved in Cs + accumulation in this species is absent. Here, we show that in tomato plants, high concentrations of Cs + produce deficiency of K + but do not induce high-affinity K + uptake or the gene encoding the high-affinity K + transporter SlHAK5. At these concentrations, Cs + uptake takes place through a Ca 2+ -sensitive pathway, probably a non-selective cation channel. At micromolar concentrations, Cs + is accumulated by a high-affinity uptake system upregulated in K + -starved plants. This high-affinity Cs + uptake shares features with high-affinity K + uptake. It is sensitive to NH 4 + and insensitive to Ba 2+ and Ca 2+ and its presence parallels the pattern of SlHAK5 expression. Moreover, blockers of reactive oxygen species and ethylene action repress SlHAK5 and negatively regulate both high-affinity K + and Cs + uptake. Thus, we propose that SlHAK5 contributes to Cs + uptake from micromolar concentrations in tomato plants and can constitute a pathway for radiocesium transfer from contaminated areas to the food chain. © 2017 Scandinavian Plant Physiology Society.

Comparison of acellular dermal graft and palatal autograft in the reconstruction of keratinized gingiva around dental implants: a case report.

PubMed

Yan, Ji-Jong; Tsai, Alex Yi-Min; Wong, Man-Ying; Hou, Lein-Tuan

2006-06-01

The use of autogenous gingival grafts has proved to be an effective and predictable way to increase the amount of keratinized gingiva. However, discomfort and pain at the donor site are unavoidable. Acellular dermal matrix (ADM) allograft can be used as a donor tissue to eliminate the need for another surgical site and alleviate pain and trauma. The purpose of this study was to evaluate the effectiveness of ADM allograft in increasing the width of keratinized gingiva around dental implants. A patient with inadequate keratinized gingiva around dental implants in maxillary and mandibular anterior regions received either an ADM graft or palatal autograft by random allocation. The width of keratinized gingiva and other clinical periodontal parameters were recorded initially and at 3 and 6 months after surgery. Both grafts provided satisfactory results. The width of keratinized tissues was increased by using the ADM allograft, but by a lesser amount than seen with the autogenous gingival graft.

Intermediate filament structure in fully differentiated (oxidised) trichocyte keratin.

PubMed

Fraser, R D Bruce; Parry, David A D

2017-10-01

For the past 50years there has been considerable debate over the sub-structure of the fully differentiated (oxidised) trichocyte keratin intermediate filament. Depending on the staining and preparative procedures employed, IF observed in transverse section in the transmission electron microscope have varied in appearance between that of a "ring" and a "ring-core" structure, corresponding to the so-called (8+0) and (7+1) protofilament arrangements. In a new analysis of the fine structure of the 1nm equatorial region of the X-ray diffraction pattern of quill we show that the observed pattern is consistent with the (8+0) model and we are also able to assign values to the various parameters. In contrast, we show that the observed X-ray pattern is inconsistent with a (7+1) arrangement. Furthermore, in the (7+1) model steric hindrance would be encountered between the core protofilament and those constituting the ring. The appearance of a central "core" in transverse TEM sections, previously attributed to a central protofilament, is explained in terms of portions of the apolar, disulfide-bonded head and/or tail domains of the trichocyte keratin IF molecules, including the conserved H subdomains, lying along the axis of the IF, thereby decreasing the efficacy of the reducing agents used prior to staining. The H1 subdomain, previously shown to be important in the assembly of epidermal IF molecules at the two- to four-molecule level, is likely to have a similar role for the trichocyte keratins and may form part of a central scaffold on which the molecules assemble into fully functional IF. Copyright © 2017 Elsevier Inc. All rights reserved.

Mutations of the EGFR, K-ras, EML4-ALK, and BRAF genes in resected pathological stage I lung adenocarcinoma.

PubMed

Ohba, Taro; Toyokawa, Gouji; Osoegawa, Atsushi; Hirai, Fumihiko; Yamaguchi, Masafumi; Taguchi, Ken-Ichi; Seto, Takashi; Takenoyama, Mitsuhiro; Ichinose, Yukito; Sugio, Kenji

2016-09-01

The EGFR, K-ras, EML4-ALK, and BRAF genes are oncogenic drivers of lung adenocarcinoma. We conducted this study to analyze the mutations of these genes in stage I adenocarcinoma. The subjects of this retrospective study were 256 patients with resected stage I lung adenocarcinoma. We analyzed mutations of the EGFR, K-ras, and BRAF genes, and the EML4-ALK fusion gene. We also assessed disease-free survival (DFS) to evaluate the prognostic value and overall survival (OS) to evaluate the predictive value of treatment after recurrence. Mutations of the EGFR, K-ras, EML4-ALK, and BRAF genes were detected in 120 (46.8 %), 14 (5.5 %), 6 (2.3 %), and 2 (0.8 %) of the 256 tumors. Two tumors had double mutations (0.8 %). The incidence of EGFR mutations was significantly higher in women than in men. The EML4-ALK fusion gene was detected only in younger patients. The DFS and OS of the K-ras mutant group were significantly worse than those of the EGFR mutant group, the EML4-ALK fusion gene group, and the wild-type group. Six of the seven patients with the EML4-ALK fusion gene are still alive without recurrent disease. In patients with stage I adenocarcinoma, mutation of the K-ras gene was a poor prognostic factor for recurrence. The presence of a mutation of the EGFR or EML4-ALK gene was not a prognostic factor.

Characterization of the Lymantria dispar nucleopolyhedrovirus 25K FP gene

Treesearch

David S. Bischoff; James M. Slavicek

1996-01-01

The Lymantria dispar nucleopolyhedrovirus (LdMNPV) gene encoding the 25K FP protein has been cloned and sequenced. The 25KFP gene codes for a 217 amino acid protein with a predicted molecular mass of 24870 Da. Expression of the 25K FP protein in a rabbit reticulocyte system generated a 27 kDa protein, in close agreement with the...

Tissue-specific expression of the gene coding for human Clara cell 10-kD protein, a phospholipase A2-inhibitory protein.

PubMed Central

Peri, A; Cordella-Miele, E; Miele, L; Mukherjee, A B

1993-01-01

Clara cell 10-kD protein (cc10kD), a secretory phospholipase A2 inhibitor, is suggested to be the human counterpart of rabbit uteroglobin (UG). Because cc10kD is expressed constitutively at a very high level in the human respiratory epithelium, the 5' region of its gene may be useful in achieving organ-specific expression of recombinant DNA in gene therapy of diseases such as cystic fibrosis. However, it is important to establish the tissue-specific expression of this gene before designing gene transfer experiments. Since the UG gene in the rabbit is expressed in many other organs besides the lung and the endometrium, we investigated the organ and tissue specificity of human cc10kD gene expression using polymerase chain reaction, nucleotide sequence analysis, immunofluorescence, and Northern blotting. Our results indicate that, in addition to the lung, cc10kD is expressed in several nonrespiratory organs, with a distribution pattern very similar, if not identical, to that of UG in the rabbit. These results underscore the necessity for more detailed analyses of the 5' region of the human cc10kD gene before its usefulness in gene therapy could be fully assessed. These data also suggest that cc10kD and UG may have similar physiological function(s). Images PMID:8227325

Amplification of Chromosome 1q Genes Encoding the Phosphoinositide Signalling Enzymes PI4KB, AKT3, PIP5K1A and PI3KC2B in Breast Cancer

PubMed Central

Waugh, Mark G.

2014-01-01

Little is known about the possible oncogenic roles of genes encoding for the phosphatidylinositol 4-kinases, a family of enzymes that regulate an early step in phosphoinositide signalling. To address this issue, the mutational status of all four human phosphatidylinositol 4-kinases genes was analyzed across 852 breast cancer samples using the COSMIC data resource. Point mutations in the phosphatidylinositol 4-kinase genes were uncommon and appeared in less than 1% of the patient samples however, 62% of the tumours had increases in gene copy number for PI4KB which encodes the phosphatidylinositol 4-kinase IIIbeta isozyme. Extending this analysis to subsequent enzymes in the phosphoinositide signalling cascades revealed that the only PIP5K1A, PI3KC2B and AKT3 genes exhibited similar patterns of gene copy number variation. By comparison, gene copy number increases for established oncogenes such as EGFR and HER2/Neu were only evident in 20% of the samples. The PI4KB, PIP5K1A, PI3KC2B and AKT3 genes are related in that they all localize to chromosome 1q which is often structurally and numerically abnormal in breast cancer. These results demonstrate that a gene quartet encoding a potential phosphoinositide signalling pathway is amplified in a subset of breast cancers. PMID:25368680

Keratin hydrogel carrier system for simultaneous delivery of exogenous growth factors and muscle progenitor cells1,2,3

PubMed Central

Tomblyn, Seth; Kneller, Elizabeth Pettit; Walker, Stephen J.; Ellenburg, Mary D.; Kowalczewski, Christine J.; Van Dyke, Mark; Burnett, Luke; Saul, Justin M.

2017-01-01

Ideal material characteristics for tissue engineering or regenerative medicine approaches to volumetric muscle loss (VML) include the ability to deliver cells, growth factors and molecules that support tissue formation from a system with a tunable degradation profile. Two different types of human hair-derived keratins were tested as options to fulfill these VML design requirements: (1) oxidatively extracted keratin (keratose) characterized by a lack of covalent crosslinking between cysteine residues, and (2) reductively extracted keratin (kerateine) characterized by disulfide crosslinks. Human skeletal muscle myoblasts cultured on coatings of both types of keratin had increased numbers of multinucleated cells compared to collagen or Matrigel™ and adhesion levels greater than collagen. Rheology showed elastic moduli from 102 – 105 Pa and viscous moduli from 101 – 104 Pa depending on gel concentration and keratin type. Kerateine and keratose showed differing rates of degradation due to the presence or absence of disulfide crosslinks, which likely contributed to observed differences in release profiles of several growth factors. In vivo testing in a subcutaneous mouse model showed that keratose hydrogels can be used to deliver mouse muscle progenitor cells and growth factors. Histological assessment showed minimal inflammatory responses and an increase in markers of muscle formation. PMID:25953729

Immunohistochemical localisation of keratin and luminal epithelial antigen in myoepithelial and luminal epithelial cells of human mammary and salivary gland tumours.

PubMed

Nathrath, W B; Wilson, P D; Trejdosiewicz, L K

1982-01-01

Rabbit antisera to human 40-63 000 MW epidermal keratin, one batch with restricted distribution of reactivity from an initial (aK1) and one with "broad spectrum" distribution of reactivity from a late bleeding (aK), and to "luminal epithelial antigen" (aLEA) were applied to formalin fixed paraffin embedded sections of human normal and neoplastic mammary and salivary glands using an indirect immunoperoxidase method. aK1 reacted with myoepithelial cells, aLEA with luminal epithelial cells and aK with both cell types in normal mammary and salivary gland. In breast carcinomas the majority of intraluminal and infiltrating carcinoma cells reacted with aLEA but not with aK1 which reacted only with surrounding myoepithelial cells. aK reacted with both myoepithelial cells and with intraluminal and infiltrating tumour cells. In the salivary gland adenomas the majority of cells reacted with aK, and those cells arranged in a tubular fashion reacted with aLEA.

Studies on the expression of an H-2K/human growth hormone fusion gene in giant transgenic mice.

PubMed Central

Morello, D; Moore, G; Salmon, A M; Yaniv, M; Babinet, C

1986-01-01

Transgenic mice carrying the H-2K/human growth hormone (hGH) fusion gene were produced by microinjecting into the pronucleus of fertilized eggs DNA molecules containing 2 kb of the 5' flanking sequences (including promoter) of the class I H-2Kb gene joined to the coding sequences of the hGH gene. Thirteen transgenic mice were obtained which all contained detectable levels of hGH hormone in their blood. Nine grew larger than their control litter-mates. Endogenous H-2Kb and exogenous hGH mRNA levels were analysed by S1 nuclease digestion experiments. hGH transcripts were found in all the tissues examined and the pattern of expression paralleled that of endogenous H-2K gene expression, being high in liver and lymphoid organs and low in muscle and brain. Thus 2 kb of the 5' promoter/regulatory region of the H-2K gene are sufficient to ensure regulated expression of hGH in transgenic mice. This promoter may therefore be of use to target the expression of different exogenous genes in most tissues of transgenic mice and to study the biological role of the corresponding proteins in different cellular environments. Images Fig. 2. Fig. 3. Fig. 4. Fig. 5. PMID:3019667

Comparative Study of Ultrasonication-Induced and Naturally Self-Assembled Silk Fibroin-Wool Keratin Hydrogel Biomaterials

PubMed Central

Vu, Trang; Xue, Ye; Vuong, Trinh; Erbe, Matthew; Bennet, Christopher; Palazzo, Ben; Popielski, Lucas; Rodriguez, Nelson; Hu, Xiao

2016-01-01

This study reports the formation of biocompatible hydrogels using protein polymers from natural silk cocoon fibroins and sheep wool keratins. Silk fibroin protein contains β-sheet secondary structures, allowing for the formation of physical cross-linkers in the hydrogels. Comparative studies were performed on two groups of samples. In the first group, ultrasonication was used to induce a quick gelation of a protein aqueous solution, enhancing the ability of Bombyx mori silk fibroin chains to quickly entrap the wool keratin protein molecules homogenously. In the second group, silk/keratin mixtures were left at room temperature for days, resulting in naturally-assembled gelled solutions. It was found that silk/wool blended solutions can form hydrogels at different mixing ratios, with perfectly interconnected gel structure when the wool content was less than 30 weight percent (wt %) for the first group (ultrasonication), and 10 wt % for the second group (natural gel). Differential scanning calorimetry (DSC) and temperature modulated DSC (TMDSC) were used to confirm that the fibroin/keratin hydrogel system was well-blended without phase separation. Fourier transform infrared spectroscopy (FTIR) was used to investigate the secondary structures of blended protein gels. It was found that intermolecular β-sheet contents significantly increase as the system contains more silk for both groups of samples, resulting in stable crystalline cross-linkers in the blended hydrogel structures. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to analyze the samples’ characteristic morphology on both micro- and nanoscales, which showed that ultrasonic waves can significantly enhance the cross-linker formation and avoid phase separation between silk and keratin molecules in the blended systems. With the ability to form cross-linkages non-chemically, these silk/wool hydrogels may be economically useful for various biomedical applications, thanks to the

Multicenter Clinical Trial of Keratin Biomaterial for Peripheral Nerve Regeneration

DTIC Science & Technology

2012-10-01

Drug Evaluation Research (CDER), the Center for Biologics E valuation Research (CBER), and the Center for Devices and Radiological Health ( CDRH ) on May...Research (CBER) , and the Center for Devices and Radiological Health ( CDRH ) to clar ify the designation of the keratin hydrogel. During this m eeting

Keratin-based products for effective wound care management in superficial and partial thickness burns injuries.

PubMed

Loan, Fiona; Cassidy, Sharon; Marsh, Clive; Simcock, Jeremy

2016-05-01

This n=40 cohort study on superficial and partial thickness burns compares novel keratin-based products with the standard products used at our facility. The keratin products are found to facilitate healing with minimal scarring, be well tolerated with minimal pain and itch, be easy to use for the health professional and be cost effective for the health care provider. For these reasons they are being adopted into use at our facility. Copyright © 2015 Elsevier Ltd and ISBI. All rights reserved.

Differential expression of cyclin D1 in keratin-producing odontogenic cysts.

PubMed

Vera-Sirera, Beatriz; Forner-Navarro, Leopoldo; Vera-Sempere, Francisco

2015-01-01

The aim of the present study was to analyze the expression levels of Cyclin D1 (CCD1), a nuclear protein that plays a crucial role in cell cycle progression, in a series of keratin-producing odontogenic cysts. A total of 58 keratin-producing odontogenic cysts, diagnosed over ten years and classified according to the WHO 2005 criteria, were immunohistochemically analyzed in terms of CCD1 expression, which was quantified in the basal, suprabasal and intermediate/superficial epithelial compartments. The extent of immunostaining was measured as a proportion of total epithelial thickness. Quantified immunohistochemical data were correlated with clinicopathological features and clinical recurrence. Keratin-producing odontogenic cysts were classified as 6 syndromic keratocystic odontogenic tumors (S-KCOT), 40 sporadic or non-syndromic KCOT (NS-KCOT) and 12 orthokeratinized odontogenic cysts (OOC). Immunohistochemically, CCD1 staining was evident predominantly in the parabasal region of all cystic lesions, but among-lesion differences were apparent, showing a clear expansion of parabasal compartment especially in the S-KCOT, followed to a lesser extent in the NS-KCOT, and being much more reduced in the OOC, which had the greatest average epithelial thickness. The differential expression of CCD1 noted in the present study suggests that dysregulation of cell cycle progression from G1 to the S phase contributes to the different aggressiveness of these lesions. However, CCD1 expression levels did not predict NS-KCOT recurrence, which is likely influenced by factors unrelated to lesion biology.

Oncosuppressive suicide gene virotherapy "PVH1-yCD/5-FC" for pancreatic peritoneal carcinomatosis treatment: NFκB and Akt/PI3K involvement.

PubMed

Réjiba, Soukaina; Bigand, Christelle; Parmentier, Celine; Masmoudi, Ahmed; Hajri, Amor

2013-01-01

Peritoneal carcinomatosis is common in advanced pancreatic cancer. Despite current standard treatment, patients with this disease until recently were considered incurable. Cancer gene therapy using oncolytic viruses have generated much interest over the past few years. Here, we investigated a new gene directed enzyme prodrug therapy (GDEPT) approach for an oncosuppressive virotherapy strategy using parvovirus H1 (PV-H1) which preferentially replicates and kills malignant cells. Although, PV-H1 is not potent enough to destroy tumors, it represents an attractive vector for cancer gene therapy. We therefore sought to determine whether the suicide gene/prodrug system, yCD/5-FC could be rationally combined to PV-H1 augmenting its intrinsic oncolytic activity for pancreatic cancer prevention and treatment. We showed that the engineered recombinant parvovirus rPVH1-yCD with 5-FC treatment increased significantly the intrinsic cytotoxic effect and resulted in potent induction of apoptosis and tumor growth inhibition in chemosensitive and chemoresistant cells. Additionally, the suicide gene-expressing PV-H1 infection reduced significantly the constitutive activities of NFκB and Akt/PI3K. Combination of their pharmacological inhibitors (MG132 and LY294002) with rPVH1-yCD/5-FC resulted in substantial increase of antitumor activity. In vivo, high and sustained expression of NS1 and yCD was observed in the disseminated tumor nodules and absent in normal tissues. Treatment of mice bearing intraperitoneal pancreatic carcinomatosis with rPVH1-yCD/5-FC resulted in a drastic inhibition of tumor cell spreading and subsequent increase in long-term survival. Together, the presented data show the improved oncolytic activity of wPV-H1 by yCD/5-FC and thus provides valuable effective and promising virotherapy strategy for prevention of tumor recurrence and treatment. In the light of this study, the suicide gene parvovirotherapy approach represents a new weapon in the war against

Extensive keratinized tissue augmentation during implant rehabilitation after Le Fort I osteotomy: using a new porcine collagen membrane (Mucoderm).

PubMed

Nocini, Pier Francesco; Castellani, Roberto; Zanotti, Guglielmo; Gelpi, Federico; Covani, Ugo; Marconcini, Simone; de Santis, Daniele

2014-05-01

The aim of this study was to test a new collagen matrix (Mucoderm) positioned during oral implant abutment connection. A patient previously treated with Le Fort I for bone augmentation and 8 implants showing minimal amount of keratinized tissue was selected for an extensive keratinized tissue augmentation and deepening of the oral vestibule by apically positioning a split palatal flap and palatal grafting with Mucoderm. Clinical data at 9 and 14 days and 1 and 2 months showed resorption of the collagen graft, augmentation of the keratinized tissue around the implants, and deepening of the vestibule, with minimal morbidity and reduced surgical treatment time. However, some vestibular keratinized tissue contraction was evident. The new collagen matrix may be a promising material as a substitute for an autologous gingival/connective tissue graft. Despite the preliminary results of this innovative article, before drawing any general conclusion, the benefit of the procedure should be further evaluated by prospective clinical trials.

Immunoelectron microscopic localisation of keratin and luminal epithelial antigens in normal and neoplastic urothelium.

PubMed

Wilson, P D; Nathrath, W B; Trejdosiewicz, L K

1982-01-01

Immunoelectron microscope cytochemistry was carried out on 2% paraformaldehyde fixed, 50 mu sections of normal urothelium and bladder carcinoma cells in culture using antisera raised in rabbits to human 40-63 000 MW epidermal "broad spectrum" keratin and calf urothelial "luminal epithelial antigen" (aLEA) Both the unconjugated and indirect immunoperoxidase-DAB techniques were used before routine embedding. The localisation of both keratin and luminal epithelial antigen (LEA) was similar in normal and neoplastic cells and reaction product was associated not only with tonofilaments but also lining membrane vesicles and on fine filaments in the cytoplasmic ground substance.

Preservation of keratinized mucosa around implants using a prefabricated implant-retained stent: a case-control study

PubMed Central

2016-01-01

Purpose The aim of this study was to clinically assess the impact of a prefabricated implant-retained stent clipped over healing abutments on the preservation of keratinized mucosa around implants after implant surgery, and to compare it with horizontal external mattress sutures. Methods A total of 50 patients were enrolled in this study. In the test group, a prefabricated implant-retained stent was clipped on the healing abutment after implant surgery to replace the keratinized tissue bucco-apically. In the control group, horizontal external mattress sutures were applied instead of using a stent. After the surgical procedure, the width of the buccal keratinized mucosa was measured at the mesial, middle, and distal aspects of the healing abutment. The change in the width of the buccal keratinized mucosa was assessed at 1 and 3 months. Results Healing was uneventful in both groups. The difference of width between baseline and 1 month was −0.26±0.85 mm in the test group, without any statistical significance (P=0.137). Meanwhile, the corresponding difference in the control group was −0.74±0.73 mm and it showed statistical significance (P<0.001). The difference of width between baseline and 3 months was −0.57±0.97 mm in the test group and −0.86±0.71 mm in the control group. These reductions were statistically significant (P<0.05); however, there was no difference between the 2 groups. Conclusions Using a prefabricated implant-retained stent was shown to be effective in the preservation of the keratinized mucosa around implants and it was simple and straightforward in comparison to the horizontal external mattress suture technique. PMID:27800215

Mammalian keratin associated proteins (KRTAPs) subgenomes: disentangling hair diversity and adaptation to terrestrial and aquatic environments.

PubMed

Khan, Imran; Maldonado, Emanuel; Vasconcelos, Vítor; O'Brien, Stephen J; Johnson, Warren E; Antunes, Agostinho

2014-09-10

Adaptation of mammals to terrestrial life was facilitated by the unique vertebrate trait of body hair, which occurs in a range of morphological patterns. Keratin associated proteins (KRTAPs), the major structural hair shaft proteins, are largely responsible for hair variation. We exhaustively characterized the KRTAP gene family in 22 mammalian genomes, confirming the existence of 30 KRTAP subfamilies evolving at different rates with varying degrees of diversification and homogenization. Within the two major classes of KRTAPs, the high cysteine (HS) subfamily experienced strong concerted evolution, high rates of gene conversion/recombination and high GC content. In contrast, high glycine-tyrosine (HGT) KRTAPs showed evidence of positive selection and low rates of gene conversion/recombination. Species with more hair and of higher complexity tended to have more KRATP genes (gene expansion). The sloth, with long and coarse hair, had the most KRTAP genes (175 with 141 being intact). By contrast, the "hairless" dolphin had 35 KRTAPs and the highest pseudogenization rate (74% relative to the 19% mammalian average). Unique hair-related phenotypes, such as scales (armadillo) and spines (hedgehog), were correlated with changes in KRTAPs. Gene expression variation probably also influences hair diversification patterns, for example human have an identical KRTAP repertoire as apes, but much less hair. We hypothesize that differences in KRTAP gene repertoire and gene expression, together with distinct rates of gene conversion/recombination, pseudogenization and positive selection, are likely responsible for micro and macro-phenotypic hair diversification among mammals in response to adaptations to ecological pressures.

Fabrication and Characterization of Electrospun PCL-MgO-Keratin-Based Composite Nanofibers for Biomedical Applications

PubMed Central

Boakye, Maame A. D.; Rijal, Nava P.; Adhikari, Udhab; Bhattarai, Narayan

2015-01-01

Polymeric nanofibers are of great interest in biomedical applications, such as tissue engineering, drug delivery and wound healing, due to their ability to mimic and restore the function of natural extracellular matrix (ECM) found in tissues. Electrospinning has been heavily used to fabricate nanofibers because of its reliability and effectiveness. In our research, we fabricated poly(ε-caprolactone)-(PCL), magnesium oxide-(MgO) and keratin (K)-based composite nanofibers by electrospinning a blend solution of PCL, MgO and/or K. The electrospun nanofibers were analyzed by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), mechanical tensile testing and inductively-coupled plasma optical emission spectroscopy (ICP-OES). Nanofibers with diameters in the range of 0.2–2.2 µm were produced by using different ratios of PCL/MgO and PCL-K/MgO. These fibers showed a uniform morphology with suitable mechanical properties; ultimate tensile strength up to 3 MPa and Young’s modulus 10 MPa. The structural integrity of nanofiber mats was retained in aqueous and phosphate buffer saline (PBS) medium. This study provides a new composite material with structural and material properties suitable for potential application in tissue engineering. PMID:28793426

Effects of scalp dermatitis on chemical property of hair keratin

NASA Astrophysics Data System (ADS)

Kim, Kyung Sook; Shin, Min Kyung; Park, Hun-Kuk

2013-05-01

The effects of scalp dermatitis (seborrheic dermatitis (SD), psoriasis, and atopic dermatitis (AD)) on chemical properties of hair keratin were investigated by Fourier transform infrared (FT-IR) spectroscopy. Hairs were collected from lesional regions affected by SD, psoriasis, and AD and non-lesional regions separately. The hairs with SD were taken from patients with ages of 16-80 years. The ages of patients with psoriasis ranged from 8 to 67 years, and all patients exhibited moderate disease. Hairs with AD were taken from the patients with ages of 24-45 years and the average SCORing atopic dermatitis (SCORAD) was 48.75. Hairs from 20 normal adults were collected as a control. The FT-IR absorbance bands were analyzed by the Gaussian model to obtain the center frequency, half width, height, and area of each band. The height and area of all bands in the spectra were normalized to the amide I centered at 1652 cm-1 to quantitatively analyze the chemical composition of keratin. The spectra of hair with scalp dermatitis were different with that of control, the amide A components centered at 3278 cm-1 were smaller than those of the control. The psoriasis hair showed a large difference in the IR absorbance band between lesional and non-lesional hairs indicating good agreement with the morphological changes. The hairs with diseases did not show differences in the content of cystine, which was centered at 1054 cm-1, from the control. The chemical properties of keratin were not significantly different between the hairs affected by SD, psoriasis, and AD. However, the changes induced by scalp dermatitis were different with weathering. Therefore, FT-IR analysis could be used to screen differences between the physiological and pathological conditions of scalp hair.

RB inactivation in keratin 18 positive thymic epithelial cells promotes non-cell autonomous T cell hyperproliferation in genetically engineered mice.

PubMed

Song, Yurong; Sullivan, Teresa; Klarmann, Kimberly; Gilbert, Debra; O'Sullivan, T Norene; Lu, Lucy; Wang, Sophie; Haines, Diana C; Van Dyke, Terry; Keller, Jonathan R

2017-01-01

Thymic epithelial cells (TEC), as part of thymic stroma, provide essential growth factors/cytokines and self-antigens to support T cell development and selection. Deletion of Rb family proteins in adult thymic stroma leads to T cell hyperplasia in vivo. To determine whether deletion of Rb specifically in keratin (K) 18 positive TEC was sufficient for thymocyte hyperplasia, we conditionally inactivated Rb and its family members p107 and p130 in K18+ TEC in genetically engineered mice (TgK18GT121; K18 mice). We found that thymocyte hyperproliferation was induced in mice with Rb inactivation in K18+ TEC, while normal T cell development was maintained; suggesting that inactivation of Rb specifically in K18+ TEC was sufficient and responsible for the phenotype. Transplantation of wild type bone marrow cells into mice with Rb inactivation in K18+ TEC resulted in donor T lymphocyte hyperplasia confirming the non-cell autonomous requirement for Rb proteins in K18+ TEC in regulating T cell proliferation. Our data suggests that thymic epithelial cells play an important role in regulating lymphoid proliferation and thymus size.

Oncosuppressive Suicide Gene Virotherapy “PVH1-yCD/5-FC” for Pancreatic Peritoneal Carcinomatosis Treatment: NFκB and Akt/PI3K Involvement

PubMed Central

Réjiba, Soukaina; Bigand, Christelle; Parmentier, Celine; Masmoudi, Ahmed; Hajri, Amor

2013-01-01

Peritoneal carcinomatosis is common in advanced pancreatic cancer. Despite current standard treatment, patients with this disease until recently were considered incurable. Cancer gene therapy using oncolytic viruses have generated much interest over the past few years. Here, we investigated a new gene directed enzyme prodrug therapy (GDEPT) approach for an oncosuppressive virotherapy strategy using parvovirus H1 (PV-H1) which preferentially replicates and kills malignant cells. Although, PV-H1 is not potent enough to destroy tumors, it represents an attractive vector for cancer gene therapy. We therefore sought to determine whether the suicide gene/prodrug system, yCD/5-FC could be rationally combined to PV-H1 augmenting its intrinsic oncolytic activity for pancreatic cancer prevention and treatment. We showed that the engineered recombinant parvovirus rPVH1-yCD with 5-FC treatment increased significantly the intrinsic cytotoxic effect and resulted in potent induction of apoptosis and tumor growth inhibition in chemosensitive and chemoresistant cells. Additionally, the suicide gene-expressing PV-H1 infection reduced significantly the constitutive activities of NFκB and Akt/PI3K. Combination of their pharmacological inhibitors (MG132 and LY294002) with rPVH1-yCD/5-FC resulted in substantial increase of antitumor activity. In vivo, high and sustained expression of NS1 and yCD was observed in the disseminated tumor nodules and absent in normal tissues. Treatment of mice bearing intraperitoneal pancreatic carcinomatosis with rPVH1-yCD/5-FC resulted in a drastic inhibition of tumor cell spreading and subsequent increase in long-term survival. Together, the presented data show the improved oncolytic activity of wPV-H1 by yCD/5-FC and thus provides valuable effective and promising virotherapy strategy for prevention of tumor recurrence and treatment. In the light of this study, the suicide gene parvovirotherapy approach represents a new weapon in the war against

TGM5 mutations impact epidermal differentiation in acral peeling skin syndrome.

PubMed

Pigors, Manuela; Kiritsi, Dimitra; Cobzaru, Cristina; Schwieger-Briel, Agnes; Suárez, Jose; Faletra, Flavio; Aho, Heikki; Mäkelä, Leeni; Kern, Johannes S; Bruckner-Tuderman, Leena; Has, Cristina

2012-10-01

Acral peeling skin syndrome (APSS) is an autosomal recessive skin disorder characterized by acral blistering and peeling of the outermost layers of the epidermis. It is caused by mutations in the gene for transglutaminase 5, TGM5. Here, we report on clinical and molecular findings in 11 patients and extend the TGM5 mutation database by four, to our knowledge, previously unreported mutations: p.M1T, p.L41P, p.L214CfsX15, and p.S604IfsX9. The recurrent mutation p.G113C was found in 9 patients, but also in 3 of 100 control individuals in a heterozygous state, indicating that APSS might be more widespread than hitherto expected. Using quantitative real-time PCR, immunoblotting, and immunofluorescence analysis, we demonstrate that expression and distribution of several epidermal differentiation markers and corneodesmosin (CDSN) is altered in APSS keratinocytes and skin. Although the expression of transglutaminases 1 and 3 was not changed, we found an upregulation of keratin 1, keratin 10, involucrin, loricrin, and CDSN, probably as compensatory mechanisms for stabilization of the epidermal barrier. Our results give insights into the consequences of TGM5 mutations on terminal epidermal differentiation.

CTCF-KDM4A complex correlates with histone modifications that negatively regulate CHD5 gene expression in cancer cell lines

PubMed Central

Guerra-Calderas, Lissania; González-Barrios, Rodrigo; Patiño, Carlos César; Alcaraz, Nicolás; Salgado-Albarrán, Marisol; de León, David Cantú; Hernández, Clementina Castro; Sánchez-Pérez, Yesennia; Maldonado-Martínez, Héctor Aquiles; De la Rosa-Velazquez, Inti A.; Vargas-Romero, Fernanda; Herrera, Luis A.; García-Carrancá, Alejandro; Soto-Reyes, Ernesto

2018-01-01

Histone demethylase KDM4A is involved in H3K9me3 and H3K36me3 demethylation, which are epigenetic modifications associated with gene silencing and RNA Polymerase II elongation, respectively. KDM4A is abnormally expressed in cancer, affecting the expression of multiple targets, such as the CHD5 gene. This enzyme localizes at the first intron of CHD5, and the dissociation of KDM4A increases gene expression. In vitro assays showed that KDM4A-mediated demethylation is enhanced in the presence of CTCF, suggesting that CTCF could increase its enzymatic activity in vivo, however the specific mechanism by which CTCF and KDM4A might be involved in the CHD5 gene repression is poorly understood. Here, we show that CTCF and KDM4A form a protein complex, which is recruited into the first intron of CHD5. This is related to a decrease in H3K36me3/2 histone marks and is associated with its transcriptional downregulation. Depletion of CTCF or KDM4A by siRNA, triggered the reactivation of CHD5 expression, suggesting that both proteins are involved in the negative regulation of this gene. Furthermore, the knockout of KDM4A restored the CHD5 expression and H3K36me3 and H3K36me2 histone marks. Such mechanism acts independently of CHD5 promoter DNA methylation. Our findings support a novel mechanism of epigenetic repression at the gene body that does not involve promoter silencing. PMID:29682202

Expression of the Caulobacter heat shock gene dnaK is developmentally controlled during growth at normal temperatures.

PubMed Central

Gomes, S L; Gober, J W; Shapiro, L

1990-01-01

Caulobacter crescentus has a single dnaK gene that is highly homologous to the hsp70 family of heat shock genes. Analysis of the cloned and sequenced dnaK gene has shown that the deduced amino acid sequence could encode a protein of 67.6 kilodaltons that is 68% identical to the DnaK protein of Escherichia coli and 49% identical to the Drosophila and human hsp70 protein family. A partial open reading frame 165 base pairs 3' to the end of dnaK encodes a peptide of 190 amino acids that is 59% identical to DnaJ of E. coli. Northern blot analysis revealed a single 4.0-kilobase mRNA homologous to the cloned fragment. Since the dnaK coding region is 1.89 kilobases, dnaK and dnaJ may be transcribed as a polycistronic message. S1 mapping and primer extension experiments showed that transcription initiated at two sites 5' to the dnaK coding sequence. A single start site of transcription was identified during heat shock at 42 degrees C, and the predicted promoter sequence conformed to the consensus heat shock promoters of E. coli. At normal growth temperature (30 degrees C), a different start site was identified 3' to the heat shock start site that conformed to the E. coli sigma 70 promoter consensus sequence. S1 protection assays and analysis of expression of the dnaK gene fused to the lux transcription reporter gene showed that expression of dnaK is temporally controlled under normal physiological conditions and that transcription occurs just before the initiation of DNA replication. Thus, in both human cells (I. K. L. Milarski and R. I. Morimoto, Proc. Natl. Acad. Sci. USA 83:9517-9521, 1986) and in a simple bacterium, the transcription of a hsp70 gene is temporally controlled as a function of the cell cycle under normal growth conditions. Images PMID:2345134

The expression of keratins, vimentin, neurofilament proteins, smooth muscle actin, neuron-specific enolase, and synaptophysin in tumors of the specific glands in the canine anal region.

PubMed

Vos, J H; van den Ingh, T S; Ramaekers, F C; Molenbeek, R F; de Neijs, M; van Mil, F N; Ivanyi, D

1993-07-01

Eight canine tumors originating from specific glandular structures in the anal region, as well as metastatic tumor tissue of two of these cases (case Nos. 7, 8), were immunohistochemically analyzed using various monoclonal antibodies (MoAbs) directed against human keratin types, vimentin, neurofilament proteins, and alpha-smooth muscle actin. These tumors also were stained for the broad-spectrum neuroendocrine markers neuron-specific enolase (NSE) and synaptophysin. In histologically normal canine anal structures, alpha-smooth muscle actin and NSE antibodies stained basally localized (probably myoepithelial) cells in the anal glands and the anal sac glands. NSE staining also was present in a limited number of luminal cells in both anal glands and anal sac glands. Synaptophysin labeling was not observed in any of these glandular structures. Histologically, the tumors were differentiated into well- and moderately differentiated perianal gland tumors (n = 5) and carcinomas without perianal gland differentiation (n = 3), corresponding to the so-called apocrine carcinomas of the anal region. Immunohistochemically, the perianal gland tumors could be differentiated from the carcinomas by marked differences in staining pattern with the various keratin MoAbs, particularly MoAbs directed against human keratin types 7 and 18. The keratin-staining characteristics of the carcinomas suggest a glandular luminal cell origin. Metastases of the carcinomas showed loss of some keratin-staining characteristics as compared with the primary tumor. Staining for NSE was only observed in solitary cells and small cell clusters in the carcinomas and their metastases, whereas the alpha-smooth muscle actin antibody did not react with the carcinoma cells. None of the tumors stained for neurofilament proteins or synaptophysin. An unequivocal neuroendocrine nature of the carcinomas could not be substantiated by our immunohistochemical study, although the presence of a population of neuroendocrine

Two T7-like Bacteriophages, K5-2 and K5-4, Each Encodes Two Capsule Depolymerases: Isolation and Functional Characterization.

PubMed

Hsieh, Pei-Fang; Lin, Hsiao-Hsuan; Lin, Tzu-Lung; Chen, Yi-Yin; Wang, Jin-Town

2017-07-04

Two Klebsiella bacteriophages K5-2 and K5-4, which are able to infect and grow on either capsular types K30/K69 and K5 or K8 and K5 of Klebsiella strains, were isolated and characterized. Each phage contained two open reading frames (ORFs), which encoded two putative capsule depolymerases, respectively. The first ORF encoded tail fiber proteins, which have K30/K69 depolymerase and K8 depolymerase activities. The second ORF encoded hypothetical proteins, which are almost identical in amino acid sequences, and have K5 depolymerase activity. Alcian blue staining of enzyme-treated capsular polysaccharides (CPS) showed that purified depolymerases can cleave purified Klebsiella CPS in vitro and liberate monosaccharaides. Capsule K5 deletion mutants were not lysed by either phage, suggesting that the capsule was essential for phage infection. Bacterial killing was observed when incubated Klebsiella strains with phages but not with purified depolymerases. Treatment with the K5-4 phage significantly increased the survival of mice infected with a K. pneumoniae K5 strain. In conclusion, two dual host-specific Klebsiella phages and their tailspikes exhibit capsule depolymerase activity were characterized. Each phage and phage-encoded depolymerase has specificity for capsular type K30/K69, K8 or K5, and could be used for the typing and treatment of K. pneumoniae infection.

ATXR5 and ATXR6 are novel H3K27 monomethyltransferases required for chromatin structure and gene silencing

PubMed Central

Jacob, Yannick; Feng, Suhua; LeBlanc, Chantal A.; Bernatavichute, Yana V.; Stroud, Hume; Cokus, Shawn; Johnson, Lianna M.; Pellegrini, Matteo; Jacobsen, Steven E.; Michaels, Scott D.

2009-01-01

Constitutive heterochromatin in Arabidopsis thaliana is marked by repressive chromatin modifications including DNA methylation, histone H3 dimethylation at lysine 9 (H3K9me2), and monomethylation at lysine 27 (H3K27me1). The enzymes catalyzing DNA methylation and H3K9me2 have been identified and mutations in these proteins lead to the reactivation of silenced heterochromatic elements. The enzymes responsible for heterochromatic H3K27me1, in contrast, remain unknown. Here we show that the divergent SET-domain proteins ARABIDOPSIS TRITHORAX-RELATED PROTEIN5 (ATXR5) and ATXR6 exhibit H3K27 monomethyltransferase activity and double mutants have reduced H3K27me1 in vivo and show partial heterochromatin decondensation. Mutations in atxr5 and atxr6 also lead to transcriptional activation of repressed heterochromatic elements. Interestingly, H3K9me2 and DNA methylation are unaffected in the double mutant. These results indicate that ATXR5 and ATXR6 form a novel class of H3K27 methyltransferases and that H3K27me1 represents a new pathway required for transcriptional repression in Arabidopsis. PMID:19503079

An additional k-means clustering step improves the biological features of WGCNA gene co-expression networks.

PubMed

Botía, Juan A; Vandrovcova, Jana; Forabosco, Paola; Guelfi, Sebastian; D'Sa, Karishma; Hardy, John; Lewis, Cathryn M; Ryten, Mina; Weale, Michael E

2017-04-12

Weighted Gene Co-expression Network Analysis (WGCNA) is a widely used R software package for the generation of gene co-expression networks (GCN). WGCNA generates both a GCN and a derived partitioning of clusters of genes (modules). We propose k-means clustering as an additional processing step to conventional WGCNA, which we have implemented in the R package km2gcn (k-means to gene co-expression network, https://github.com/juanbot/km2gcn ). We assessed our method on networks created from UKBEC data (10 different human brain tissues), on networks created from GTEx data (42 human tissues, including 13 brain tissues), and on simulated networks derived from GTEx data. We observed substantially improved module properties, including: (1) few or zero misplaced genes; (2) increased counts of replicable clusters in alternate tissues (x3.1 on average); (3) improved enrichment of Gene Ontology terms (seen in 48/52 GCNs) (4) improved cell type enrichment signals (seen in 21/23 brain GCNs); and (5) more accurate partitions in simulated data according to a range of similarity indices. The results obtained from our investigations indicate that our k-means method, applied as an adjunct to standard WGCNA, results in better network partitions. These improved partitions enable more fruitful downstream analyses, as gene modules are more biologically meaningful.

The Utility of Naphthyl-Keratin Adducts as Biomarkers for Jet-Fuel Exposure

EPA Science Inventory

We investigated the association between biomarkers of dermal exposure, naphthyl-keratin adducts (NKA), and urine naphthalene biomarker levels in 105 workers routinely exposed to jet-fuel. A moderate correlation was observed between NKA and urine naphthalene levels (p = 0.061). Th...

A novel keratin18 promoter that drives reporter gene expression in the intrahepatic and extrahepatic biliary system allows isolation of cell-type specific transcripts from zebrafish liver

PubMed Central

Wilkins, Benjamin J.; Gong, Weilong; Pack, Michael

2015-01-01

Heritable and acquired biliary disorders are an important cause of acute and chronic human liver disease. Biliary development and physiology have been studied extensively in rodent models and more recently, zebrafish have been used to uncover pathogenic mechanisms and potential therapies for these conditions. Here we report development of novel transgenic lines labeling the intrahepatic and extrahepatic biliary system of zebrafish larvae that can be used for lineage tracing and isolation of biliary-specific RNAs from mixed populations of liver cells. We show that GFP expression driven by a 4.4 kilobase promoter fragment from the zebrafish keratin18 (krt18) gene allows visualization of all components of the developing biliary system as early as 3 days post-fertilization. In addition, expression of a ribosomal fusion protein (EGFP-Rpl10a) in krt18:TRAP transgenic fish allows for enrichment of translated biliary cell mRNAs via translating ribosome affinity purification (TRAP). Future studies utilizing these reagents will enhance our understanding of the morphologic and molecular processes involved in biliary development and disease. PMID:24394404

IBTK Differently Modulates Gene Expression and RNA Splicing in HeLa and K562 Cells.

PubMed

Fiume, Giuseppe; Scialdone, Annarita; Rizzo, Francesca; De Filippo, Maria Rosaria; Laudanna, Carmelo; Albano, Francesco; Golino, Gaetanina; Vecchio, Eleonora; Pontoriero, Marilena; Mimmi, Selena; Ceglia, Simona; Pisano, Antonio; Iaccino, Enrico; Palmieri, Camillo; Paduano, Sergio; Viglietto, Giuseppe; Weisz, Alessandro; Scala, Giuseppe; Quinto, Ileana

2016-11-07

The IBTK gene encodes the major protein isoform IBTKα that was recently characterized as substrate receptor of Cul3-dependent E3 ligase, regulating ubiquitination coupled to proteasomal degradation of Pdcd4, an inhibitor of translation. Due to the presence of Ankyrin-BTB-RCC1 domains that mediate several protein-protein interactions, IBTKα could exert expanded regulatory roles, including interaction with transcription regulators. To verify the effects of IBTKα on gene expression, we analyzed HeLa and K562 cell transcriptomes by RNA-Sequencing before and after IBTK knock-down by shRNA transduction. In HeLa cells, 1285 (2.03%) of 63,128 mapped transcripts were differentially expressed in IBTK -shRNA-transduced cells, as compared to cells treated with control-shRNA, with 587 upregulated (45.7%) and 698 downregulated (54.3%) RNAs. In K562 cells, 1959 (3.1%) of 63128 mapped RNAs were differentially expressed in IBTK -shRNA-transduced cells, including 1053 upregulated (53.7%) and 906 downregulated (46.3%). Only 137 transcripts (0.22%) were commonly deregulated by IBTK silencing in both HeLa and K562 cells, indicating that most IBTKα effects on gene expression are cell type-specific. Based on gene ontology classification, the genes responsive to IBTK are involved in different biological processes, including in particular chromatin and nucleosomal organization, gene expression regulation, and cellular traffic and migration. In addition, IBTK RNA interference affected RNA maturation in both cell lines, as shown by the evidence of alternative 3'- and 5'-splicing, mutually exclusive exons, retained introns, and skipped exons. Altogether, these results indicate that IBTK differently modulates gene expression and RNA splicing in HeLa and K562 cells, demonstrating a novel biological role of this protein.

IBTK Differently Modulates Gene Expression and RNA Splicing in HeLa and K562 Cells

PubMed Central

Fiume, Giuseppe; Scialdone, Annarita; Rizzo, Francesca; De Filippo, Maria Rosaria; Laudanna, Carmelo; Albano, Francesco; Golino, Gaetanina; Vecchio, Eleonora; Pontoriero, Marilena; Mimmi, Selena; Ceglia, Simona; Pisano, Antonio; Iaccino, Enrico; Palmieri, Camillo; Paduano, Sergio; Viglietto, Giuseppe; Weisz, Alessandro; Scala, Giuseppe; Quinto, Ileana

2016-01-01

The IBTK gene encodes the major protein isoform IBTKα that was recently characterized as substrate receptor of Cul3-dependent E3 ligase, regulating ubiquitination coupled to proteasomal degradation of Pdcd4, an inhibitor of translation. Due to the presence of Ankyrin-BTB-RCC1 domains that mediate several protein-protein interactions, IBTKα could exert expanded regulatory roles, including interaction with transcription regulators. To verify the effects of IBTKα on gene expression, we analyzed HeLa and K562 cell transcriptomes by RNA-Sequencing before and after IBTK knock-down by shRNA transduction. In HeLa cells, 1285 (2.03%) of 63,128 mapped transcripts were differentially expressed in IBTK-shRNA-transduced cells, as compared to cells treated with control-shRNA, with 587 upregulated (45.7%) and 698 downregulated (54.3%) RNAs. In K562 cells, 1959 (3.1%) of 63128 mapped RNAs were differentially expressed in IBTK-shRNA-transduced cells, including 1053 upregulated (53.7%) and 906 downregulated (46.3%). Only 137 transcripts (0.22%) were commonly deregulated by IBTK silencing in both HeLa and K562 cells, indicating that most IBTKα effects on gene expression are cell type-specific. Based on gene ontology classification, the genes responsive to IBTK are involved in different biological processes, including in particular chromatin and nucleosomal organization, gene expression regulation, and cellular traffic and migration. In addition, IBTK RNA interference affected RNA maturation in both cell lines, as shown by the evidence of alternative 3′- and 5′-splicing, mutually exclusive exons, retained introns, and skipped exons. Altogether, these results indicate that IBTK differently modulates gene expression and RNA splicing in HeLa and K562 cells, demonstrating a novel biological role of this protein. PMID:27827994

Multi-layered bird beaks: a finite-element approach towards the role of keratin in stress dissipation

PubMed Central

Soons, Joris; Herrel, Anthony; Genbrugge, Annelies; Adriaens, Dominique; Aerts, Peter; Dirckx, Joris

2012-01-01

Bird beaks are layered structures, which contain a bony core and an outer keratin layer. The elastic moduli of this bone and keratin were obtained in a previous study. However, the mechanical role and interaction of both materials in stress dissipation during seed crushing remain unknown. In this paper, a multi-layered finite-element (FE) model of the Java finch's upper beak (Padda oryzivora) is established. Validation measurements are conducted using in vivo bite forces and by comparing the displacements with those obtained by digital speckle pattern interferometry. Next, the Young modulus of bone and keratin in this FE model was optimized in order to obtain the smallest peak von Mises stress in the upper beak. To do so, we created a surrogate model, which also allows us to study the impact of changing material properties of both tissues on the peak stresses. The theoretically best values for both moduli in the Java finch are retrieved and correspond well with previous experimentally obtained values, suggesting that material properties are tuned to the mechanical demands imposed during seed crushing. PMID:22337628

Delineation of Matriptase Protein Expression by Enzymatic Gene Trapping Suggests Diverging Roles in Barrier Function, Hair Formation, and Squamous Cell Carcinogenesis

PubMed Central

List, Karin; Szabo, Roman; Molinolo, Alfredo; Nielsen, Boye Schnack; Bugge, Thomas H.

2006-01-01

The membrane serine protease matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma. PMID:16651618

Delineation of matriptase protein expression by enzymatic gene trapping suggests diverging roles in barrier function, hair formation, and squamous cell carcinogenesis.

PubMed

List, Karin; Szabo, Roman; Molinolo, Alfredo; Nielsen, Boye Schnack; Bugge, Thomas H

2006-05-01

The membrane serine protease matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma.

The non-anticoagulant heparin-like K5 polysaccharide derivative K5-N,OSepi attenuates myocardial ischaemia/reperfusion injury

PubMed Central

Collino, Massimo; Pini, Alessandro; Mastroianni, Rosanna; Benetti, Elisa; Lanzi, Cecilia; Bani, Daniele; Chini, Jacopo; Manoni, Marco; Fantozzi, Roberto; Masini, Emanuela

2012-01-01

Heparin and low molecular weight heparins have been demonstrated to reduce myocardial ischaemia/reperfusion (I/R) injury, although their use is hampered by the risk of haemorrhagic and thrombotic complications. Chemical and enzymatic modifications of K5 polysaccharide have shown the possibility of producing heparin-like compounds with low anticoagulant activity and strong anti-inflammatory effects. Using a rat model of regional myocardial I/R, we investigated the effects of an epimerized N-,O-sulphated K5 polysaccharide derivative, K5-N,OSepi, on infarct size and histological signs of myocardial injury caused by 30 min. ligature of the left anterior descending coronary artery followed by 1 or 24 h reperfusion. K5-N,OSepi (0.1–1 mg/kg given i.v. 15 min. before reperfusion) significantly reduced the extent of myocardial damage in a dose-dependent manner. Furthermore, we investigated the potential mechanism(s) of the cardioprotective effect(s) afforded by K5-N,OSepi. In left ventricular samples, I/R induced mast cell degranulation and a robust increase in lipid peroxidation, free radical-induced DNA damage and calcium overload. Markers of neutrophil infiltration and activation were also induced by I/R in rat hearts, specifically myeloperoxidase activity, intercellular-adhesion-molecule-1 expression, prostaglandin-E2 and tumour-necrosis-factor-α production. The robust increase in oxidative stress and inflammatory markers was blunted by K5-N,OSepi, in a dose-dependent manner, with maximum at 1 mg/kg. Furthermore, K5-N,OSepi administration attenuated the increase in caspase 3 activity, Bid and Bax activation and ameliorated the decrease in expression of Bcl-2 within the ischaemic myocardium. In conclusion, we demonstrate that the cardioprotective effect of the non-anticoagulant K5 derivative K5-N,OSepi is secondary to a combination of anti-apoptotic and anti-inflammatory effects. PMID:22248092

Arid5b facilitates chondrogenesis by recruiting the histone demethylase Phf2 to Sox9-regulated genes

NASA Astrophysics Data System (ADS)

Hata, Kenji; Takashima, Rikako; Amano, Katsuhiko; Ono, Koichiro; Nakanishi, Masako; Yoshida, Michiko; Wakabayashi, Makoto; Matsuda, Akio; Maeda, Yoshinobu; Suzuki, Yutaka; Sugano, Sumio; Whitson, Robert H.; Nishimura, Riko; Yoneda, Toshiyuki

2013-11-01

Histone modification, a critical step for epigenetic regulation, is an important modulator of biological events. Sox9 is a transcription factor critical for endochondral ossification; however, proof of its epigenetic regulation remains elusive. Here we identify AT-rich interactive domain 5b (Arid5b) as a transcriptional co-regulator of Sox9. Arid5b physically associates with Sox9 and synergistically induces chondrogenesis. Growth of Arid5b-/- mice is retarded with delayed endochondral ossification. Sox9-dependent chondrogenesis is attenuated in Arid5b-deficient cells. Arid5b recruits Phf2, a histone lysine demethylase, to the promoter region of Sox9 target genes and stimulates H3K9me2 demethylation of these genes. In the promoters of chondrogenic marker genes, H3K9me2 levels are increased in Arid5b-/- chondrocytes. Finally, we show that Phf2 knockdown inhibits Sox9-induced chondrocyte differentiation. Our findings establish an epigenomic mechanism of skeletal development, whereby Arid5b promotes chondrogenesis by facilitating Phf2-mediated histone demethylation of Sox9-regulated chondrogenic gene promoters.

Melanotic Translocation Renal Cell Carcinoma With a Novel ARID1B-TFE3 Gene Fusion.

PubMed

Antic, Tatjana; Taxy, Jerome B; Alikhan, Mir; Segal, Jeremy

2017-11-01

A 36-year-old male was found to have a 7.0 cm left upper pole renal mass on renal ultrasound. Following nephrectomy, the mass was grossly ill-demarcated, friable and red-brown, invading renal parenchyma, hilar fat and the renal vein. Microscopically, the tumor had a nested and papillary architecture. The cells demonstrated abundant clear and eosinophilic cytoplasm and focal intracytoplasmic melanin pigment. Nucleoli were prominent. By immunohistochemistry, the tumor was positive for TFE3; HMB-45 stained approximately 5% of tumor cells corresponding to the histologic melanin pigment, which was confirmed with Fontana-Masson stain with bleach. Immunostains for PAX8, CD10, MiTF, and CAIX were negative; keratins Cam 5.2 and AE1/AE3 were focally positive. Targeted next-generation sequencing revealed an ARID1B-TFE3 gene fusion. Melanotic Xp11 renal cell carcinoma is a rare, pigment containing translocation variant demonstrating overlapping features with melanoma and is usually associated with an SFPQ-TFE3 gene fusion. The patient is alive and without evidence of disease 7 years after his diagnosis. The combination of high grade histopathology, the presence of melanin, absent PAX8, keratin positivity, and relatively indolent clinical behavior with a unique translocation may warrant recognition as a distinct renal cell carcinoma translocation subtype.

An acellular dermal matrix allograft (Alloderm®) for increasing keratinized attached gingiva: A case series

PubMed Central

Agarwal, Chitra; Kumar, Baron Tarun; Mehta, Dhoom Singh

2015-01-01

Context: Adequate amount of keratinized gingiva is necessary to keep gingiva healthy and free of inflammation. Autografts have been used for years with great success to increase the width of attached gingiva. Autografts, however, have the disadvantage of increasing postoperative morbidity and improper color match with the adjacent tissues. Alloderm® allograft has been introduced as an alternative to autografts to overcome these disadvantages. Aim: In this study, the efficacy of alloderm® in increasing the width of attached gingiva and the stability of gained attached gingiva was evaluated clinically. Materials and Methods: Five patients with sites showing inadequate width of attached gingiva (≤1 mm) were enrolled for the study. The width of keratinized gingiva and other clinical parameters were recorded at baseline and 9th month postoperatively. Result: In all cases, there is the average increase of about 2.5 mm of attached gingiva and was maintained for 9-month. Percentage shrinkage of the graft is about 75% at the end of 3rd month in all cases. Excellent colors match with adjacent tissue has been obtained. Conclusion: The study signifies that Alloderm® results in an adequate increase in the amount of attached gingiva and therefore can be used successfully in place of autografts. PMID:26015676

Changes in nail keratin observed by Raman spectroscopy after Nd:YAG laser treatment.

PubMed

Shin, Min Kyung; Kim, Tae In; Kim, Wan Sun; Park, Hun-Kuk; Kim, Kyung Sook

2017-04-01

Lasers and photodynamic therapy have been considered a convergence treatment for onychomycosis, which is a fungal infection on the nail bed and nail plate. Laser therapies have shown satisfactory results without significant complications for onychomycosis; however, the mechanism of clearing remains unknown. In this work, we investigated changes in the chemical structure of nail keratin induced by Nd:YAG laser using Raman spectroscopy. Toe nails with onychomycosis were treated with 1064 nm Nd:YAG laser. After laser treatment, the disulfide band (490-590 cm -1 ) of nail keratin was rarely observed or was reduced in intensity. The amide I band (1500-1700 cm -1 ) also showed changes induced by the laser. The α-helical (1652 cm -1 ) structures dominated the β-sheet (1673 cm -1 ) in nontreated nail, but the opposite phenomenon was observed after laser treatment. © 2016 Wiley Periodicals, Inc.

Keratin 8 limits TLR-triggered inflammatory responses through inhibiting TRAF6 polyubiquitination

PubMed Central

Dong, Xiao-Ming; Liu, En-Dong; Meng, Yun-Xiao; Liu, Chao; Bi, Ya-Lan; Wu, Huan-Wen; Jin, Yan-Chao; Yao, Jing-Hui; Tang, Liu-Jun; Wang, Jian; Li, Min; Zhang, Chao; Yu, Miao; Zhan, Yi-Qun; Chen, Hui; Ge, Chang-Hui; Yang, Xiao-Ming; Li, Chang-Yan

2016-01-01

Toll-like receptors (TLRs) have critical roles in innate immunity and inflammation and the detailed mechanisms by which TLR signaling is fine tuned remain unclear. Keratin 8 (CK8) belongs to the type II keratin family and is the major compontent of the intermediate filaments of simple or single-layered epithelia. Here we report that down-regulation of CK8 in mice enhanced TLR-mediated responses, rendering mice more susceptible to lipopolysaccharide (LPS)-induced endotoxin shock and Escherichia coli–caused septic peritonitis with reduced survival, elevated levels of inflammation cytokines and more severe tissue damage. We found that CK8 suppressed TLR-induced nuclear factor (NF)-κB activation and interacted with the adaptor tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) to prevent its polyubiquitination. Our findings demonstrate a novel role of CK8 in negative regulation of TLR/NF-κB signaling and highlight a previously unidentified nonclassical function for CK8 in limiting inflammatory responses. PMID:27586056

Structural genes for thiamine biosynthetic enzymes (thiCEFGH) in Escherichia coli K-12.

PubMed Central

Vander Horn, P B; Backstrom, A D; Stewart, V; Begley, T P

1993-01-01

Escherichia coli K-12 synthesizes thiamine pyrophosphate (vitamin B1) de novo. Two precursors [4-methyl-5-(beta-hydroxyethyl)thiazole monophosphate and 4-amino-5-hydroxymethyl-2-methylpyrimidine pyrophosphate] are coupled to form thiamine monophosphate, which is then phosphorylated to make thiamine pyrophosphate. Previous studies have identified two classes of thi mutations, clustered at 90 min on the genetic map, which result in requirements for the thiazole or the hydroxymethylpryimidine. We report here our initial molecular genetic analysis of the thi cluster. We cloned the thi cluster genes and examined their organization, structure, and function by a combination of phenotypic testing, complementation analysis, polypeptide expression, and DNA sequencing. We found five tightly linked genes, designated thiCEFGH. The thiC gene product is required for the synthesis of the hydroxymethylpyrimidine. The thiE, thiF, thiG, and thiH gene products are required for synthesis of the thiazole. These mutants did not respond to 1-deoxy-D-threo-2-pentulose, indicating that they are blocked in the conversion of this precursor compound to the thiazole itself. Images PMID:8432721

MoSET1 (Histone H3K4 Methyltransferase in Magnaporthe oryzae) Regulates Global Gene Expression during Infection-Related Morphogenesis

PubMed Central

Pham, Kieu Thi Minh; Inoue, Yoshihiro; Vu, Ba Van; Nguyen, Hanh Hieu; Nakayashiki, Toru; Ikeda, Ken-ichi; Nakayashiki, Hitoshi

2015-01-01

Here we report the genetic analyses of histone lysine methyltransferase (KMT) genes in the phytopathogenic fungus Magnaporthe oryzae. Eight putative M. oryzae KMT genes were targeted for gene disruption by homologous recombination. Phenotypic assays revealed that the eight KMTs were involved in various infection processes at varying degrees. Moset1 disruptants (Δmoset1) impaired in histone H3 lysine 4 methylation (H3K4me) showed the most severe defects in infection-related morphogenesis, including conidiation and appressorium formation. Consequently, Δmoset1 lost pathogenicity on wheat host plants, thus indicating that H3K4me is an important epigenetic mark for infection-related gene expression in M. oryzae. Interestingly, appressorium formation was greatly restored in the Δmoset1 mutants by exogenous addition of cAMP or of the cutin monomer, 16-hydroxypalmitic acid. The Δmoset1 mutants were still infectious on the super-susceptible barley cultivar Nigrate. These results suggested that MoSET1 plays roles in various aspects of infection, including signal perception and overcoming host-specific resistance. However, since Δmoset1 was also impaired in vegetative growth, the impact of MoSET1 on gene regulation was not infection specific. ChIP-seq analysis of H3K4 di- and tri-methylation (H3K4me2/me3) and MoSET1 protein during infection-related morphogenesis, together with RNA-seq analysis of the Δmoset1 mutant, led to the following conclusions: 1) Approximately 5% of M. oryzae genes showed significant changes in H3K4-me2 or -me3 abundance during infection-related morphogenesis. 2) In general, H3K4-me2 and -me3 abundance was positively associated with active transcription. 3) Lack of MoSET1 methyltransferase, however, resulted in up-regulation of a significant portion of the M. oryzae genes in the vegetative mycelia (1,491 genes), and during infection-related morphogenesis (1,385 genes), indicating that MoSET1 has a role in gene repression either directly or more

The i5k Workspace@NAL—enabling genomic data access, visualization and curation of arthropod genomes

PubMed Central

Poelchau, Monica; Childers, Christopher; Moore, Gary; Tsavatapalli, Vijaya; Evans, Jay; Lee, Chien-Yueh; Lin, Han; Lin, Jun-Wei; Hackett, Kevin

2015-01-01

The 5000 arthropod genomes initiative (i5k) has tasked itself with coordinating the sequencing of 5000 insect or related arthropod genomes. The resulting influx of data, mostly from small research groups or communities with little bioinformatics experience, will require visualization, dissemination and curation, preferably from a centralized platform. The National Agricultural Library (NAL) has implemented the i5k Workspace@NAL (http://i5k.nal.usda.gov/) to help meet the i5k initiative's genome hosting needs. Any i5k member is encouraged to contact the i5k Workspace with their genome project details. Once submitted, new content will be accessible via organism pages, genome browsers and BLAST search engines, which are implemented via the open-source Tripal framework, a web interface for the underlying Chado database schema. We also implement the Web Apollo software for groups that choose to curate gene models. New content will add to the existing body of 35 arthropod species, which include species relevant for many aspects of arthropod genomic research, including agriculture, invasion biology, systematics, ecology and evolution, and developmental research. PMID:25332403

NMR spectroscopy reveals the presence and association of lipids and keratin in adhesive gecko setae

PubMed Central

Jain, Dharamdeep; Stark, Alyssa Y.; Niewiarowski, Peter H.; Miyoshi, Toshikazu; Dhinojwala, Ali

2015-01-01

Lipid and protein aggregates are one of the fundamental materials of biological systems. Examples include cell membranes, insect cuticle, vertebrate epidermis, feathers, hair and adhesive structures known as ‘setae’ on gecko toes. Until recently gecko setae were assumed to be composed entirely of keratin, but analysis of footprints left behind by geckos walking on surfaces revealed that setae include various kinds of lipids. However, the arrangement and molecular-level behavior of lipids and keratin in the setae is still not known. In the present study we demonstrate, for the first time, the use of Nuclear Magnetic Resonance (NMR) spectroscopy techniques to confirm the presence of lipids and investigate their association with keratin in ‘pristine' sheds, or natural molts of the adhesive toe pad and non-adhesive regions of the skin. Analysis was also carried on the sheds after they were ‘delipidized’ to remove surface lipids. Our results show a distribution of similar lipids in both the skin and toe shed but with different dynamics at a molecular level. The present study can help us understand the gecko system both biologically and for design of synthetic adhesives, but the findings may be relevant to the characteristics of lipid-protein interactions in other biological systems. PMID:25902194

Medium Calcium Concentration Determines Keratin Intermediate Filament Density and Distribution in Immortalized Cultured Thymic Epithelial Cells (TECs)

NASA Astrophysics Data System (ADS)

Sands, Sandra S.; Meek, William D.; Hayashi, Jun; Ketchum, Robert J.

2005-08-01

Isolation and culture of thymic epithelial cells (TECs) using conventional primary tissue culture techniques under conditions employing supplemented low calcium medium yielded an immortalized cell line derived from the LDA rat (Lewis [Rt1l] cross DA [Rt1a]) that could be manipulated in vitro. Thymi were harvested from 4 5-day-old neonates, enzymically digested using collagenase (1 mg/ml, 37°C, 1 h) and cultured in low calcium WAJC404A medium containing cholera toxin (20 ng/ml), dexamethasone (10 nM), epidermal growth factor (10 ng/ml), insulin (10 [mu]g/ml), transferrin (10 [mu]g/ml), 2% calf serum, 2.5% Dulbecco's Modified Eagle's Medium (DMEM), and 1% antibiotic/antimycotic. TECs cultured in low calcium displayed round to spindle-shaped morphology, distinct intercellular spaces (even at confluence), and dense reticular-like keratin patterns. In high calcium (0.188 mM), TECs formed cobblestone-like confluent monolayers that were resistant to trypsinization (0.05%) and displayed keratin intermediate filaments concentrated at desmosomal junctions between contiguous cells. Changes in cultured TEC morphology were quantified by an analysis of desmosome/membrane relationships in high and low calcium media. Desmosomes were significantly increased in the high calcium medium. These studies may have value when considering the growth conditions of cultured primary cell lines like TECs.

Testosterone regulates keratin 33B expression in rat penis growth through androgen receptor signaling

PubMed Central

Ma, Yan-Min; Wu, Kai-Jie; Dang, Qiang; Shi, Qi; Gao, Yang; Guo, Peng; Xu, Shan; Wang, Xin-Yang; He, Da-Lin; Gong, Yong-Guang

2014-01-01

Androgen therapy is the mainstay of treatment for the hypogonadotropic hypogonadal micropenis because it obviously enhances penis growth in prepubescent microphallic patients. However, the molecular mechanisms of androgen treatment leading to penis growth are still largely unknown. To clarify this well-known phenomenon, we successfully generated a castrated male Sprague Dawley rat model at puberty followed by testosterone administration. Interestingly, compared with the control group, testosterone treatment stimulated a dose-dependent increase of penis weight, length, and width in castrated rats accompanied with a dramatic recovery of the pathological changes of the penis. Mechanistically, testosterone administration substantially increased the expression of androgen receptor (AR) protein. Increased AR protein in the penis could subsequently initiate transcription of its target genes, including keratin 33B (Krt33b). Importantly, we demonstrated that KRT33B is generally expressed in the rat penis and that most KRT33B expression is cytoplasmic. Furthermore, AR could directly modulate its expression by binding to a putative androgen response element sequence of the Krt33b promoter. Overall, this study reveals a novel mechanism facilitating penis growth after testosterone treatment in precastrated prepubescent animals, in which androgen enhances the expression of AR protein as well as its target genes, such as Krt33b. PMID:24994782

Testosterone regulates keratin 33B expression in rat penis growth through androgen receptor signaling.

PubMed

Ma, Yan-Min; Wu, Kai-Jie; Dang, Qiang; Shi, Qi; Gao, Yang; Guo, Peng; Xu, Shan; Wang, Xin-Yang; He, Da-Lin; Gong, Yong-Guang

2014-01-01

Androgen therapy is the mainstay of treatment for the hypogonadotropic hypogonadal micropenis because it obviously enhances penis growth in prepubescent microphallic patients. However, the molecular mechanisms of androgen treatment leading to penis growth are still largely unknown. To clarify this well-known phenomenon, we successfully generated a castrated male Sprague Dawley rat model at puberty followed by testosterone administration. Interestingly, compared with the control group, testosterone treatment stimulated a dose-dependent increase of penis weight, length, and width in castrated rats accompanied with a dramatic recovery of the pathological changes of the penis. Mechanistically, testosterone administration substantially increased the expression of androgen receptor (AR) protein. Increased AR protein in the penis could subsequently initiate transcription of its target genes, including keratin 33B (Krt33b). Importantly, we demonstrated that KRT33B is generally expressed in the rat penis and that most KRT33B expression is cytoplasmic. Furthermore, AR could directly modulate its expression by binding to a putative androgen response element sequence of the Krt33b promoter. Overall, this study reveals a novel mechanism facilitating penis growth after testosterone treatment in precastrated prepubescent animals, in which androgen enhances the expression of AR protein as well as its target genes, such as Krt33b.

Simultaneous detection of 5-fluorocytosine and 5-fluorouracil in human cells carrying CD/5-FC suicide gene system by using capillary zone electrophoresis.

PubMed

Liu, Yajing; Zhu, Pan; Huang, Zhiwei; Zhou, Li; Shi, Ping

2018-02-15

A well-known suicide gene therapy approach, cytosine deaminase (CD) in combination with prodrug 5-flurocytosine (5-FC), has become an effective strategy of tumor treatment. However, there are short of simple and convenient detection methods to evaluate the efficiency of 5-FC conversion to 5-fluorouracil (5-FU) in human cells carrying various CD/5-FC systems. In this study, we developed an effective capillary zone electrophoresis (CZE) method to simultaneously measure 5-FC and 5-FU in cells carrying CD/5-FC suicide gene system. Under the condition of 60 mM borate buffer (pH 9.5) and 25 kV separation voltage with 0.5 psi × 15 s injection in 210 nm, the separation of 5-FC and 5-FU could be completely achieved within 15 min. The linearity of the calibration curve of standard 5-FC and 5-FU was in the range from 1 to 1000 μM (r 2  > 0.999) and their recoveries were 98.4% and 96.0%, respectively. Due to the simple sample preparation and easy detection, this method is suitable for the study of the conversion efficiency of CD/5-FC suicide gene system. It aims to intuitively evaluate CD/5-FC systems and helps to guide the improvement of more effective CD/5-FC suicide gene systems. Copyright © 2018. Published by Elsevier B.V.

The 5-kW arcjet power electronics

NASA Technical Reports Server (NTRS)

Gruber, R. P.; Gott, R. W.; Haag, T. W.

1989-01-01

The initial design and evaluation of a 5 kW arcjet power electronics breadboard which as been integrated with a modified 1 kW design laboratory arcjet is presented. A single stage, 5 kW full bridge, pulse width modulated (PWM), power converter was developed which was phase shift regulated. The converter used metal oxide semiconductor field effect transistor (MOSFET) power switches and incorporated current mode control and an integral arcjet pulse ignition circuit. The unoptimized power efficiency was 93.5 and 93.9 percent at 5 kW and 50A output at input voltages of 130 and 150V, respectively. Line and load current regulation at 50A output was within one percent. The converter provided up to 6.6 kW to the arcjet with simulated ammonia used as a propellant.

Avian malaria in a boreal resident species: long-term temporal variability, and increased prevalence in birds with avian keratin disorder

USGS Publications Warehouse

Wilkinson, Laura C.; Handel, Colleen M.; Van Hemert, Caroline R.; Loiseau, Claire; Sehgal, Ravinder N. M.

2016-01-01

The prevalence of vector-borne parasitic diseases is widely influenced by biological and ecological factors. Environmental conditions such as temperature and precipitation can have a marked effect on haemosporidian parasites (Plasmodium spp.) that cause malaria and those that cause other malaria-like diseases in birds. However, there have been few long-term studies monitoring haemosporidian infections in birds in northern latitudes, where weather conditions can be highly variable and the effects of climate change are becoming more pronounced. We used molecular methods to screen more than 2,000 blood samples collected from black-capped chickadees (Poecile atricapillus), a resident passerine bird. Samples were collected over a 10 year period, mostly during the non-breeding season, at seven sites in Alaska, USA. We tested for associations between Plasmodium prevalence and local environmental conditions including temperature, precipitation, site, year and season. We also evaluated the relationship between parasite prevalence and individual host factors of age, sex and presence or absence of avian keratin disorder. This disease, which causes accelerated keratin growth in the beak, provided a natural study system in which to test the interaction between disease state and malaria prevalence. Prevalence of Plasmodium infection varied by year, site, age and individual disease status but there was no support for an effect of sex or seasonal period. Significantly, birds with avian keratin disorder were 2.6 times more likely to be infected by Plasmodium than birds without the disorder. Interannual variation in the prevalence of Plasmodium infection at different sites was positively correlated with summer temperatures at the local but not statewide scale. Sequence analysis of the parasite cytochrome b gene revealed a single Plasmodiumspp. lineage, P43. Our results demonstrate associations between prevalence of avian malaria and a variety of biological and

Multicenter Clinical Trial of Keratin Biomaterial for Peripheral Nerve Regeneration

DTIC Science & Technology

2015-12-01

Radiological Health ( CDRH ) to clarify the designation of the hydrogel. As a result of this meeting, steps required for an IND for the keratin...the Center for Biologics Evaluation Research (CBER), and the Center for Devices and Radiological Health 8 ( CDRH ) to clarify the designation of the...application to the CDRH for a new product. This new product is the material that is produced in the validated manufacturing facility at KeraNetics. This

H3K27me3 and H3K4me3 chromatin environment at super-induced dehydration stress memory genes of Arabidopsis thaliana.

PubMed

Liu, Ning; Fromm, Michael; Avramova, Zoya

2014-03-01

Pre-exposure to a stress may alter the plant's cellular, biochemical, and/or transcriptional responses during future encounters as a 'memory' from the previous stress. Genes increasing transcription in response to a first dehydration stress, but producing much higher transcript levels in a subsequent stress, represent the super-induced 'transcription memory' genes in Arabidopsis thaliana. The chromatin environment (histone H3 tri-methylations of Lys 4 and Lys 27, H3K4me3, and H3K27me3) studied at five dehydration stress memory genes revealed existence of distinct memory-response subclasses that responded differently to CLF deficiency and displayed different transcriptional activities during the watered recovery periods. Among the most important findings is the novel aspect of the H3K27me3 function observed at specific dehydration stress memory genes. In contrast to its well-known role as a chromatin repressive mechanism at developmentally regulated genes, H3K27me3 did not prevent transcription from the dehydration stress-responding genes. The high H3K27me3 levels present during transcriptionally inactive states did not interfere with the transition to active transcription and with H3K4me3 accumulation. H3K4me3 and H3K27me3 marks function independently and are not mutually exclusive at the dehydration stress-responding memory genes.

Escherichia coli K-12 and B contain functional bacteriophage P2 ogr genes.

PubMed Central

Slettan, A; Gebhardt, K; Kristiansen, E; Birkeland, N K; Lindqvist, B H

1992-01-01

The bacteriophage P2 ogr gene encodes an essential 72-amino-acid protein which acts as a positive regulator of P2 late transcription. A P2 ogr deletion phage, which depends on the supply of Ogr protein in trans for lytic growth on Escherichia coli C, has previously been constructed. E. coli B and K-12 were found to support the growth of the ogr-defective P2 phage because of the presence of functional ogr genes located in cryptic P2-like prophages in these strains. The cryptic ogr genes were cloned and sequenced. Compared with the P2 wild-type ogr gene, the ogr genes in the B and K-12 strains are conserved, containing mostly silent base substitutions. One of the base substitutions in the K-12 ogr gene results in replacement of an alanine with valine at position 57 in the Ogr protein but does not seem to affect the function of Ogr as a transcriptional activator. The cryptic ogr genes are constitutively transcribed, apparently at a higher level than the wild-type ogr gene in a P2 lysogen. Images PMID:1597424

Colour-producing β-keratin nanofibres in blue penguin (Eudyptula minor) feathers

PubMed Central

D'Alba, Liliana; Saranathan, Vinodkumar; Clarke, Julia A.; Vinther, Jakob A.; Prum, Richard O.; Shawkey, Matthew D.

2011-01-01

The colours of living organisms are produced by the differential absorption of light by pigments (e.g. carotenoids, melanins) and/or by the physical interactions of light with biological nanostructures, referred to as structural colours. Only two fundamental morphologies of non-iridescent nanostructures are known in feathers, and recent work has proposed that they self-assemble by intracellular phase separation processes. Here, we report a new biophotonic nanostructure in the non-iridescent blue feather barbs of blue penguins (Eudyptula minor) composed of parallel β-keratin nanofibres organized into densely packed bundles. Synchrotron small angle X-ray scattering and two-dimensional Fourier analysis of electron micrographs of the barb nanostructure revealed short-range order in the organization of fibres at the appropriate size scale needed to produce the observed colour by coherent scattering. These two-dimensional quasi-ordered penguin nanostructures are convergent with similar arrays of parallel collagen fibres in avian and mammalian skin, but constitute a novel morphology for feathers. The identification of a new class of β-keratin nanostructures adds significantly to the known mechanisms of colour production in birds and suggests additional complexity in their self-assembly. PMID:21307042

Colour-producing [beta]-keratin nanofibres in blue penguin (Eudyptula minor) feathers

DOE Office of Scientific and Technical Information (OSTI.GOV)

D; Alba, Liliana; Saranathan, Vinodkumar

2012-03-26

The colours of living organisms are produced by the differential absorption of light by pigments (e.g. carotenoids, melanins) and/or by the physical interactions of light with biological nanostructures, referred to as structural colours. Only two fundamental morphologies of non-iridescent nanostructures are known in feathers, and recent work has proposed that they self-assemble by intracellular phase separation processes. Here, we report a new biophotonic nanostructure in the non-iridescent blue feather barbs of blue penguins (Eudyptula minor) composed of parallel {beta}-keratin nanofibres organized into densely packed bundles. Synchrotron small angle X-ray scattering and two-dimensional Fourier analysis of electron micrographs of the barbmore » nanostructure revealed short-range order in the organization of fibres at the appropriate size scale needed to produce the observed colour by coherent scattering. These two-dimensional quasi-ordered penguin nanostructures are convergent with similar arrays of parallel collagen fibres in avian and mammalian skin, but constitute a novel morphology for feathers. The identification of a new class of {beta}-keratin nanostructures adds significantly to the known mechanisms of colour production in birds and suggests additional complexity in their self-assembly.« less

Gene expression profiling for nitric oxide prodrug JS-K to kill HL-60 myeloid leukemia cells.

PubMed

Liu, Jie; Malavya, Swati; Wang, Xueqian; Saavedra, Joseph E; Keefer, Larry K; Tokar, Erik; Qu, Wei; Waalkes, Michael P; Shami, Paul J

2009-07-01

The nitric oxide (NO) prodrug JS-K is shown to have anticancer activity. To profile the molecular events associated with the anticancer effects of JS-K, HL-60 leukemia cells were treated with JS-K and subjected to microarray and real-time RT-PCR analysis. JS-K induced concentration- and time-dependent gene expression changes in HL-60 cells corresponding to the cytolethality effects. The apoptotic genes (caspases, Bax, and TNF-alpha) were induced, and differentiation-related genes (CD14, ITGAM, and VIM) were increased. For acute phase protein genes, some were increased (TP53, JUN) while others were suppressed (c-myc, cyclin E). The expression of anti-angiogenesis genes THBS1 and CD36 and genes involved in tumor cell migration such as tissue inhibitors of metalloproteinases, were also increased by JS-K. Confocal analysis confirmed key gene changes at the protein levels. Thus, multiple molecular events are associated with JS-K effects in killing HL-60, which could be molecular targets for this novel anticancer NO prodrug.

A role for repressive complexes and H3K9 di-methylation in PRDM5-associated brittle cornea syndrome.

PubMed

Porter, Louise F; Galli, Giorgio G; Williamson, Sally; Selley, Julian; Knight, David; Elcioglu, Nursel; Aydin, Ali; Elcioglu, Mustafa; Venselaar, Hanka; Lund, Anders H; Bonshek, Richard; Black, Graeme C; Manson, Forbes D

2015-12-01

Type 2 brittle cornea syndrome (BCS2) is an inherited connective tissue disease with a devastating ocular phenotype caused by mutations in the transcription factor PR domain containing 5 (PRDM5) hypothesized to exert epigenetic effects through histone and DNA methylation. Here we investigate clinical samples, including skin fibroblasts and retinal tissue from BCS2 patients, to elucidate the epigenetic role of PRDM5 and mechanisms of its dysregulation in disease. First we report abnormal retinal vascular morphology in the eyes of two cousins with BCS2 (PRDM5 Δ exons 9-14) using immunohistochemistry, and mine data from skin fibroblast expression microarrays from patients with PRDM5 mutations p.Arg590* and Δ exons 9-14, as well as from a PRDM5 ChIP-sequencing experiment. Gene ontology analysis of dysregulated PRDM5-target genes reveals enrichment for extracellular matrix (ECM) genes supporting vascular integrity and development. Q-PCR and ChIP-qPCR confirm upregulation of critical mediators of ECM stability in vascular structures (COL13A1, COL15A1, NTN1, CDH5) in patient fibroblasts. We identify H3K9 di-methylation (H3K9me2) at these PRDM5-target genes in fibroblasts, and demonstrate that the BCS2 mutation p.Arg83Cys diminishes interaction of PRDM5 with repressive complexes, including NuRD complex protein CHD4, and the repressive chromatin interactor HP1BP3, by co-immunoprecipitation combined with mass spectrometry. We observe reduced heterochromatin protein 1 binding protein 3 (HP1BP3) staining in the retinas of two cousins lacking exons 9-14 by immunohistochemistry, and dysregulated H3K9me2 in skin fibroblasts of three patients (p.Arg590*, p.Glu134* and Δ exons 9-14) by western blotting. These findings suggest that defective interaction of PRDM5 with repressive complexes, and dysregulation of H3K9me2, play a role in PRDM5-associated disease. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Molecular analysis of the genes involved in the biosynthesis of serotype specific polysaccharide in the novel serotype k strains of Streptococcus mutans.

PubMed

Nomura, R; Nakano, K; Ooshima, T

2005-10-01

We previously reported the new serotype k of Streptococcus mutans, which, compared to serotypes c, e, and f, features a drastic reduction in the length of the glucose side chain linked to the rhamnose backbone of the serotype specific polysaccharide. The 5' region of the rgpF gene of serotype k strains contains a distinctive nucleotide sequence, which suggests that an alteration of the rgpF gene in serotype k strains may explain the shortened glucose side chain. However, in the present study, expression of the rgpF gene of MT8148 (serotype c) in serotype k isolates was not found to lead to serotype conversion. Furthermore, mRNA expression of rgpE, known to be associated with glucose side chain formation, was not detected in any of the tested serotype k isolates with an RT-PCR method. The nucleotide alignment of all genes known to be involved in the biosynthesis of serotype specific polysaccharide in serotype k strains was shown to be quite similar to that of serotype c strains, as compared to serotype e and f strains, especially in the region downstream of rgpF. Our results indicate that the common characteristics of serotype k isolates may be caused by a lack of expression of the gene involved in glucose side chain formation.

Production of Proteolytic Enzymes by a Keratin-Degrading Aspergillus niger

PubMed Central

Lopes, Fernanda Cortez; Silva, Lucas André Dedavid e; Tichota, Deise Michele; Daroit, Daniel Joner; Velho, Renata Voltolini; Pereira, Jamile Queiroz; Corrêa, Ana Paula Folmer; Brandelli, Adriano

2011-01-01

A fungal isolate with capability to grow in keratinous substrate as only source of carbon and nitrogen was identified as Aspergillus niger using the sequencing of the ITS region of the rDNA. This strain produced a slightly acid keratinase and an acid protease during cultivation in feather meal. The peak of keratinolytic activity occurred in 48 h and the maximum proteolytic activity in 96 h. These enzymes were partly characterized as serine protease and aspartic protease, respectively. The effects of feather meal concentration and initial pH on enzyme production were evaluated using a central composite design combined with response surface methodology. The optimal conditions were determined as pH 5.0 for protease and 7.8 for keratinase and 20 g/L of feather meal, showing that both models were predictive. Production of keratinases by A. niger is a less-exploited field that might represent a novel and promising biotechnological application for this microorganism. PMID:22007293

Vitamin K2 alleviates type 2 diabetes in rats by induction of osteocalcin gene expression.

PubMed

Hussein, Atef G; Mohamed, Randa H; Shalaby, Sally M; Abd El Motteleb, Dalia M

2018-03-01

The biological mechanisms behind the association between vitamin K (Vit K) and glucose metabolism are uncertain. We aimed to analyze the expression of insulin 1 (Ins 1), insulin 2 (Ins 2) and cyclin D2, the expression of adiponectin and UCP-1 . In addition, we aimed to estimate the doses of Vit K2 able to affect various aspects of glucose and energy metabolism in type 2 diabetes. Thirty adult male rats were allocated equally into five groups: control group, diabetes mellitus group, and groups 3, 4, and 5, which received Vit K 2 at three daily dose levels (10, 15, and 30 mg/kg, respectively) for 8 wk. At the end of the study, blood samples were collected to quantify total osteocalcin, fasting plasma glucose, fasting insulin, and relevant variables. The expression of OC, Ins 1, Ins 2, cyclin D2, adiponectin, UCP-1 genes was analyzed by real-time polymerase chain reaction. After administration of Vit K 2 , a dose-dependent decrease in fasting plasma glucose, hemoglobin A1c and homeostatic model assessment method insulin resistance, and a dose-dependent increase in fasting insulin and homeostatic model assessment method β cell function levels, when compared with diabetes mellitus rats, were detected. There was significant upregulation of OC, Ins 1, Ins 2, or cyclin D2 gene expression in the three treated groups in a dose-dependent manner when compared with the diabetic rats. However, expression of adiponectin and UCP-1 were significantly increased at the highest dose (30 mg/kg daily) only. Vit K 2 administration could improve glycemic status in type 2 diabetic rats by induction of OC gene expression. Osteocalcin could increase β-cell proliferation, energy expenditure, and adiponectin expression. Different concentrations of Vit K 2 were required to affect glucose metabolism and insulin sensitivity. Copyright © 2017 Elsevier Inc. All rights reserved.

Plasminogen fragments K 1-3 and K 5 bind to different sites in fibrin fragment DD.

PubMed

Grinenko, T V; Kapustianenko, L G; Yatsenko, T A; Yusova, O I; Rybachuk, V N

2016-01-01

Specific plasminogen-binding sites of fibrin molecule are located in Аα148-160 regions of C-terminal domains. Plasminogen interaction with these sites initiates the activation process of proenzyme and subsequent fibrin lysis. In this study we investigated the binding of plasminogen fragments K 1-3 and K 5 with fibrin fragment DD and their effect on Glu-plasminogen interaction with DD. It was shown that the level of Glu-plasminogen binding to fibrin fragment DD is decreased by 50-60% in the presence of K 1-3 and K 5. Fragments K 1-3 and K 5 have high affinity to fibrin fragment DD (Kd is 0.02 for K 1-3 and 0.054 μМ for K 5). K 5 interaction is independent and K 1-3 is partly dependent on C-terminal lysine residues. K 1-3 interacts with complex of fragment DD-immobilized K 5 as well as K 5 with complex of fragment DD-immobilized K 1-3. The plasminogen fragments do not displace each other from binding sites located in fibrin fragment DD, but can compete for the interaction. The results indicate that fibrin fragment DD contains different binding sites for plasminogen kringle fragments K 1-3 and K 5, which can be located close to each other. The role of amino acid residues of fibrin molecule Аα148-160 region in interaction with fragments K 1-3 and K 5 is discussed.

The CBL-Interacting Protein Kinase CIPK23 Regulates HAK5-Mediated High-Affinity K+ Uptake in Arabidopsis Roots.

PubMed

Ragel, Paula; Ródenas, Reyes; García-Martín, Elena; Andrés, Zaida; Villalta, Irene; Nieves-Cordones, Manuel; Rivero, Rosa M; Martínez, Vicente; Pardo, Jose M; Quintero, Francisco J; Rubio, Francisco

2015-12-01

Plant growth and development requires efficient acquisition of essential elements. Potassium (K(+)) is an important macronutrient present in the soil solution at a wide range of concentrations. Regulation of the K(+) uptake systems in the roots is essential to secure K(+) supply. It has been shown in Arabidopsis (Arabidopsis thaliana) that when the external K(+) concentration is very low (<10 µm), K(+) nutrition depends exclusively on the high-affinity K(+) transporter5 (HAK5). Low-K(+)-induced transcriptional activation of the gene encoding HAK5 has been previously reported. Here, we show the posttranscriptional regulation of HAK5 transport activity by phosphorylation. Expression in a heterologous system showed that the Ca(2+) sensors calcineurin B-like (CBL1), CBL8, CBL9, and CBL10, together with CBL-interacting protein kinase23 (CIPK23), activated HAK5 in vivo. This activation produced an increase in the affinity and the Vmax of K(+) transport. In vitro experiments show that the N terminus of HAK5 is phosphorylated by CIPK23. This supports the idea that phosphorylation of HAK5 induces a conformational change that increases its affinity for K(+). Experiments of K(+) (Rb(+)) uptake and growth measurements in low-K(+) medium with Arabidopsis single mutants hak5, akt1, and cipk23, double mutants hak5 akt1, hak5 cipk23, and akt1 cipk23, and the triple mutant hak5 akt1 cipk23 confirmed the regulatory role of CIPK23 in planta. © 2015 American Society of Plant Biologists. All Rights Reserved.

A clinical and histologic evaluation of gingival fibroblasts seeding on a chitosan-based scaffold and its effect on the width of keratinized gingiva in dogs.

PubMed

Lotfi, Ghogha; Shokrgozar, Mohammad Ali; Mofid, Rasoul; Abbas, Fatemeh Mashhadi; Ghanavati, Farzin; Bagheban, Alireza Akbarzadeh; Shariati, Ramin Pajoum

2011-09-01

Finding biocompatible matrix materials capable of enhancing the procedures of gingival augmentation is a major concern in periodontal research. This has prompted the investigation of a safe grafting technique by means of synthetic or natural polymers. The objective of this study is to examine the effect of a gingival fibroblast cultured on a naturally derived (i.e., chitosan-based) scaffold on the width of keratinized gingiva in dogs. Gingival fibroblasts were cultured from a small portion of hard palates of five dogs. A bilayered chitosan scaffold was seeded with the gingival fibroblasts and transferred to dogs. Surgery was performed bilaterally, and the regions were randomly divided into two groups: chitosan only (control site) and chitosan + fibroblast (test site). Periodontal parameters, including probing depth and width of keratinized and attached gingiva, were measured at baseline and 3 months after surgery. A histologic evaluation was also performed on the healed grafted sites. Comparison of width of keratinized and attached gingiva in control and test sites showed that the mean width of keratinized and attached gingiva increased in each group after surgery. However, the difference between control and test groups was not statistically significant. Concerning the existence of the keratinized epithelium, exocytosis, and epithelium thickness, no significant difference was observed in test and control sites. The difference was significant in relation to rete ridge formation. The tissue-engineered graft consisting of chitosan + fibroblast was applied to gingival augmentation procedures and generated keratinized tissue without any complications usually associated with donor-site surgery.

The Molecular Architecture for the Intermediate Filaments of Hard α -Keratin Based on the Superlattice Data Obtained from a Study of Mammals Using Synchrotron Fibre Diffraction

DOE Office of Scientific and Technical Information (OSTI.GOV)

James, Veronica

High- and low-angle X-ray diffraction studies of hard α -keratin have been studied, and various models have been proposed over the last 70 years. Most of these studies have been confined to one or two forms of alpha keratin. This high- and low-angle synchrotron fibre diffraction study extends the study to cover all available data for all known forms of hard α -keratin including hairs, fingernails, hooves, horn, and quills from mammals, marsupials, and a monotreme, and it confirms that the model proposed is universally acceptable for all mammals. A complete Bragg analysis of the meridional diffraction patterns, including multiple-timemore » exposures to verify any weak reflections, verified the existence of a superlattice consisting of two infinite lattices and three finite lattices. An analysis of the equatorial patterns establishes the radii of the oligomeric levels of dimers, tetramers, and intermediate filaments (IFs) together with the centre to centre distance for the IFs, thus confirming the proposed helices within helices molecular architecture for hard α -keratin. The results verify that the structure proposed by Feughelman and James meets the criteria for a valid α -keratin structure.« less

The Molecular Architecture for the Intermediate Filaments of Hard α -Keratin Based on the Superlattice Data Obtained from a Study of Mammals Using Synchrotron Fibre Diffraction

DOE PAGES

James, Veronica

2011-01-01

High- and low-angle X-ray diffraction studies of hard α -keratin have been studied, and various models have been proposed over the last 70 years. Most of these studies have been confined to one or two forms of alpha keratin. This high- and low-angle synchrotron fibre diffraction study extends the study to cover all available data for all known forms of hard α -keratin including hairs, fingernails, hooves, horn, and quills from mammals, marsupials, and a monotreme, and it confirms that the model proposed is universally acceptable for all mammals. A complete Bragg analysis of the meridional diffraction patterns, including multiple-timemore » exposures to verify any weak reflections, verified the existence of a superlattice consisting of two infinite lattices and three finite lattices. An analysis of the equatorial patterns establishes the radii of the oligomeric levels of dimers, tetramers, and intermediate filaments (IFs) together with the centre to centre distance for the IFs, thus confirming the proposed helices within helices molecular architecture for hard α -keratin. The results verify that the structure proposed by Feughelman and James meets the criteria for a valid α -keratin structure.« less

The Molecular Architecture for the Intermediate Filaments of Hard [alpha]-Keratin Based on the Superlattice Data Obtained from a Study ofMammals Using Synchrotron Fibre Diffraction

DOE Office of Scientific and Technical Information (OSTI.GOV)

James, Veronica

2014-09-24

High- and low-angle X-ray diffraction studies of hard {alpha}-keratin have been studied, and various models have been proposed over the last 70 years. Most of these studies have been confined to one or two forms of alpha keratin. This high- and low-angle synchrotron fibre diffraction study extends the study to cover all available data for all known forms of hard {alpha}-keratin including hairs, fingernails, hooves, horn, and quills from mammals, marsupials, and a monotreme, and it confirms that the model proposed is universally acceptable for all mammals. A complete Bragg analysis of the meridional diffraction patterns, including multiple-time exposures tomore » verify any weak reflections, verified the existence of a superlattice consisting of two infinite lattices and three finite lattices. An analysis of the equatorial patterns establishes the radii of the oligomeric levels of dimers, tetramers, and intermediate filaments (IFs) together with the centre to centre distance for the IFs, thus confirming the proposed helices within helices molecular architecture for hard {alpha}-keratin. The results verify that the structure proposed by Feughelman and James meets the criteria for a valid {alpha}-keratin structure.« less

Down-regulation of Pax6 is associated with abnormal differentiation of corneal epithelial cells in severe ocular surface diseases

PubMed Central

Li, W; Chen, Y-T; Hayashida, Y; Blanco, G; Kheirkah, A; He, H; Chen, S-Y; Liu, C-Y; Tseng, SCG

2010-01-01

Pax6 is the universal master control gene for eye morphogenesis. Other than retina and lens, Pax6 also expressed in the ocular surface epithelium from early gestation until the postnatal stage, in which little is known about the function of Pax6. In this study, corneal pannus tissues from patients with ocular surface diseases such as Stevens–Johnson syndrome (SJS), chemical burn, aniridia and recurrent pterygium were investigated. Our results showed that normal ocular surface epithelial cells expressed Pax6. However, corneal pannus epithelial cells from the above patients showed a decline or absence of Pax6 expression, accompanied by a decline or absence of K12 keratin but an increase of K10 keratin and filaggrin expression. Pannus basal epithelial cells maintained nuclear p63 expression and showed activated proliferation, evidenced by positive Ki67 and K16 keratin staining. On 3T3 fibroblast feeder layers, Pax6 immunostaining was negative in clones generated from epithelial cells harvested from corneal pannus from SJS or aniridia, but positive in those from the normal limbal epithelium; whereas western blots showed that some epithelial clones expanded from pannus retained Pax6 expression. Transient transfection of an adenoviral vector carrying EGFP–Pax6 transgenes into these Pax6− clones increased both Pax6 and K12 keratin expression. These results indicate that Pax6 helps to maintain the normal corneal epithelial phenotype postnatally, and that down-regulation of Pax6 is associated with abnormal epidermal differentiation in severe ocular surface diseases. Reintroduction of activation of the Pax6 gene might be useful in treating squamous metaplasia of the ocular surface epithelium. PMID:18027901

Interaction of Escherichia coli K1 and K5 with Acanthamoeba castellanii Trophozoites and Cysts

PubMed Central

Matin, Abdul

2011-01-01

The existence of symbiotic relationships between Acanthamoeba and a variety of bacteria is well-documented. However, the ability of Acanthamoeba interacting with host bacterial pathogens has gained particular attention. Here, to understand the interactions of Escherichia coli K1 and E. coli K5 strains with Acanthamoeba castellanii trophozoites and cysts, association assay, invasion assay, survival assay, and the measurement of bacterial numbers from cysts were performed, and nonpathogenic E. coli K12 was also applied. The association ratio of E. coli K1 with A. castellanii was 4.3 cfu per amoeba for 1 hr but E. coli K5 with A. castellanii was 1 cfu per amoeba for 1 hr. By invasion and survival assays, E. coli K5 was recovered less than E. coli K1 but still alive inside A. castellanii. E. coli K1 and K5 survived and multiplied intracellularly in A. castellanii. The survival assay was performed under a favourable condition for 22 hr and 43 hr with the encystment of A. castellanii. Under the favourable condition for the transformation of trophozoites into cysts, E. coli K5 multiplied significantly. Moreover, the pathogenic potential of E. coli K1 from A. castellanii cysts exhibited no changes as compared with E. coli K1 from A. castellanii trophozoites. E. coli K5 was multiplied in A. castellanii trophozoites and survived in A. castellanii cysts. Therefore, this study suggests that E. coli K5 can use A. castellanii as a reservoir host or a vector for the bacterial transmission. PMID:22355201

Interaction of Escherichia coli K1 and K5 with Acanthamoeba castellanii trophozoites and cysts.

PubMed

Matin, Abdul; Jung, Suk-Yul

2011-12-01

The existence of symbiotic relationships between Acanthamoeba and a variety of bacteria is well-documented. However, the ability of Acanthamoeba interacting with host bacterial pathogens has gained particular attention. Here, to understand the interactions of Escherichia coli K1 and E. coli K5 strains with Acanthamoeba castellanii trophozoites and cysts, association assay, invasion assay, survival assay, and the measurement of bacterial numbers from cysts were performed, and nonpathogenic E. coli K12 was also applied. The association ratio of E. coli K1 with A. castellanii was 4.3 cfu per amoeba for 1 hr but E. coli K5 with A. castellanii was 1 cfu per amoeba for 1 hr. By invasion and survival assays, E. coli K5 was recovered less than E. coli K1 but still alive inside A. castellanii. E. coli K1 and K5 survived and multiplied intracellularly in A. castellanii. The survival assay was performed under a favourable condition for 22 hr and 43 hr with the encystment of A. castellanii. Under the favourable condition for the transformation of trophozoites into cysts, E. coli K5 multiplied significantly. Moreover, the pathogenic potential of E. coli K1 from A. castellanii cysts exhibited no changes as compared with E. coli K1 from A. castellanii trophozoites. E. coli K5 was multiplied in A. castellanii trophozoites and survived in A. castellanii cysts. Therefore, this study suggests that E. coli K5 can use A. castellanii as a reservoir host or a vector for the bacterial transmission.

Selective biodegradation of keratin matrix in feather rachis reveals classic bioengineering

PubMed Central

Lingham-Soliar, Theagarten; Bonser, Richard H. C.; Wesley-Smith, James

2010-01-01

Flight necessitates that the feather rachis is extremely tough and light. Yet, the crucial filamentous hierarchy of the rachis is unknown—study hindered by the tight chemical bonding between the filaments and matrix. We used novel microbial biodegradation to delineate the fibres of the rachidial cortex in situ. It revealed the thickest keratin filaments known to date (factor >10), approximately 6 µm thick, extending predominantly axially but with a small outer circumferential component. Near-periodic thickened nodes of the fibres are staggered with those in adjacent fibres in two- and three-dimensional planes, creating a fibre–matrix texture with high attributes for crack stopping and resistance to transverse cutting. Close association of the fibre layer with the underlying ‘spongy’ medulloid pith indicates the potential for higher buckling loads and greater elastic recoil. Strikingly, the fibres are similar in dimensions and form to the free filaments of the feather vane and plumulaceous and embryonic down, the syncitial barbules, but, identified for the first time in 140+ years of study in a new location—as a major structural component of the rachis. Early in feather evolution, syncitial barbules were consolidated in a robust central rachis, definitively characterizing the avian lineage of keratin. PMID:20018788

A universal method for the mutational analysis of K-ras and p53 gene in non-small-cell lung cancer using formalin-fixed paraffin-embedded tissue.

PubMed

Sarkar, F H; Valdivieso, M; Borders, J; Yao, K L; Raval, M M; Madan, S K; Sreepathi, P; Shimoyama, R; Steiger, Z; Visscher, D W

1995-12-01

The p53 tumor suppressor gene has been found to be altered in almost all human solid tumors, whereas K-ras gene mutations have been observed in a limited number of human cancers (adenocarcinoma of colon, pancreas, and lung). Studies of mutational inactivation for both genes in the same patient's sample on non-small-cell lung cancer have been limited. In an effort to perform such an analysis, we developed and compared methods (for the mutational detection of p53 and K-ras gene) that represent a modified and universal protocol, in terms of DNA extraction, polymerase chain reaction (PCR) amplification, and nonradioisotopic PCR-single-strand conformation polymorphism (PCR-SSCP) analysis, which is readily applicable to either formalin-fixed, paraffin-embedded tissues or frozen tumor specimens. We applied this method to the evaluation of p53 (exons 5-8) and K-ras (codon 12 and 13) gene mutations in 55 cases of non-small-cell lung cancer. The mutational status in the p53 gene was evaluated by radioisotopic PCR-SSCP and compared with PCR-SSCP utilizing our standardized nonradioisotopic detection system using a single 6-microns tissue section. The mutational patterns observed by PCR-SSCP were subsequently confirmed by PCR-DNA sequencing. The mutational status in the K-ras gene was similarly evaluated by PCR-SSCP, and the specific mutation was confirmed by Southern slot-blot hybridization using 32P-labeled sequence-specific oligonucleotide probes for codons 12 and 13. Mutational changes in K-ras (codon 12) were found in 10 of 55 (18%) of non-small-cell lung cancers. Whereas adenocarcinoma showed K-ras mutation in 33% of the cases at codon 12, only one mutation was found at codon 13. As expected, squamous cell carcinoma samples (25 cases) did not show K-ras mutations. Mutations at exons 5-8 of the p53 gene were documented in 19 of 55 (34.5%) cases. Ten of the 19 mutations were single nucleotide point mutations, leading to amino acid substitution. Six showed insertional

Cysteic Acid in Dietary Keratin is Metabolized to Glutathione and Liver Taurine in a Rat Model of Human Digestion

PubMed Central

Wolber, Frances M.; McGrath, Michelle; Jackson, Felicity; Wylie, Kim; Broomfield, Anne

2016-01-01

Poultry feathers, consisting largely of keratin, are a low-value product of the poultry industry. The safety and digestibility of a dietary protein produced from keratin (KER) was compared to a cysteine-supplemented casein-based diet in a growing rat model for four weeks. KER proved to be an effective substitute for casein at 50% of the total dietary protein, with no changes in the rats’ food intake, weight gain, organ weight, bone mineral density, white blood cell counts, liver glutathione, or blood glutathione. Inclusion of KER in the diet reduced total protein digestibility from 94% to 86% but significantly increased total dietary cysteine uptake and subsequent liver taurine levels. The KER diet also significantly increased caecum weight and significantly decreased fat digestibility, resulting in a lower proportion of body fat, and induced a significant increase in blood haemoglobin. KER is therefore a safe and suitable protein substitute for casein, and the cysteic acid in keratin is metabolised to maintain normal liver and blood glutathione levels. PMID:26907334

K-shuff: A Novel Algorithm for Characterizing Structural and Compositional Diversity in Gene Libraries.

PubMed

Jangid, Kamlesh; Kao, Ming-Hung; Lahamge, Aishwarya; Williams, Mark A; Rathbun, Stephen L; Whitman, William B

2016-01-01

K-shuff is a new algorithm for comparing the similarity of gene sequence libraries, providing measures of the structural and compositional diversity as well as the significance of the differences between these measures. Inspired by Ripley's K-function for spatial point pattern analysis, the Intra K-function or IKF measures the structural diversity, including both the richness and overall similarity of the sequences, within a library. The Cross K-function or CKF measures the compositional diversity between gene libraries, reflecting both the number of OTUs shared as well as the overall similarity in OTUs. A Monte Carlo testing procedure then enables statistical evaluation of both the structural and compositional diversity between gene libraries. For 16S rRNA gene libraries from complex bacterial communities such as those found in seawater, salt marsh sediments, and soils, K-shuff yields reproducible estimates of structural and compositional diversity with libraries greater than 50 sequences. Similarly, for pyrosequencing libraries generated from a glacial retreat chronosequence and Illumina® libraries generated from US homes, K-shuff required >300 and 100 sequences per sample, respectively. Power analyses demonstrated that K-shuff is sensitive to small differences in Sanger or Illumina® libraries. This extra sensitivity of K-shuff enabled examination of compositional differences at much deeper taxonomic levels, such as within abundant OTUs. This is especially useful when comparing communities that are compositionally very similar but functionally different. K-shuff will therefore prove beneficial for conventional microbiome analysis as well as specific hypothesis testing.

K-shuff: A Novel Algorithm for Characterizing Structural and Compositional Diversity in Gene Libraries

PubMed Central

Jangid, Kamlesh; Kao, Ming-Hung; Lahamge, Aishwarya; Williams, Mark A.; Rathbun, Stephen L.; Whitman, William B.

2016-01-01

K-shuff is a new algorithm for comparing the similarity of gene sequence libraries, providing measures of the structural and compositional diversity as well as the significance of the differences between these measures. Inspired by Ripley’s K-function for spatial point pattern analysis, the Intra K-function or IKF measures the structural diversity, including both the richness and overall similarity of the sequences, within a library. The Cross K-function or CKF measures the compositional diversity between gene libraries, reflecting both the number of OTUs shared as well as the overall similarity in OTUs. A Monte Carlo testing procedure then enables statistical evaluation of both the structural and compositional diversity between gene libraries. For 16S rRNA gene libraries from complex bacterial communities such as those found in seawater, salt marsh sediments, and soils, K-shuff yields reproducible estimates of structural and compositional diversity with libraries greater than 50 sequences. Similarly, for pyrosequencing libraries generated from a glacial retreat chronosequence and Illumina® libraries generated from US homes, K-shuff required >300 and 100 sequences per sample, respectively. Power analyses demonstrated that K-shuff is sensitive to small differences in Sanger or Illumina® libraries. This extra sensitivity of K-shuff enabled examination of compositional differences at much deeper taxonomic levels, such as within abundant OTUs. This is especially useful when comparing communities that are compositionally very similar but functionally different. K-shuff will therefore prove beneficial for conventional microbiome analysis as well as specific hypothesis testing. PMID:27911946

Effects of tissue fixation conditions and protease pretreatment on immunohistochemical performance of a large series of new anti-keratin monoclonal antibodies: value in oncopathology.

PubMed

Bártková, J; Bártek, J; Lukás, Z; Vojtĕsek, B; Stasková, Z; Bursová, H; Pavlovská, R; Rejthar, A; Kovarík, J

1991-01-01

A comparative study with 21 recently raised monoclonal antibodies (3 of which are reported here for the first time) to human keratin polypeptides was performed on a wide range of paraffin-embedded tissues and tumors, aimed at the examination of effects of four different fixatives and protease pretreatment on the immunohistochemical detection of keratins. Our data demonstrated that: (a) formaldehyde-based fixatives modified by acidification and/or addition of methanol gave results superior to those achieved by routinely used formol saline; (b) relatively rare antibodies (4 out of 21) could be identified which gave reliable immunostaining patterns even on routine formalin-fixed material; (c) a proteolytic digestion step preceding the immunostaining was beneficial for the performance of the majority of antibodies in our panel. Additional options which could potentially lead to further improvement of keratin immunohistochemistry in paraffin embedded specimens are also suggested. This work provides the necessary basis for wider application of the anti-keratin antibodies of the C-series in both routine oncopathology and research-oriented retrospective studies.

The ESR signals in silk fibroin and wool keratin under both the effect of UV-irradiation and without any external effects and the formation of free radicals.

PubMed

Mamedov, Sh V; Aktas, B; Cantürk, M; Aksakal, B; Alekperov, V; Bülbül, F; Yilgin, R; Aslanov, R B

2002-08-01

ESR studies have been done on natural and UV-irradiated silk fibroins and wool keratins at the temperature range of -196 degrees C to 20 C. The intensities of ESR signals obtained from the irradiated samples at -196 C remarkably increase with respect to those of natural samples. While the signals mainly consist of triplet peaks at -196 C. a doublet arises around the room temperatures. For the first time, at room temperature without any external effect the complicated ESR spectra of fibrous proteins (wool keratin and silk fibroin) whose components are as follows have been observed: (1) (for white wool keratin) a central doublet with deltaHm = 1.1 mT and g = 2.0075; deltaHm = 5mT and g = 2.1911; (2) a wide peak with deltaHm approximately 66 mT and g approximately 2.1575; (3) the 'sulfur' peak given in the literature with deltaHm = 2.2 mT and g = 2.0218; (4) the signal with deltaHm = 0.6 mT and g = 2.0065, and for silk fibroin, (a) a very wide signal with deltaHm approximately 70 mT and g approximately 2.084; (b) a very sharp signal with deltaHm approximately 1.1 mT and g approximately 2.01; and (c) relatively narrower signal with deltaHm approximately 5 mT and g approximately 2.336. It has been shown by recombination kinetic method that 30-50% of the free radicals formed by UV-irradiation do not undergo recombination up to 220 degrees C and 15 degrees C for silk libroin and wool keratin, respectively, even they keep their concentration constant for long period of time (weeks, months, even longer). In this article, considering above-mentioned results, the mechanism of signals observed in natural wool keratin and silk fibroin without any external effects is examined. We can briefly explain the role of the subject of the article, by considering fibrous proteins and some applications of the reactions by free radical occurring in these proteins tinder the effects of different factors in medicine and biology and the important role of oxidation and the other kinds of

Evaluating the clinical and esthetic outcome of apically positioned flap technique in augmentation of keratinized gingiva around dental implants

PubMed Central

Reddy, Vineela Katam; Parthasarathy, Harinath; Lochana, Priya

2013-01-01

Purpose: Dental implants though a successful treatment modality there exists controversies regarding the relationship between the adequacy of the keratinized gingiva (KG) and peri-implant health. The presence of an adequate amount of peri-implant KG reduces gingival inflammation and hence soft-tissue augmentation should be frequently considered. Among the various periodontal plastic surgical procedures, the apically displaced flap increases the width of keratinized tissue with reduced patient morbidity. The current study aims at evaluating the esthetic improvement in KG around dental implants applying apically positioned flap (APF) technique. Materials and Methods: A total of 10 endosseous dental implants were placed in eight systemically healthy patients. APF surgery was performed at the implant site on the buccal aspect either at the time of implant placement (one stage surgical protocol) or during the implant recovery stage (two stage surgical protocols) for increasing the width of KG and reviewed until 12 weeks post-operatively. The width of KG was evaluated at baseline and at the end of 12 weeks after surgery. Paired t-test was performed to evaluate the changes in the width of KG at baseline and at 12 weeks post-operatively. In addition, soft-tissue esthetic outcome was assessed by using visual analog scale (VAS). Results: The mean width of KG at baseline was 1.47 mm and 12 weeks post-operatively was 5.42 mm. The gain in KG from baseline was 3.95 mm with the P value of 0.000, which was highly statistically significant. The assessment of esthetic outcome using VAS gave an average score of 7.1 indicating good esthetics. Conclusion: The technique of APF yielded a significant improvement in keratinized tissue, which is both functionally and esthetically acceptable. PMID:24124297

Molecular forms of HMGB1 and Keratin-18 as mechanistic biomarkers for mode of cell death and prognosis during clinical acetaminophen hepatotoxicity

PubMed Central

Antoine, Daniel J; Jenkins, Rosalind E; Dear, James W; Williams, Dominic P; McGill, Mitchell R; Sharpe, Matthew R; Craig, Darren G; Simpson, Kenneth J; Jaeschke, Hartmut; Park, B. Kevin

2014-01-01

Background & Aims Full length keratin-18 (FL-K18) and High Mobility Group Box-1 (HMGB1) represent circulating indicators of necrosis during acetaminophen (APAP) hepatotoxicity in vivo. In addition, the caspase-cleaved fragment of K18 (cK18) and hyper-acetylated HMGB1 represent serum indicators of apoptosis and immune cell activation respectively. The study aim was to assess their mechanistic utility to establish the balance between apoptosis, necrosis and immune cell activation throughout the time course of clinical APAP hepatotoxicity. Methods HMGB1 (total, acetylated) and K18 (apoptotic, necrotic) were identified and quantified by novel LC-MS/MS assays in APAP overdose patients (n=78). Results HMGB1 (total; 15.4±1.9ng/ml, p<0.01, acetylated; 5.4±2.6ng/ml, p<0.001), cK18 (5649.8±721.0U/l, p<0.01) and FL-K18 (54770.2±6717.0U/l, p<0.005) were elevated in the sera of APAP overdose patients with liver injury compared to overdose patients without liver injury and healthy volunteers. HMGB1 and FL-K18 correlated with alanine aminotransferase (ALT) activity (R2=0.60 and 0.58 respectively, p<0.0001) and prothrombin time (R2=0.62 and 0.71 respectively, p<0.0001). Increased total and acetylated HMGB1 and FL-K18 were associated with worse prognosis (King’s College Criteria) or patients that died/required liver transplant compared to spontaneous survivors (all p<0.05-0.001), a finding not reflected by ALT and supported by ROC analysis. Acetylated HMGB1 was a better predictor of outcome than the other markers of cell death. Conclusion K18 and HMGB1 represent blood-based tools to investigate the cell death balance clinical APAP hepatotoxicity. Activation of the immune response was seen later in the time course as shown by the distinct profile of acetylated HMGB1 and was associated with worse outcome. PMID:22266604

De Novo RNA Sequencing and Transcriptome Analysis of Monascus purpureus and Analysis of Key Genes Involved in Monacolin K Biosynthesis

PubMed Central

Zhang, Chan; Liang, Jian; Yang, Le; Sun, Baoguo; Wang, Chengtao

2017-01-01

Monascus purpureus is an important medicinal and edible microbial resource. To facilitate biological, biochemical, and molecular research on medicinal components of M. purpureus, we investigated the M. purpureus transcriptome by RNA sequencing (RNA-seq). An RNA-seq library was created using RNA extracted from a mixed sample of M. purpureus expressing different levels of monacolin K output. In total 29,713 unigenes were assembled from more than 60 million high-quality short reads. A BLAST search revealed hits for 21,331 unigenes in at least one of the protein or nucleotide databases used in this study. The 22,365 unigenes were categorized into 48 functional groups based on Gene Ontology classification. Owing to the economic and medicinal importance of M. purpureus, most studies on this organism have focused on the pharmacological activity of chemical components and the molecular function of genes involved in their biogenesis. In this study, we performed quantitative real-time PCR to detect the expression of genes related to monacolin K (mokA-mokI) at different phases (2, 5, 8, and 12 days) of M. purpureus M1 and M1-36. Our study found that mokF modulates monacolin K biogenesis in M. purpureus. Nine genes were suggested to be associated with the monacolin K biosynthesis. Studies on these genes could provide useful information on secondary metabolic processes in M. purpureus. These results indicate a detailed resource through genetic engineering of monacolin K biosynthesis in M. purpureus and related species. PMID:28114365

5C analysis of the Epidermal Differentiation Complex locus reveals distinct chromatin interaction networks between gene-rich and gene-poor TADs in skin epithelial cells

PubMed Central

Malashchuk, Igor; Lajoie, Brian R.; Mardaryev, Andrei N.; Gdula, Michal R.; Sharov, Andrey A.; Kohwi-Shigematsu, Terumi; Fessing, Michael Y.

2017-01-01

Mammalian genomes contain several dozens of large (>0.5 Mbp) lineage-specific gene loci harbouring functionally related genes. However, spatial chromatin folding, organization of the enhancer-promoter networks and their relevance to Topologically Associating Domains (TADs) in these loci remain poorly understood. TADs are principle units of the genome folding and represents the DNA regions within which DNA interacts more frequently and less frequently across the TAD boundary. Here, we used Chromatin Conformation Capture Carbon Copy (5C) technology to characterize spatial chromatin interaction network in the 3.1 Mb Epidermal Differentiation Complex (EDC) locus harbouring 61 functionally related genes that show lineage-specific activation during terminal keratinocyte differentiation in the epidermis. 5C data validated by 3D-FISH demonstrate that the EDC locus is organized into several TADs showing distinct lineage-specific chromatin interaction networks based on their transcription activity and the gene-rich or gene-poor status. Correlation of the 5C results with genome-wide studies for enhancer-specific histone modifications (H3K4me1 and H3K27ac) revealed that the majority of spatial chromatin interactions that involves the gene-rich TADs at the EDC locus in keratinocytes include both intra- and inter-TAD interaction networks, connecting gene promoters and enhancers. Compared to thymocytes in which the EDC locus is mostly transcriptionally inactive, these interactions were found to be keratinocyte-specific. In keratinocytes, the promoter-enhancer anchoring regions in the gene-rich transcriptionally active TADs are enriched for the binding of chromatin architectural proteins CTCF, Rad21 and chromatin remodeler Brg1. In contrast to gene-rich TADs, gene-poor TADs show preferential spatial contacts with each other, do not contain active enhancers and show decreased binding of CTCF, Rad21 and Brg1 in keratinocytes. Thus, spatial interactions between gene promoters and

Genomic organization of human fetal specific P-450IIIA7 (cytochrome P-450HFLa)-related gene(s) and interaction of transcriptional regulatory factor with its DNA element in the 5' flanking region.

PubMed

Itoh, S; Yanagimoto, T; Tagawa, S; Hashimoto, H; Kitamura, R; Nakajima, Y; Okochi, T; Fujimoto, S; Uchino, J; Kamataki, T

1992-03-24

P-450IIIA7 is a form of cytochrome P-450 which was isolated from human fetal livers and termed P-450HFLa. This form has been clarified to be expressed during fetal life specifically (Komori, M., Nishio, K., Kitada, M., Shiramatsu, K., Muroya, K., Soma, M., Nagashima, K. and Kamataki, T. (1990) Biochemistry 29, 4430-4433). In the present study, we isolated five independent clones which probably corresponded to the human P-450IIIA7 gene. These clones were completely sequenced, all exons, exon-intron junctions and the 5' flanking region from the cap site to-869. Although the sequences in the coding region were completely identical to P-450IIIA7, it is possible that genomic fragments sequenced in this study encode portions of other P-450IIIA7-related genes since we could not obtain a complete overlapping set of genomic clones. Within its 5' flanking sequence, the putative binding sites of several transcriptional regulatory factors existed. Among them, it was shown that a basic transcription element binding factor (BTEB) actually interacted with the 5' flanking region of this gene.

U.K. science budget. Gene jocks, data crunchers hit jackpot.

PubMed

Pickrell, J

2000-12-01

Unveiling its science spending plan for the next 3 years, the U.K. government last week announced major new investments in three key areas: tracking disease genes, leveraging the Internet for data analysis, and supporting emerging industries such as nanotechnology and bioengineering. Although these programs cut across a range of disciplines funded by the U.K.'s science councils, the government also bestowed a long-anticipated gift on astronomers: membership in the European Southern Observatory, which will give U.K. researchers access to the world's largest optical telescope.

Regulation of K-Cl cotransport: from function to genes.

PubMed

Adragna, N C; Di Fulvio, M; Lauf, P K

2004-10-01

cotransporter and the cytoskeleton appears to depend on the cellular origin and experimental conditions. Pathophysiologically, K-Cl COT is altered in sickle cell anemia and neuropathies, and it has also been proposed to play a role in blood pressure control. Four closely related human genes code for KCCs (KCC1-4). Although considerable information is accumulating on tissue distribution, function and pathologies associated with the different isoforms, little is known about the genetic regulation of the KCC genes in terms of transcriptional and post-transcriptional regulation. A few reports indicate that the NO/cGMP/PKG signaling pathway regulates KCC1 and KCC3 mRNA expression in VSMCs at the post-transcriptional level. However, the detailed mechanisms of post-transcriptional regulation of KCC genes and of regulation of KCC2 and KCC4 mRNA expression are unknown. The K-Cl COT field is expected to expand further over the next decades, as new isoforms and/or regulatory pathways are discovered and its implication in health and disease is revealed.

Physiological Responses and Gene Co-Expression Network of Mycorrhizal Roots under K+ Deprivation1[OPEN

PubMed Central

Roy, Sushmita

2017-01-01

Arbuscular mycorrhizal (AM) associations enhance the phosphorous and nitrogen nutrition of host plants, but little is known about their role in potassium (K+) nutrition. Medicago truncatula plants were cocultured with the AM fungus Rhizophagus irregularis under high and low K+ regimes for 6 weeks. We determined how K+ deprivation affects plant development and mineral acquisition and how these negative effects are tempered by the AM colonization. The transcriptional response of AM roots under K+ deficiency was analyzed by whole-genome RNA sequencing. K+ deprivation decreased root biomass and external K+ uptake and modulated oxidative stress gene expression in M. truncatula roots. AM colonization induced specific transcriptional responses to K+ deprivation that seem to temper these negative effects. A gene network analysis revealed putative key regulators of these responses. This study confirmed that AM associations provide some tolerance to K+ deprivation to host plants, revealed that AM symbiosis modulates the expression of specific root genes to cope with this nutrient stress, and identified putative regulators participating in these tolerance mechanisms. PMID:28159827

Mammalian Adaptation in the PB2 Gene of Avian H5N1 Influenza Virus

PubMed Central

Min, Ji-Young; Santos, Celia; Fitch, Adam; Twaddle, Alan; Toyoda, Yoshiko; DePasse, Jay V.

2013-01-01

The substitution of glutamic acid (E) for lysine (K) at position 627 of the PB2 protein of avian H5N1 viruses has been identified as a virulence and host range determinant for infection of mammals. Here, we report that the E-to-K host-adaptive mutation in the PB2 gene appeared from day 4 and 5 along the respiratory tracts of mice and was complete by day 6 postinoculation. This mutation correlated with efficient replication of the virus in mice. PMID:23864613

Shear Stress Induced Reorganization of the Keratin Intermediate Filament Network Requires Phosphorylation by Protein Kinase C ζ

PubMed Central

Sivaramakrishnan, Sivaraj; Schneider, Jaime L.; Sitikov, Albert; Goldman, Robert D.

2009-01-01

Keratin intermediate filaments (KIFs) form a fibrous polymer network that helps epithelial cells withstand external mechanical forces. Recently, we established a correlation between the structure of the KIF network and its local mechanical properties in alveolar epithelial cells. Shear stress applied across the cell surface resulted in the structural remodeling of KIF and a substantial increase in the elastic modulus of the network. This study examines the mechanosignaling that regulates the structural remodeling of the KIF network. We report that the shear stress–mediated remodeling of the KIF network is facilitated by a twofold increase in the dynamic exchange rate of KIF subunits, which is regulated in a PKC ζ and 14-3-3–dependent manner. PKC ζ phosphorylates K18pSer33, and this is required for the structural reorganization because the KIF network in A549 cells transfected with a dominant negative PKC ζ, or expressing the K18Ser33Ala mutation, is unchanged. Blocking the shear stress–mediated reorganization results in reduced cellular viability and increased apoptotic levels. These data suggest that shear stress mediates the phosphorylation of K18pSer33, which is required for the reorganization of the KIF network, resulting in changes in mechanical properties of the cell that help maintain the integrity of alveolar epithelial cells. PMID:19357195

Insulin Inhibits Nrf2 Gene Expression via Heterogeneous Nuclear Ribonucleoprotein F/K in Diabetic Mice

PubMed Central

Ghosh, Anindya; Abdo, Shaaban; Zhao, Shuiling; Wu, Chin-Han; Shi, Yixuan; Lo, Chao-Sheng; Chenier, Isabelle; Alquier, Thierry; Filep, Janos G.; Ingelfinger, Julie R.; Zhang, Shao-Ling

2017-01-01

Oxidative stress induces endogenous antioxidants via nuclear factor erythroid 2–related factor 2 (Nrf2), potentially preventing tissue injury. We investigated whether insulin affects renal Nrf2 expression in type 1 diabetes (T1D) and studied its underlying mechanism. Insulin normalized hyperglycemia, hypertension, oxidative stress, and renal injury; inhibited renal Nrf2 and angiotensinogen (Agt) gene expression; and upregulated heterogeneous nuclear ribonucleoprotein F and K (hnRNP F and hnRNP K) expression in Akita mice with T1D. In immortalized rat renal proximal tubular cells, insulin suppressed Nrf2 and Agt but stimulated hnRNP F and hnRNP K gene transcription in high glucose via p44/42 mitogen-activated protein kinase signaling. Transfection with small interfering RNAs of p44/42 MAPK, hnRNP F, or hnRNP K blocked insulin inhibition of Nrf2 gene transcription. Insulin curbed Nrf2 promoter activity via a specific DNA-responsive element that binds hnRNP F/K, and hnRNP F/K overexpression curtailed Nrf2 promoter activity. In hyperinsulinemic-euglycemic mice, renal Nrf2 and Agt expression was downregulated, whereas hnRNP F/K expression was upregulated. Thus, the beneficial actions of insulin in diabetic nephropathy appear to be mediated, in part, by suppressing renal Nrf2 and Agt gene transcription and preventing Nrf2 stimulation of Agt expression via hnRNP F/K. These findings identify hnRNP F/K and Nrf2 as potential therapeutic targets in diabetes. PMID:28324005

Phosphatidylinositol (4,5)-bisphosphate dynamically regulates the K2P background K+ channel TASK-2

PubMed Central

Niemeyer, María Isabel; Cid, L. Pablo; Paulais, Marc; Teulon, Jacques; Sepúlveda, Francisco V.

2017-01-01

Two-pore domain K2P K+ channels responsible for the background K+ conductance and the resting membrane potential, are also finely regulated by a variety of chemical, physical and physiological stimuli. Hormones and transmitters acting through Gq protein-coupled receptors (GqPCRs) modulate the activity of various K2P channels but the signalling involved has remained elusive, in particular whether dynamic regulation by membrane PI(4,5)P2, common among other classes of K+ channels, affects K2P channels is controversial. Here we show that K2P K+ channel TASK-2 requires PI(4,5)P2 for activity, a dependence that accounts for its run down in the absence of intracellular ATP and its full recovery by addition of exogenous PI(4,5)P2, its inhibition by low concentrations of polycation PI scavengers, and inhibition by PI(4,5)P2 depletion from the membrane. Comprehensive mutagenesis suggests that PI(4,5)P2 interaction with TASK-2 takes place at C-terminus where three basic aminoacids are identified as being part of a putative binding site. PMID:28358046

Phosphatidylinositol (4,5)-bisphosphate dynamically regulates the K2P background K+ channel TASK-2.

PubMed

Niemeyer, María Isabel; Cid, L Pablo; Paulais, Marc; Teulon, Jacques; Sepúlveda, Francisco V

2017-03-30

Two-pore domain K 2P K + channels responsible for the background K + conductance and the resting membrane potential, are also finely regulated by a variety of chemical, physical and physiological stimuli. Hormones and transmitters acting through Gq protein-coupled receptors (GqPCRs) modulate the activity of various K 2P channels but the signalling involved has remained elusive, in particular whether dynamic regulation by membrane PI(4,5)P 2 , common among other classes of K + channels, affects K 2P channels is controversial. Here we show that K 2P K + channel TASK-2 requires PI(4,5)P 2 for activity, a dependence that accounts for its run down in the absence of intracellular ATP and its full recovery by addition of exogenous PI(4,5)P 2 , its inhibition by low concentrations of polycation PI scavengers, and inhibition by PI(4,5)P 2 depletion from the membrane. Comprehensive mutagenesis suggests that PI(4,5)P 2 interaction with TASK-2 takes place at C-terminus where three basic aminoacids are identified as being part of a putative binding site.

The PDGFRα-laminin B1-keratin 19 cascade drives tumor progression at the invasive front of human hepatocellular carcinoma

PubMed Central

Govaere, O; Petz, M; Wouters, J; Vandewynckel, Y-P; Scott, E J; Topal, B; Nevens, F; Verslype, C; Anstee, Q M; Van Vlierberghe, H; Mikulits, W; Roskams, T

2017-01-01

Human hepatocellular carcinomas (HCCs) expressing the biliary/hepatic progenitor cell marker keratin 19 (K19) have been linked with a poor prognosis and exhibit an increase in platelet-derived growth factor receptor α (PDGFRα) and laminin beta 1 (LAMB1) expression. PDGFRα has been reported to induce de novo synthesis of LAMB1 protein in a Sjogren syndrome antigen B (La/SSB)-dependent manner in a murine metastasis model. However, the role of this cascade in human HCC remains unclear. This study focused on the functional role of the PDGFRα-La/SSB-LAMB1 pathway and its molecular link to K19 expression in human HCC. In surgical HCC specimens from a cohort of 136 patients, PDGFRα expression correlated with K19 expression, microvascular invasion and metastatic spread. In addition, PDGFRα expression in pre-operative needle biopsy specimens predicted poor overall survival during a 5-year follow-up period. Consecutive histological staining demonstrated that the signaling components of the PDGFRα-La/SSB-LAMB1 pathway were strongly expressed at the invasive front. K19-positive HCC cells displayed high levels of α2β1 integrin (ITG) receptor, both in vitro and in vivo. In vitro activation of PDGFRα signaling triggered the translocation of nuclear La/SSB into the cytoplasm, enhanced the protein synthesis of LAMB1 by activating its internal ribosome entry site, which in turn led to increased secretion of laminin-111. This effect was abrogated by the PDGFRα-specific inhibitor crenolanib. Importantly LAMB1 stimulated ITG-dependent focal adhesion kinase/Src proto-oncogene non-receptor tyrosine kinase signaling. It also promoted the ITG-specific downstream target Rho-associated coiled-coil containing protein kinase 2, induced K19 expression in an autocrine manner, invadopodia formation and cell invasion. Finally, we showed that the knockdown of LAMB1 or K19 in subcutaneous xenograft mouse models resulted in significant loss of cells invading the surrounding stromal tissue

5. OVERHEAD VIEW OF GENE CAMP LOOKING SOUTH. GENE PUMP ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

5. OVERHEAD VIEW OF GENE CAMP LOOKING SOUTH. GENE PUMP PLANT IS AT CENTER WITH ADMINISTRATIVE COMPLEX IN FOREGROUND AND RESIDENTIAL AREA BEYOND PLANT. - Gene Pump Plant, South of Gene Wash Reservoir, 2 miles west of Whitsett Pump Plant, Parker Dam, San Bernardino County, CA

Variation of Keratin 7 Expression and Other Phenotypic Characteristics of Independent Isolates of Cadmium Transformed Human Urothelial Cells (UROtsa)

PubMed Central

Somji, Seema; Zhou, Xu Dong; Mehus, Aaron; Sens, Mary Ann; Garrett, Scott H.; Lutz, Krista L.; Dunlevy, Jane R.; Zheng, Yun; Sens, Donald. A.

2009-01-01

This laboratory has shown that a human urothelial cell line (UROtsa) transformed by cadmium (Cd+2) produced subcutaneous tumor heterotransplants that resemble human transitional cell carcinoma (TCC). In the present study, additional Cd+2 transformed cell lines were isolated to determine if independent exposures of the cell line to Cd+2 would result in malignantly transformed cell lines possessing similar phenotypic properties. Seven independent isolates were isolated and assessed for their doubling times, morphology, ability to heterotransplant subcutaneously and in the peritoneal cavity of nude mice and for the expression keratin 7. The 7 cell lines all displayed an epithelial morphology with no evidence of squamous differentiation. Doubling times were variable among the isolates, being significantly reduced or similar to the parental cells. All 7 isolates were able to form subcutaneous tumor heterotransplants with a TCC morphology and all heterotransplants displayed areas of squamous differentiation of the transitional cells. The degree of squamous differentiation varied among the isolates. In contrast to subcutaneous tumor formation, only 1 isolate of the Cd+2 transformed cells (UTCd#1) was able to effectively colonize multiple sites within the peritoneal cavity. An analysis of keratin 7 expression showed no correlation with squamous differentiation for the subcutaneous heterotransplants generated from the 7 cell lines. Keratin 7 was expressed in 6 of the 7 cell lines and their subcutaneous tumor heterotransplants. Keratin 7 was not expressed in the cell line that was able to form tumors within the peritoneal cavity. These results show that individual isolates of Cd+2 transformed cells have both similarities and differences in their phenotype. PMID:19921857

DNA variation in the genes of the Na,K-adenosine triphosphatase and its relation with resting metabolic rate, respiratory quotient, and body fat.

PubMed

Dériaz, O; Dionne, F; Pérusse, L; Tremblay, A; Vohl, M C; Côté, G; Bouchard, C

1994-02-01

The aim of this study was to investigate in 261 subjects from 58 families the association between DNA variation at the genes coding for the Na,K-ATPase peptides and resting metabolic rate (RMR), respiratory quotient (RQ), and percent body fat (%FAT). Five restriction fragment length polymorphisms (RFLP) at three Na,K-ATPase genes were determined: one at the alpha 1 locus (BglII), and two at the beta locus (beta MspI and beta PvuII). Haplotypes were determined from the two variable sites of the alpha 2 gene (alpha 2 haplotypes) and the beta gene (beta haplotypes). There was a strong trend for %FAT to be related to the RFLP generated by BglII at the alpha 2 exons 21-22 in males (P = 0.06) and females (P = 0.05). RQ was (a) associated with the BglII RFLP at the alpha 2 exon 1 (P = 0.02) and with the alpha 2 8.0 kb/4.3 kb haplotype (P = 0.04) and (b) linked with the beta gene MspI marker (P = 0.04) and with the beta 5.3 kb/5.1 kb haplotype (P = 0.008) based on sib-pair analysis. The present study suggests that the genes encoding Na,K-ATPase may be associated or linked with RQ and perhaps with %FAT but not with RMR.

DNA variation in the genes of the Na,K-adenosine triphosphatase and its relation with resting metabolic rate, respiratory quotient, and body fat.

PubMed Central

Dériaz, O; Dionne, F; Pérusse, L; Tremblay, A; Vohl, M C; Côté, G; Bouchard, C

1994-01-01

The aim of this study was to investigate in 261 subjects from 58 families the association between DNA variation at the genes coding for the Na,K-ATPase peptides and resting metabolic rate (RMR), respiratory quotient (RQ), and percent body fat (%FAT). Five restriction fragment length polymorphisms (RFLP) at three Na,K-ATPase genes were determined: one at the alpha 1 locus (BglII), and two at the beta locus (beta MspI and beta PvuII). Haplotypes were determined from the two variable sites of the alpha 2 gene (alpha 2 haplotypes) and the beta gene (beta haplotypes). There was a strong trend for %FAT to be related to the RFLP generated by BglII at the alpha 2 exons 21-22 in males (P = 0.06) and females (P = 0.05). RQ was (a) associated with the BglII RFLP at the alpha 2 exon 1 (P = 0.02) and with the alpha 2 8.0 kb/4.3 kb haplotype (P = 0.04) and (b) linked with the beta gene MspI marker (P = 0.04) and with the beta 5.3 kb/5.1 kb haplotype (P = 0.008) based on sib-pair analysis. The present study suggests that the genes encoding Na,K-ATPase may be associated or linked with RQ and perhaps with %FAT but not with RMR. PMID:7509349

[Carcinogenesis and its mechanism of mutant-type[12Asp]K-ras4B gene].

PubMed

Gui, Li-ming; Wei, Li-hui; Zhang, Ying-mei; Wang, Jian-liu; Wang, Ying; Chen, Ying; Ma, Da-long

2002-01-01

Ras gene plays an important role in the extra- and intra-cellular signal transduction pathway. It mediates series cascade reactions, and eventually actives transcriptional factors in nucleus. It is unknown on the mechanism of carcinogenesis of Ras gene in endometrial carcinoma, though K-ras mutant is very common in endometrial atypical hyperplasia and carcinoma. On basis of discovering the mutation in 12th codon of K-ras in endometrial carcinoma cell line, HEC-1A, we explored the carcinogenesis and molecular mechanism of mutant-type [12Asp] K-ras4B gene. (1) Full-length [12Asp]K-ras4B cDNA was amplified with RT-PCR, then inserted into pcDI eukaryotic expressive vector. (2) Morphological change, growth kinetics in vitro and tumorigencity in nude mice in vivo after-before transfection were observed. (3) To test the cell growth kinetics by methyl thiazolium tetrazolium (MTT) and [3H]thymidine incorporation method. (1) The authors have successfully constructed eukaryotic expression plasmid pcDI-[12Asp] K-ras4B; (2) To confirm that [12Asp] K-ras4B mutant can trigger the neoplastic transformation of NIH3T3 cells by test in vitro and in vivo. (3) After pMCV-RasN17 plasmid, a Ras mutant were transfected into pcDI-[12Asp] K-ras4B cells, the growth of this cell were restrained significantly in comparison with control group. (4) These findings indicate the expression of RafS621A resulted in remarkable inhibition in proliferation of pcDI-[12Asp]K-ras4B cell (P < 0.05). However, RafCAAX mutant can enhance pcDI-[12Asp]K-ras4B cell growth (P < 0.05). (1) [12Asp]K-ras4B gene alone is able to cause neoplastic transformation in NIH3T3 cells in vitro and in vivo. (2) [12Asp]K-ras4B-induced NIH3T3 cells neoplastic transformation required Raf signaling pathway.

[K198N polymorphism in the EDN1 gene in patients with pulmonary arterial hypertension].

PubMed

Pousada, Guillermo; Baloira, Adolfo; Vilariño, Carlos; Valverde, Diana

2015-04-20

In pulmonary arterial hypertension (PAH) an association with a polymorphism in the endothelin gene (EDN1) has been described. The main objective of this study was to analyze the polymorphism K198N in the gene EDN1 in patients with PAH, correlating the results with clinical and hemodynamic parameters. We compared 41 patients diagnosed with idiopathic and associated PAH of group i with 50 healthy controls. Polymerase chain reaction and direct sequencing were used to analyze the polymorphism K198N. We compared the genotype distribution and searched for a correlation with clinical, hemodynamic and therapeutic response. Genotype GG was present in 42% of patients in this study and 65% of controls. The GT+TT genotypes appeared in 58% of patients and in 35% of controls. Statistically significant differences between patients and controls (P=.032) were detected, with a relative risk in carriers of having the T allele of 2.51 (95% CI 1.07 to 5.86). The analysis by PolyPhen software defined K198N change as pathogenic. No significant differences in the response to treatment at medium term were found. The genotype analysis of the EDN1 gene polymorphism shows statistically significant differences in patients with PAH compared to healthy individuals. Individuals carrying at least one T allele exhibit a higher relative significant risk to develop HAP. Copyright © 2013 Elsevier España, S.L.U. All rights reserved.

Genes In Space-5

NASA Image and Video Library

2018-04-13

iss055e020319 (April 13, 2018) --- Flight Engineer Ricky Arnold processes of samples inside the Miniature Polymerase Chain Reaction (miniPCR) for the Genes In Space-5 experiment. The research gathered from Genes in Space-5 may be valuable in the development of procedures to maintain astronaut health and prevent an increased risk of cancer on deep space missions. The investigation also provides a deeper understanding of the human immune system, while giving student researchers a direct connection to the space program and offering hands-on educational experiences on Earth and promoting involvement in STEM fields.

Genetic diversity of K-antigen gene clusters of Escherichia coli and their molecular typing using a suspension array.

PubMed

Yang, Shuang; Xi, Daoyi; Jing, Fuyi; Kong, Deju; Wu, Junli; Feng, Lu; Cao, Boyang; Wang, Lei

2018-04-01

Capsular polysaccharides (CPSs), or K-antigens, are the major surface antigens of Escherichia coli. More than 80 serologically unique K-antigens are classified into 4 groups (Groups 1-4) of capsules. Groups 1 and 4 contain the Wzy-dependent polymerization pathway and the gene clusters are in the order galF to gnd; Groups 2 and 3 contain the ABC-transporter-dependent pathway and the gene clusters consist of 3 regions, regions 1, 2 and 3. Little is known about the variations among the gene clusters. In this study, 9 serotypes of K-antigen gene clusters (K2ab, K11, K20, K24, K38, K84, K92, K96, and K102) were sequenced and correlated with their CPS chemical structures. On the basis of sequence data, a K-antigen-specific suspension array that detects 10 distinct CPSs, including the above 9 CPSs plus K30, was developed. This is the first report to catalog the genetic features of E. coli K-antigen variations and to develop a suspension array for their molecular typing. The method has a number of advantages over traditional bacteriophage and serum agglutination methods and lays the foundation for straightforward identification and detection of additional K-antigens in the future.

Quantitative fucK gene polymerase chain reaction on sputum and nasopharyngeal secretions to detect Haemophilus influenzae pneumonia.

PubMed

Abdeldaim, Guma M K; Strålin, Kristoffer; Olcén, Per; Blomberg, Jonas; Mölling, Paula; Herrmann, Björn

2013-06-01

A quantitative polymerase chain reaction (PCR) for the fucK gene was developed for specific detection of Haemophilus influenzae. The method was tested on sputum and nasopharyngeal aspirate (NPA) from 78 patients with community-acquired pneumonia (CAP). With a reference standard of sputum culture and/or serology against the patient's own nasopharyngeal isolate, H. influenzae etiology was detected in 20 patients. Compared with the reference standard, fucK PCR (using the detection limit 10(5) DNA copies/mL) on sputum and NPA showed a sensitivity of 95.0% (19/20) in both cases, and specificities of 87.9% (51/58) and 89.5% (52/58), respectively. In a receiver operating characteristic curve analysis, sputum fucK PCR was found to be significantly superior to sputum P6 PCR for detection of H. influenzae CAP. NPA fucK PCR was positive in 3 of 54 adult controls without respiratory symptoms. In conclusion, quantitative fucK real-time PCR provides a sensitive and specific identification of H. influenzae in respiratory secretions. Copyright © 2013 Elsevier Inc. All rights reserved.

Social isolation stress induces ATF-7 phosphorylation and impairs silencing of the 5-HT 5B receptor gene

PubMed Central

Maekawa, Toshio; Kim, Seungjoon; Nakai, Daisuke; Makino, Chieko; Takagi, Tsuyoshi; Ogura, Hiroo; Yamada, Kazuyuki; Chatton, Bruno; Ishii, Shunsuke

2010-01-01

Many symptoms induced by isolation rearing of rodents may be relevant to neuropsychiatric disorders, including depression. However, identities of transcription factors that regulate gene expression in response to chronic social isolation stress remain elusive. The transcription factor ATF-7 is structurally related to ATF-2, which is activated by various stresses, including inflammatory cytokines. Here, we report that Atf-7-deficient mice exhibit abnormal behaviours and increased 5-HT receptor 5B (Htr5b) mRNA levels in the dorsal raphe nuclei. ATF-7 silences the transcription of Htr5B by directly binding to its 5′-regulatory region, and mediates histone H3-K9 trimethylation via interaction with the ESET histone methyltransferase. Isolation-reared wild-type (WT) mice exhibit abnormal behaviours that resemble those of Atf-7-deficient mice. Upon social isolation stress, ATF-7 in the dorsal raphe nucleus is phosphorylated via p38 and is released from the Htr5b promoter, leading to the upregulation of Htr5b. Thus, ATF-7 may have a critical role in gene expression induced by social isolation stress. PMID:19893493

RNA Sequencing-Based Genome Reannotation of the Dermatophyte Arthroderma benhamiae and Characterization of Its Secretome and Whole Gene Expression Profile during Infection

PubMed Central

De Coi, Niccolò; Feuermann, Marc; Schmid-Siegert, Emanuel; Băguţ, Elena-Tatiana; Mignon, Bernard; Waridel, Patrice; Peter, Corinne; Pradervand, Sylvain

2016-01-01

ABSTRACT Dermatophytes are the most common agents of superficial mycoses in humans and animals. The aim of the present investigation was to systematically identify the extracellular, possibly secreted, proteins that are putative virulence factors and antigenic molecules of dermatophytes. A complete gene expression profile of Arthroderma benhamiae was obtained during infection of its natural host (guinea pig) using RNA sequencing (RNA-seq) technology. This profile was completed with those of the fungus cultivated in vitro in two media containing either keratin or soy meal protein as the sole source of nitrogen and in Sabouraud medium. More than 60% of transcripts deduced from RNA-seq data differ from those previously deposited for A. benhamiae. Using these RNA-seq data along with an automatic gene annotation procedure, followed by manual curation, we produced a new annotation of the A. benhamiae genome. This annotation comprised 7,405 coding sequences (CDSs), among which only 2,662 were identical to the currently available annotation, 383 were newly identified, and 15 secreted proteins were manually corrected. The expression profile of genes encoding proteins with a signal peptide in infected guinea pigs was found to be very different from that during in vitro growth when using keratin as the substrate. Especially, the sets of the 12 most highly expressed genes encoding proteases with a signal sequence had only the putative vacuolar aspartic protease gene PEP2 in common, during infection and in keratin medium. The most upregulated gene encoding a secreted protease during infection was that encoding subtilisin SUB6, which is a known major allergen in the related dermatophyte Trichophyton rubrum. IMPORTANCE Dermatophytoses (ringworm, jock itch, athlete’s foot, and nail infections) are the most common fungal infections, but their virulence mechanisms are poorly understood. Combining transcriptomic data obtained from growth under various culture conditions with data

RNA Sequencing-Based Genome Reannotation of the Dermatophyte Arthroderma benhamiae and Characterization of Its Secretome and Whole Gene Expression Profile during Infection.

PubMed

Tran, Van Du T; De Coi, Niccolò; Feuermann, Marc; Schmid-Siegert, Emanuel; Băguţ, Elena-Tatiana; Mignon, Bernard; Waridel, Patrice; Peter, Corinne; Pradervand, Sylvain; Pagni, Marco; Monod, Michel

2016-01-01

Dermatophytes are the most common agents of superficial mycoses in humans and animals. The aim of the present investigation was to systematically identify the extracellular, possibly secreted, proteins that are putative virulence factors and antigenic molecules of dermatophytes. A complete gene expression profile of Arthroderma benhamiae was obtained during infection of its natural host (guinea pig) using RNA sequencing (RNA-seq) technology. This profile was completed with those of the fungus cultivated in vitro in two media containing either keratin or soy meal protein as the sole source of nitrogen and in Sabouraud medium. More than 60% of transcripts deduced from RNA-seq data differ from those previously deposited for A. benhamiae . Using these RNA-seq data along with an automatic gene annotation procedure, followed by manual curation, we produced a new annotation of the A. benhamiae genome. This annotation comprised 7,405 coding sequences (CDSs), among which only 2,662 were identical to the currently available annotation, 383 were newly identified, and 15 secreted proteins were manually corrected. The expression profile of genes encoding proteins with a signal peptide in infected guinea pigs was found to be very different from that during in vitro growth when using keratin as the substrate. Especially, the sets of the 12 most highly expressed genes encoding proteases with a signal sequence had only the putative vacuolar aspartic protease gene PEP2 in common, during infection and in keratin medium. The most upregulated gene encoding a secreted protease during infection was that encoding subtilisin SUB6, which is a known major allergen in the related dermatophyte Trichophyton rubrum . IMPORTANCE Dermatophytoses (ringworm, jock itch, athlete's foot, and nail infections) are the most common fungal infections, but their virulence mechanisms are poorly understood. Combining transcriptomic data obtained from growth under various culture conditions with data obtained

On the so-called membrane coating granules in keratinized lichen planus lesions of the buccal mucosa.

PubMed

El-Labban, N G; Wood, R D

1982-11-01

Serial sections of the so-called membrane-coating granules have been examined in keratinized oral epithelium of lichen planus lesions. As with 'granules' apparent in non-keratinized epithelium, it is found they do not represent specialized intra-cytoplasmic organelles, but are the result of sectioning at different areas, levels and planes through the plasma membrane of interdigitating cell processes. Such 'granules' appear mostly in the superficial, but not deep, part of the cytoplasm of the upper prickle cells. This is considered to be due to topographic differences between the upper and under surfaces of these cells and the presence of narrower intercellular spaces than those between deeper epithelial cells. Such arrangement often results in cell processes in sections appearing free in the superficial part of the cell below. The appearance of 'granules' arises when the plane of section is not at right angles to the two plasma membranes surrounding these processes.

Identification of the Genes Involved in the Biofilm-like Structures on Actinomyces oris K20, a Clinical Isolate from an Apical Lesion

DTIC Science & Technology

2013-01-01

protein conserved in Actinobacteria M206‡ AoriK_010100005764 ZP_08125978 Hypothetical protein AoriK_010100005769 ZP_08125979 TransRDD family protein M155...conserved in Actinobacteria . In mutant 4 (designated strain M206), we found that EZ-Tn5 was integrated into an intergenic region between 2 genes in divergent

Therapeutic Application of Phage Capsule Depolymerases against K1, K5, and K30 Capsulated E. coli in Mice.

PubMed

Lin, Han; Paff, Matthew L; Molineux, Ian J; Bull, James J

2017-01-01

Capsule depolymerase enzymes offer a promising class of new antibiotics. In vivo studies are encouraging but it is unclear how well this type of phage product will generalize in therapeutics, or whether different depolymerases against the same capsule function similarly. Here, in vivo efficacy was tested using cloned bacteriophage depolymerases against Escherichia coli strains with three different capsule types: K1, K5, and K30. When treating infections with the cognate capsule type in a mouse thigh model, the previously studied K1E depolymerase rescued poorly, whereas K1F, K1H, K5, and K30 depolymerases rescued well. K30 gp41 was identified as the catalytically active protein. In contrast to the in vivo studies, K1E enzyme actively degraded K1 capsule polysaccharide in vitro and sensitized K1 bacteria to serum killing. The only in vitro correlate of poor K1E performance in vivo was that the purified enzyme did not form the expected trimer. K1E appeared as an 18-mer which might limit its in vivo distribution. Overall, depolymerases were easily identified, cloned from phage genomes, and as purified proteins they proved generally effective.

Shikonin, vitamin K3 and vitamin K5 inhibit multiple glycolytic enzymes in MCF-7 cells.

PubMed

Chen, Jing; Hu, Xun; Cui, Jingjie

2018-05-01

Glycolysis is the most important source of energy for the production of anabolic building blocks in cancer cells. Therefore, glycolytic enzymes are regarded as potential targets for cancer treatment. Previously, naphthaquinones, including shikonin, vitamin K 3 and vitamin K 5 , have been proven to decrease the rate of glycolysis in cancer cells, which is partly due to suppressed pyruvate kinase activity. In the present study, enzymatic assays were performed using MCF-7 cell lysate in order to screen the profile of glycolytic enzymes in cancer cells inhibited by shikonin, vitamin K 3 and vitamin K 5 , in addition to pyruvate kinase. Results revealed that hexokinase, phosphofructokinase-1, fructose bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase produced in the process of glycolysis were inhibited by shikonin, vitamin K 3 and vitamin K 5 . The results indicated that shikonin, vitamin K 3 and vitamin K 5 are chemical inhibitors of glycolytic enzymes in cancer cells and have potential uses in translational medical applications.

Shikonin, vitamin K3 and vitamin K5 inhibit multiple glycolytic enzymes in MCF-7 cells

PubMed Central

Chen, Jing; Hu, Xun; Cui, Jingjie

2018-01-01

Glycolysis is the most important source of energy for the production of anabolic building blocks in cancer cells. Therefore, glycolytic enzymes are regarded as potential targets for cancer treatment. Previously, naphthaquinones, including shikonin, vitamin K3 and vitamin K5, have been proven to decrease the rate of glycolysis in cancer cells, which is partly due to suppressed pyruvate kinase activity. In the present study, enzymatic assays were performed using MCF-7 cell lysate in order to screen the profile of glycolytic enzymes in cancer cells inhibited by shikonin, vitamin K3 and vitamin K5, in addition to pyruvate kinase. Results revealed that hexokinase, phosphofructokinase-1, fructose bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase produced in the process of glycolysis were inhibited by shikonin, vitamin K3 and vitamin K5. The results indicated that shikonin, vitamin K3 and vitamin K5 are chemical inhibitors of glycolytic enzymes in cancer cells and have potential uses in translational medical applications. PMID:29725454

Identification and classification of genes regulated by phosphatidylinositol 3-kinase- and TRKB-mediated signalling pathways during neuronal differentiation in two subtypes of the human neuroblastoma cell line SH-SY5Y.

PubMed

Nishida, Yuichiro; Adati, Naoki; Ozawa, Ritsuko; Maeda, Aasami; Sakaki, Yoshiyuki; Takeda, Tadayuki

2008-10-28

SH-SY5Y cells exhibit a neuronal phenotype when treated with all-trans retinoic acid (RA), but the molecular mechanism of activation in the signalling pathway mediated by phosphatidylinositol 3-kinase (PI3K) is unclear. To investigate this mechanism, we compared the gene expression profiles in SK-N-SH cells and two subtypes of SH-SY5Y cells (SH-SY5Y-A and SH-SY5Y-E), each of which show a different phenotype during RA-mediated differentiation. SH-SY5Y-A cells differentiated in the presence of RA, whereas RA-treated SH-SY5Y-E cells required additional treatment with brain-derived neurotrophic factor (BDNF) for full differentiation. After exposing cells to a PI3K inhibitor, LY294002, we identified 386 genes and categorised these genes into two clusters dependent on the PI3K signalling pathway during RA-mediated differentiation in SH-SY5Y-A cells. Transcriptional regulation of the gene cluster, including 158 neural genes, was greatly reduced in SK-N-SH cells and partially impaired in SH-SY5Y-E cells, which is consistent with a defect in the neuronal phenotype of these cells. Additional stimulation with BDNF induced a set of neural genes that were down-regulated in RA-treated SH-SY5Y-E cells but were abundant in differentiated SH-SY5Y-A cells. We identified gene clusters controlled by PI3K- and TRKB-mediated signalling pathways during the differentiation of two subtypes of SH-SY5Y cells. The TRKB-mediated bypass pathway compensates for impaired neural function generated by defects in several signalling pathways, including PI3K in SH-SY5Y-E cells. Our expression profiling data will be useful for further elucidation of the signal transduction-transcriptional network involving PI3K or TRKB.

Truncated ALK derived from chromosomal translocation t(2;5)(p23;q35) binds to the SH3 domain of p85-PI3K.

PubMed

Polgar, Doris; Leisser, Christina; Maier, Susanne; Strasser, Stephan; Rüger, Beate; Dettke, Markus; Khorchide, Maya; Simonitsch, Ingrid; Cerni, Christa; Krupitza, Georg

2005-02-15

The chromosomal translocation t(2;5)(p23;q35) is associated with "Anaplastic large cell lymphomas" (ALCL), a Non Hodgkin Lymphoma occurring in childhood. The fusion of the tyrosine kinase gene-ALK (anaplastic lymphoma kinase) on chromosome 2p23 to the NPM (nucleophosmin/B23) gene on chromosome 5q35 results in a 80 kDa chimeric protein, which activates the "survival" kinase PI3K. However, the binding mechanism between truncated ALK and PI3K is poorly understood. Therefore, we attempted to elucidate the molecular interaction between ALK and the regulatory p85 subunit of PI3K. Here we provide evidence that the truncated ALK homodimer binds to the SH3 domain of p85. This finding may be useful for the development of a new target-specific intervention.

Gene Cluster Responsible for Secretion of and Immunity to Multiple Bacteriocins, the NKR-5-3 Enterocins

PubMed Central

Ishibashi, Naoki; Himeno, Kohei; Masuda, Yoshimitsu; Perez, Rodney Honrada; Iwatani, Shun; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2014-01-01

Enterococcus faecium NKR-5-3, isolated from Thai fermented fish, is characterized by the unique ability to produce five bacteriocins, namely, enterocins NKR-5-3A, -B, -C, -D, and -Z (Ent53A, Ent53B, Ent53C, Ent53D, and Ent53Z). Genetic analysis with a genome library revealed that the bacteriocin structural genes (enkA [ent53A], enkC [ent53C], enkD [ent53D], and enkZ [ent53Z]) that encode these peptides (except for Ent53B) are located in close proximity to each other. This NKR-5-3ACDZ (Ent53ACDZ) enterocin gene cluster (approximately 13 kb long) includes certain bacteriocin biosynthetic genes such as an ABC transporter gene (enkT), two immunity genes (enkIaz and enkIc), a response regulator (enkR), and a histidine protein kinase (enkK). Heterologous-expression studies of enkT and ΔenkT mutant strains showed that enkT is responsible for the secretion of Ent53A, Ent53C, Ent53D, and Ent53Z, suggesting that EnkT is a wide-range ABC transporter that contributes to the effective production of these bacteriocins. In addition, EnkIaz and EnkIc were found to confer self-immunity to the respective bacteriocins. Furthermore, bacteriocin induction assays performed with the ΔenkRK mutant strain showed that EnkR and EnkK are regulatory proteins responsible for bacteriocin production and that, together with Ent53D, they constitute a three-component regulatory system. Thus, the Ent53ACDZ gene cluster is essential for the biosynthesis and regulation of NKR-5-3 enterocins, and this is, to our knowledge, the first report that demonstrates the secretion of multiple bacteriocins by an ABC transporter. PMID:25149515

The histone demethylase Jarid1b ensures faithful mouse development by protecting developmental genes from aberrant H3K4me3.

PubMed

Albert, Mareike; Schmitz, Sandra U; Kooistra, Susanne M; Malatesta, Martina; Morales Torres, Cristina; Rekling, Jens C; Johansen, Jens V; Abarrategui, Iratxe; Helin, Kristian

2013-04-01

Embryonic development is tightly regulated by transcription factors and chromatin-associated proteins. H3K4me3 is associated with active transcription and H3K27me3 with gene repression, while the combination of both keeps genes required for development in a plastic state. Here we show that deletion of the H3K4me2/3 histone demethylase Jarid1b (Kdm5b/Plu1) results in major neonatal lethality due to respiratory failure. Jarid1b knockout embryos have several neural defects including disorganized cranial nerves, defects in eye development, and increased incidences of exencephaly. Moreover, in line with an overlap of Jarid1b and Polycomb target genes, Jarid1b knockout embryos display homeotic skeletal transformations typical for Polycomb mutants, supporting a functional interplay between Polycomb proteins and Jarid1b. To understand how Jarid1b regulates mouse development, we performed a genome-wide analysis of histone modifications, which demonstrated that normally inactive genes encoding developmental regulators acquire aberrant H3K4me3 during early embryogenesis in Jarid1b knockout embryos. H3K4me3 accumulates as embryonic development proceeds, leading to increased expression of neural master regulators like Pax6 and Otx2 in Jarid1b knockout brains. Taken together, these results suggest that Jarid1b regulates mouse development by protecting developmental genes from inappropriate acquisition of active histone modifications.

The Histone Demethylase Jarid1b Ensures Faithful Mouse Development by Protecting Developmental Genes from Aberrant H3K4me3

PubMed Central

Kooistra, Susanne M.; Malatesta, Martina; Morales Torres, Cristina; Rekling, Jens C.; Johansen, Jens V.; Abarrategui, Iratxe; Helin, Kristian

2013-01-01

Embryonic development is tightly regulated by transcription factors and chromatin-associated proteins. H3K4me3 is associated with active transcription and H3K27me3 with gene repression, while the combination of both keeps genes required for development in a plastic state. Here we show that deletion of the H3K4me2/3 histone demethylase Jarid1b (Kdm5b/Plu1) results in major neonatal lethality due to respiratory failure. Jarid1b knockout embryos have several neural defects including disorganized cranial nerves, defects in eye development, and increased incidences of exencephaly. Moreover, in line with an overlap of Jarid1b and Polycomb target genes, Jarid1b knockout embryos display homeotic skeletal transformations typical for Polycomb mutants, supporting a functional interplay between Polycomb proteins and Jarid1b. To understand how Jarid1b regulates mouse development, we performed a genome-wide analysis of histone modifications, which demonstrated that normally inactive genes encoding developmental regulators acquire aberrant H3K4me3 during early embryogenesis in Jarid1b knockout embryos. H3K4me3 accumulates as embryonic development proceeds, leading to increased expression of neural master regulators like Pax6 and Otx2 in Jarid1b knockout brains. Taken together, these results suggest that Jarid1b regulates mouse development by protecting developmental genes from inappropriate acquisition of active histone modifications. PMID:23637629

Identification and Characterization of 30 K Protein Genes Found in Bombyx mori (Lepidoptera: Bombycidae) Transcriptome

PubMed Central

Shi, Xiao-Feng; Li, Yi-Nü; Yi, Yong-Zhu; Xiao, Xing-Guo; Zhang, Zhi-Fang

2015-01-01

The 30 K proteins, the major group of hemolymph proteins in the silkworm, Bombyx mori (Lepidoptera: Bombycidae), are structurally related with molecular masses of ∼30 kDa and are involved in various physiological processes, e.g., energy storage, embryonic development, and immune responses. For this report, known 30 K protein gene sequences were used as Blastn queries against sequences in the B. mori transcriptome (SilkTransDB). Twenty-nine cDNAs (Bm30K-1–29) were retrieved, including four being previously unidentified in the Lipoprotein_11 family. The genomic structures of the 29 genes were analyzed and they were mapped to their corresponding chromosomes. Furthermore, phylogenetic analysis revealed that the 29 genes encode three types of 30 K proteins. The members increased in each type is mainly a result of gene duplication with the appearance of each type preceding the differentiation of each species included in the tree. Real-Time Quantitative Polymerase Chain Reaction (Q-PCR) confirmed that the genes could be expressed, and that the three types have different temporal expression patterns. Proteins from the hemolymph was separated by SDS-PAGE, and those with molecular mass of ∼30 kDa were isolated and identified by mass spectrometry sequencing in combination with searches of various databases containing B. mori 30K protein sequences. Of the 34 proteins identified, 13 are members of the 30 K protein family, with one that had not been found in the SilkTransDB, although it had been found in the B. mori genome. Taken together, our results indicate that the 30 K protein family contains many members with various functions. Other methods will be required to find more members of the family. PMID:26078299

Demonstration of intermediate cells during human prostate epithelial differentiation in situ and in vitro using triple-staining confocal scanning microscopy.

PubMed

van Leenders, G; Dijkman, H; Hulsbergen-van de Kaa, C; Ruiter, D; Schalken, J

2000-08-01

In human prostate epithelium, morphologically basal and luminal cells can be discriminated. The basal cell layer that putatively contains progenitor cells of the secretory epithelium is characterized by the expression of keratins (K) 5 and 14. Luminal cells represent the secretory compartment of the epithelium and express K8 and 18. We developed a technique for the simultaneous analysis of K5, 14, and 18 to identify intermediate cell stages in the prostate epithelium and to study the dynamic aspects of its differentiation in vitro. Nonmalignant prostate tissue and primary epithelial cultures were immunohistochemically characterized using triple staining with antibodies for K5, K14, and K18. Antibodies for K18 and K5 were conjugated directly with fluorochromes Alexa 488 and 546. K14 was visualized indirectly with streptavidin-Cy5. Keratin expression was analyzed by confocal scanning microscopy. The occurrence of exocrine and neuroendocrine differentiation in culture was determined via antibodies to prostate-specific antigen (PSA), chromogranin A, and serotonin. We found that basal cells expressed either K5(++)/14(++)/18+ or K5(++)/18+. The majority of luminal cells expressed K18(++), but colocalization of K5+/18(++) were recognized. Epithelial monolayer cultures predominantly revealed the basal cell phenotype K5(++)/14(++)/18+, whereas intermediate subpopulations expressing K5+/14+/18(++) and K5+/18(++) were also identified. On confluence, differentiation was induced as multicellular gland-like buds, and extensions became evident on top of the monolayer. These structures were composed of K18(++)- and K5+/18(+)-positive cell clusters surrounded by phenotypically basal cells. Few multicellular structures and cells in the monolayer showed exocrine differentiation (PSA+), but expression of chromogranin A and serotonin was absent. We conclude that simultaneous evaluation of keratin expression is useful for analyzing epithelial differentiation in the prostate. During this

Prenatal retinoic acid upregulates pulmonary gene expression of PI3K and AKT in nitrofen-induced pulmonary hypoplasia.

PubMed

Doi, Takashi; Sugimoto, Kaoru; Ruttenstock, Elke; Dingemann, Jens; Puri, Prem

2010-10-01

The precise mechanism of pulmonary hypoplasia associated with congenital diaphragmatic hernia (CDH) still remains unclear. Recently, prenatal treatment with retinoic acid (RA) has been reported to stimulate alveologenesis in hypoplastic lungs in the nitrofen model of CDH. The serine/threonine protein kinase B (AKT) plays a key role in lung morphogenesis through epithelial-mesenchymal interaction in phosphatidylinositide 3-kinase (PI3K)-dependent manner. It has been reported that the lung morphogenesis in explants in mice is interfered by inhibitors of PI3K-AKT signaling pathway. Furthermore, we have recently shown that nitrofen inhibits PI3K-AKT signaling during mid-to-late lung morphogenesis in the nitrofen-induced hypoplastic lung. We hypothesized that prenatal administration of RA upregulates pulmonary gene expression of PI3K and AKT in the nitrofen-induced hypoplastic lung. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). 5 mg/kg of RA was given on D18, D19 and D20. The fetuses were harvested on D21, and fetal lungs were obtained and divided into four groups: control, control + RA, nitrofen, nitrofen + RA. The mRNA expression levels of PI3K and AKT were analyzed in each lung by real-time RT-PCR and statistically analyzed. Immunohistochemistry was also performed to evaluate protein expression of PI3K and AKT in the fetal lungs at D21. The pulmonary gene expression levels of PI3K and AKT were significantly upregulated in nitrofen + RA group compared to nitrofen group and control + RA group (p < 0.05), whereas there were no significant differences between controls and control + RA group. Immunoreactivity of PI3K and AKT was markedly increased in nitrofen + RA lungs compared to nitrofen-induced hypoplastic lungs. Upregulation of PI3K and AKT genes after prenatal treatment with RA in the nitrofen-induced hypoplastic lung suggests that RA may have a therapeutic potential in modulating lung alveologenesis by stimulating epithelial

Porcine MAP3K5 analysis: molecular cloning, characterization, tissue expression pattern, and copy number variations associated with residual feed intake.

PubMed

Pu, L; Zhang, L C; Zhang, J S; Song, X; Wang, L G; Liang, J; Zhang, Y B; Liu, X; Yan, H; Zhang, T; Yue, J W; Li, N; Wu, Q Q; Wang, L X

2016-08-12

Mitogen-activated protein kinase kinase kinase 5 (MAP3K5) is essential for apoptosis, proliferation, differentiation, and immune responses, and is a candidate marker for residual feed intake (RFI) in pig. We cloned the full-length cDNA sequence of porcine MAP3K5 by rapid-amplification of cDNA ends. The 5451-bp gene contains a 5'-untranslated region (UTR) (718 bp), a coding region (3738 bp), and a 3'-UTR (995 bp), and encodes a peptide of 1245 amino acids, which shares 97, 99, 97, 93, 91, and 84% sequence identity with cattle, sheep, human, mouse, chicken, and zebrafish MAP3K5, respectively. The deduced MAP3K5 protein sequence contains two conserved domains: a DUF4071 domain and a protein kinase domain. Phylogenetic analysis showed that porcine MAP3K5 forms a separate branch to vicugna and camel MAP3K5. Tissue expression analysis using real-time quantitative polymerase chain reaction (qRT-PCR) revealed that MAP3K5 was expressed in the heart, liver, spleen, lung, kidney, muscle, fat, pancrea, ileum, and stomach tissues. Copy number variation was detected for porcine MAP3K5 and validated by qRT-PCR. Furthermore, a significant increase in average copy number was detected in the low RFI group when compared to the high RFI group in a Duroc pig population. These results provide useful information regarding the influence of MAP3K5 on RFI in pigs.

Dynamic association of epigenetic H3K4me3 and DNA 5hmC marks in the dorsal hippocampus and anterior cingulate cortex following reactivation of a fear memory.

PubMed

Webb, William M; Sanchez, Richard G; Perez, Gabriella; Butler, Anderson A; Hauser, Rebecca M; Rich, Megan C; O'Bierne, Aidan L; Jarome, Timothy J; Lubin, Farah D

2017-07-01

Epigenetic mechanisms such as DNA methylation and histone methylation are critical regulators of gene transcription changes during memory consolidation. However, it is unknown how these epigenetic modifications coordinate control of gene expression following reactivation of a previously consolidated memory. Here, we found that retrieval of a recent contextual fear conditioned memory increased global levels of H3 lysine 4-trimethylation (H3K4me3) and DNA 5-hydroxymethylation (5hmC) in area CA1 of the dorsal hippocampus. Further experiments revealed increased levels of H3K4me3 and DNA 5hmC within a CpG-enriched coding region of the Npas4, but not c-fos, gene. Intriguingly, retrieval of a 30-day old memory increased H3K4me3 and DNA 5hmC levels at a CpG-enriched coding region of c-fos, but not Npas4, in the anterior cingulate cortex, suggesting that while these two epigenetic mechanisms co-occur following the retrieval of a recent or remote memory, their gene targets differ depending on the brain region. Additionally, we found that in vivo siRNA-mediated knockdown of the H3K4me3 methyltransferase Mll1 in CA1 abolished retrieval-induced increases in DNA 5hmC levels at the Npas4 gene, suggesting that H3K4me3 couples to DNA 5hmC mechanisms. Consistent with this, loss of Mll1 prevented retrieval-induced increases in Npas4 mRNA levels in CA1 and impaired fear memory. Collectively, these findings suggest an important link between histone methylation and DNA hydroxymethylation mechanisms in the epigenetic control of de novo gene transcription triggered by memory retrieval. Copyright © 2017 Elsevier Inc. All rights reserved.

Effects of Vitamin K3 and K5 on Daunorubicin-resistant Human T Lymphoblastoid Leukemia Cells.

PubMed

Nakaoka, Eri; Tanaka, Sachiko; Onda, Kenji; Sugiyama, Kentaro; Hirano, Toshihiko

2015-11-01

Anticancer efficacy of vitamin K derivatives on multidrug-resistant cancer cells has been scarcely investigated. The effects of vitamins K3 and K5 on proliferation of human leukemia MOLT-4 cells and on daunorubicin-resistant MOLT-4/DNR cells were estimated by a WST assay. Apoptotic cells were detected by Annexin V and propidium iodide staining, followed by flow cytometry. Vitamins K3 and K5 significantly inhibited proliferation of leukemic cells at 10 and 100 μM (p<0.05), and these effects were almost equally observed in both MOLT-4 and MOLT/DNR drug-resistant cells. Vitamin K3 induced cell apoptosis at 10 and 100 μM in both MOLT-4 and MOLT-4/DNR cells (p<0.05). Vitamin K5 also increased apoptotic cells, while rather inducing necrotic cell death. Vitamins K3 and K5 suppress MOLT-4 and MOLT-4/DNR cell-proliferation partially through induction of apoptosis, and these vitamin derivatives can overcome drug resistance due to P-glycoprotein expression. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Polymorphisms of Plasmodium falciparum k13-propeller gene among migrant workers returning to Henan Province, China from Africa.

PubMed

Yang, Chengyun; Zhang, Hongwei; Zhou, Ruimin; Qian, Dan; Liu, Ying; Zhao, Yuling; Li, Suhua; Xu, Bianli

2017-08-10

Henan Province has been in the malaria elimination stage, with all reports of the disease being imported since 2012 and over 90% coming from Africa. Surveillance and population studies are essential for the early detection and subsequent prevention of the spread of drug resistance. The K13-propeller gene was recently identified as a proposed molecular marker of artemisinin (ART) resistance. In this study, we detected mutations of the K13-propeller gene in samples taken from imported malaria cases in Henan Province from 2012 to 2015. There were 483 samples that were obtained from Plasmodium falciparum-infected malaria migrant workers who returned to Henan Province from Africa between 2012 and 2015. The single nucleotide polymorphisms in the K13-propeller gene were assessed by nested PCR with DNA sequencing. Frequency and geographic difference of K13-propeller gene mutant types were analyzed. Of 483 patients, 476 were cured and 7 died. There were no K13-propeller mutations in the blood samples from the 7 patients who died, but there were 23 different genotypes of the K13-propeller that were observed in 24 (4.97%) of the samples. C580Y, which was the predominant one in the resistance of ART, was not detected in the samples, but R539T and P574L which have also been associated with ART resistance, were observed in two samples from Angola and Equatorial Guinea. No mutations were detected in 11 samples from North Africa. The frequency of the K13-propeller was 6.50% (8/123) in Central Africa, followed by East Africa (1/19, 5.26%), West Africa (9/198, 4.55%) and South Africa (6/132, 4.55%). There was no significant difference among these four areas (P = 0.795). R539T and P574L were found in migrant workers who traveled from Africa to Henan Province, although the frequency of the K13-propeller mutants was low. These data may enrich the molecular surveillance of antimalarial resistance and will be helpful for developing and updating the antimalarial policy in Henan Province.

The Arabidopsis thaliana HAK5 K+ transporter is required for plant growth and K+ acquisition from low K+ solutions under saline conditions.

PubMed

Nieves-Cordones, Manuel; Alemán, Fernando; Martínez, Vicente; Rubio, Francisco

2010-03-01

K(+) uptake in the high-affinity range of concentrations and its components have been widely studied. In Arabidposis thaliana, the AtHAK5 transporter and the AtAKT1 channel have been shown to be the main transport proteins involved in this process. Here, we study the role of these two systems under two important stress conditions: low K(+) supply or the presence of salinity. T-DNA insertion lines disrupting AtHAK5 and AtAKT1 are employed for long-term experiments that allow physiological characterization of the mutant lines. We found that AtHAK5 is required for K(+) absorption necessary to sustain plant growth at low K(+) in the absence as well as in the presence of salinity. Salinity greatly reduced AtHAK5 transcript levels and promoted AtAKT1-mediated K(+) efflux, resulting in an important impairment of K(+) nutrition. Although having a limited capacity, AtHAK5 plays a major role for K(+) acquisition from low K(+) concentrations in the presence of salinity.

Novel genetic associations for blood pressure identified via gene-alcohol interaction in up to 570K individuals across multiple ancestries

PubMed Central

Guo, Xiuqing; Franceschini, Nora; Cheng, Ching-Yu; Sim, Xueling; Vojinovic, Dina; Marten, Jonathan; Musani, Solomon K.; Li, Changwei; Schwander, Karen; Richard, Melissa A.; Noordam, Raymond; Aschard, Hugues; Bartz, Traci M.; Bielak, Lawrence F.; Dorajoo, Rajkumar; Fisher, Virginia; Hartwig, Fernando P.; Horimoto, Andrea R. V. R.; Lohman, Kurt K.; Manning, Alisa K.; Rankinen, Tuomo; Smith, Albert V.; Wojczynski, Mary K.; Alver, Maris; Boissel, Mathilde; Cai, Qiuyin; Divers, Jasmin; Gao, Chuan; Goel, Anuj; Harris, Sarah E.; He, Meian; Hsu, Fang-Chi; Jackson, Anne U.; Kähönen, Mika; Kasturiratne, Anuradhani; Komulainen, Pirjo; Kühnel, Brigitte; Laguzzi, Federica; Luan, Jian'an; Nolte, Ilja M.; Padmanabhan, Sandosh; Robino, Antonietta; Scott, Robert A.; Sofer, Tamar; Stančáková, Alena; Takeuchi, Fumihiko; Tayo, Bamidele O.; Varga, Tibor V.; Vitart, Veronique; Wang, Yajuan; Warren, Helen R.; Wen, Wanqing; Yanek, Lisa R.; Zhang, Weihua; Zhao, Jing Hua; Afaq, Saima; Amin, Najaf; Arking, Dan E.; Aung, Tin; Boerwinkle, Eric; Borecki, Ingrid; Broeckel, Ulrich; Brown, Morris; Brumat, Marco; Burke, Gregory L.; Chakravarti, Aravinda; Charumathi, Sabanayagam; Ida Chen, Yii-Der; Connell, John M.; Correa, Adolfo; de las Fuentes, Lisa; de Mutsert, Renée; de Silva, H. Janaka; Deng, Xuan; Ding, Jingzhong; Duan, Qing; Eaton, Charles B.; Ehret, Georg; Eppinga, Ruben N.; Faul, Jessica D.; Felix, Stephan B.; Forouhi, Nita G.; Forrester, Terrence; Franco, Oscar H.; Friedlander, Yechiel; Gandin, Ilaria; Gao, He; Ghanbari, Mohsen; Gigante, Bruna; Gu, C. Charles; Gu, Dongfeng; Hagenaars, Saskia P.; Hallmans, Göran; Harris, Tamara B.; He, Jiang; Heng, Chew-Kiat; Hirata, Makoto; Howard, Barbara V.; Ikram, M. Arfan; John, Ulrich; Katsuya, Tomohiro; Khor, Chiea Chuen; Kilpeläinen, Tuomas O.; Koh, Woon-Puay; Krieger, José E.; Kritchevsky, Stephen B.; Kubo, Michiaki; Kuusisto, Johanna; Lakka, Timo A.; Langefeld, Carl D.; Langenberg, Claudia; Launer, Lenore J.; Lehne, Benjamin; Lewis, Cora E.; Li, Yize; Lin, Shiow; Liu, Jianjun; Liu, Jingmin; Loh, Marie; Louie, Tin; Mägi, Reedik; McKenzie, Colin A.; Meitinger, Thomas; Milaneschi, Yuri; Milani, Lili; Mohlke, Karen L.; Momozawa, Yukihide; Nalls, Mike A.; Nelson, Christopher P.; Sotoodehnia, Nona; Norris, Jill M.; O'Connell, Jeff R.; Palmer, Nicholette D.; Perls, Thomas; Pedersen, Nancy L.; Peters, Annette; Peyser, Patricia A.; Poulter, Neil; Raffel, Leslie J.; Raitakari, Olli T.; Roll, Kathryn; Rose, Lynda M.; Rosendaal, Frits R.; Rotter, Jerome I.; Schmidt, Carsten O.; Schreiner, Pamela J.; Schupf, Nicole; Scott, William R.; Shi, Yuan; Sidney, Stephen; Sims, Mario; Sitlani, Colleen M.; Smith, Jennifer A.; Snieder, Harold; Starr, John M.; Strauch, Konstantin; Stringham, Heather M.; Tan, Nicholas Y. Q.; Tang, Hua; Taylor, Kent D.; Teo, Yik Ying; Tham, Yih Chung; Turner, Stephen T.; Uitterlinden, André G.; Vollenweider, Peter; Waldenberger, Melanie; Wang, Lihua; Wang, Ya Xing; Wei, Wen Bin; Williams, Christine; Yao, Jie; Yu, Caizheng; Yuan, Jian-Min; Zhao, Wei; Zonderman, Alan B.; Becker, Diane M.; Boehnke, Michael; Bowden, Donald W.; Chambers, John C.; Deary, Ian J.; Esko, Tõnu; Farrall, Martin; Franks, Paul W.; Freedman, Barry I.; Froguel, Philippe; Gasparini, Paolo; Gieger, Christian; Kamatani, Yoichiro; Kato, Norihiro; Kooner, Jaspal S.; Kutalik, Zoltán; Laakso, Markku; Laurie, Cathy C.; Leander, Karin; Lehtimäki, Terho; Study, Lifelines Cohort; Magnusson, Patrik K. E.; Oldehinkel, Albertine J.; Penninx, Brenda W. J. H.; Polasek, Ozren; Porteous, David J.; Rauramaa, Rainer; Samani, Nilesh J.; Scott, James; Shu, Xiao-Ou; van der Harst, Pim; Wagenknecht, Lynne E.; Watkins, Hugh; Weir, David R.; Wickremasinghe, Ananda R.; Wu, Tangchun; Zheng, Wei; Bouchard, Claude; Christensen, Kaare; Evans, Michele K.; Gudnason, Vilmundur; Horta, Bernardo L.; Kardia, Sharon L. R.; Liu, Yongmei; Pereira, Alexandre C.; Psaty, Bruce M.; Ridker, Paul M.; van Dam, Rob M.; Gauderman, W. James; Zhu, Xiaofeng; Mook-Kanamori, Dennis O.; Fornage, Myriam; Rotimi, Charles N.; Cupples, L. Adrienne; Kelly, Tanika N.; Fox, Ervin R.; Hayward, Caroline; van Duijn, Cornelia M.; Tai, E Shyong; Wong, Tien Yin; Kooperberg, Charles; Palmas, Walter; Morrison, Alanna C.; Caulfield, Mark J.; Munroe, Patricia B.; Rao, Dabeeru C.; Province, Michael A.; Levy, Daniel

2018-01-01

Heavy alcohol consumption is an established risk factor for hypertension; the mechanism by which alcohol consumption impact blood pressure (BP) regulation remains unknown. We hypothesized that a genome-wide association study accounting for gene-alcohol consumption interaction for BP might identify additional BP loci and contribute to the understanding of alcohol-related BP regulation. We conducted a large two-stage investigation incorporating joint testing of main genetic effects and single nucleotide variant (SNV)-alcohol consumption interactions. In Stage 1, genome-wide discovery meta-analyses in ≈131K individuals across several ancestry groups yielded 3,514 SNVs (245 loci) with suggestive evidence of association (P < 1.0 x 10−5). In Stage 2, these SNVs were tested for independent external replication in ≈440K individuals across multiple ancestries. We identified and replicated (at Bonferroni correction threshold) five novel BP loci (380 SNVs in 21 genes) and 49 previously reported BP loci (2,159 SNVs in 109 genes) in European ancestry, and in multi-ancestry meta-analyses (P < 5.0 x 10−8). For African ancestry samples, we detected 18 potentially novel BP loci (P < 5.0 x 10−8) in Stage 1 that warrant further replication. Additionally, correlated meta-analysis identified eight novel BP loci (11 genes). Several genes in these loci (e.g., PINX1, GATA4, BLK, FTO and GABBR2) have been previously reported to be associated with alcohol consumption. These findings provide insights into the role of alcohol consumption in the genetic architecture of hypertension. PMID:29912962

Novel genetic associations for blood pressure identified via gene-alcohol interaction in up to 570K individuals across multiple ancestries.

PubMed

Feitosa, Mary F; Kraja, Aldi T; Chasman, Daniel I; Sung, Yun J; Winkler, Thomas W; Ntalla, Ioanna; Guo, Xiuqing; Franceschini, Nora; Cheng, Ching-Yu; Sim, Xueling; Vojinovic, Dina; Marten, Jonathan; Musani, Solomon K; Li, Changwei; Bentley, Amy R; Brown, Michael R; Schwander, Karen; Richard, Melissa A; Noordam, Raymond; Aschard, Hugues; Bartz, Traci M; Bielak, Lawrence F; Dorajoo, Rajkumar; Fisher, Virginia; Hartwig, Fernando P; Horimoto, Andrea R V R; Lohman, Kurt K; Manning, Alisa K; Rankinen, Tuomo; Smith, Albert V; Tajuddin, Salman M; Wojczynski, Mary K; Alver, Maris; Boissel, Mathilde; Cai, Qiuyin; Campbell, Archie; Chai, Jin Fang; Chen, Xu; Divers, Jasmin; Gao, Chuan; Goel, Anuj; Hagemeijer, Yanick; Harris, Sarah E; He, Meian; Hsu, Fang-Chi; Jackson, Anne U; Kähönen, Mika; Kasturiratne, Anuradhani; Komulainen, Pirjo; Kühnel, Brigitte; Laguzzi, Federica; Luan, Jian'an; Matoba, Nana; Nolte, Ilja M; Padmanabhan, Sandosh; Riaz, Muhammad; Rueedi, Rico; Robino, Antonietta; Said, M Abdullah; Scott, Robert A; Sofer, Tamar; Stančáková, Alena; Takeuchi, Fumihiko; Tayo, Bamidele O; van der Most, Peter J; Varga, Tibor V; Vitart, Veronique; Wang, Yajuan; Ware, Erin B; Warren, Helen R; Weiss, Stefan; Wen, Wanqing; Yanek, Lisa R; Zhang, Weihua; Zhao, Jing Hua; Afaq, Saima; Amin, Najaf; Amini, Marzyeh; Arking, Dan E; Aung, Tin; Boerwinkle, Eric; Borecki, Ingrid; Broeckel, Ulrich; Brown, Morris; Brumat, Marco; Burke, Gregory L; Canouil, Mickaël; Chakravarti, Aravinda; Charumathi, Sabanayagam; Ida Chen, Yii-Der; Connell, John M; Correa, Adolfo; de Las Fuentes, Lisa; de Mutsert, Renée; de Silva, H Janaka; Deng, Xuan; Ding, Jingzhong; Duan, Qing; Eaton, Charles B; Ehret, Georg; Eppinga, Ruben N; Evangelou, Evangelos; Faul, Jessica D; Felix, Stephan B; Forouhi, Nita G; Forrester, Terrence; Franco, Oscar H; Friedlander, Yechiel; Gandin, Ilaria; Gao, He; Ghanbari, Mohsen; Gigante, Bruna; Gu, C Charles; Gu, Dongfeng; Hagenaars, Saskia P; Hallmans, Göran; Harris, Tamara B; He, Jiang; Heikkinen, Sami; Heng, Chew-Kiat; Hirata, Makoto; Howard, Barbara V; Ikram, M Arfan; John, Ulrich; Katsuya, Tomohiro; Khor, Chiea Chuen; Kilpeläinen, Tuomas O; Koh, Woon-Puay; Krieger, José E; Kritchevsky, Stephen B; Kubo, Michiaki; Kuusisto, Johanna; Lakka, Timo A; Langefeld, Carl D; Langenberg, Claudia; Launer, Lenore J; Lehne, Benjamin; Lewis, Cora E; Li, Yize; Lin, Shiow; Liu, Jianjun; Liu, Jingmin; Loh, Marie; Louie, Tin; Mägi, Reedik; McKenzie, Colin A; Meitinger, Thomas; Metspalu, Andres; Milaneschi, Yuri; Milani, Lili; Mohlke, Karen L; Momozawa, Yukihide; Nalls, Mike A; Nelson, Christopher P; Sotoodehnia, Nona; Norris, Jill M; O'Connell, Jeff R; Palmer, Nicholette D; Perls, Thomas; Pedersen, Nancy L; Peters, Annette; Peyser, Patricia A; Poulter, Neil; Raffel, Leslie J; Raitakari, Olli T; Roll, Kathryn; Rose, Lynda M; Rosendaal, Frits R; Rotter, Jerome I; Schmidt, Carsten O; Schreiner, Pamela J; Schupf, Nicole; Scott, William R; Sever, Peter S; Shi, Yuan; Sidney, Stephen; Sims, Mario; Sitlani, Colleen M; Smith, Jennifer A; Snieder, Harold; Starr, John M; Strauch, Konstantin; Stringham, Heather M; Tan, Nicholas Y Q; Tang, Hua; Taylor, Kent D; Teo, Yik Ying; Tham, Yih Chung; Turner, Stephen T; Uitterlinden, André G; Vollenweider, Peter; Waldenberger, Melanie; Wang, Lihua; Wang, Ya Xing; Wei, Wen Bin; Williams, Christine; Yao, Jie; Yu, Caizheng; Yuan, Jian-Min; Zhao, Wei; Zonderman, Alan B; Becker, Diane M; Boehnke, Michael; Bowden, Donald W; Chambers, John C; Deary, Ian J; Esko, Tõnu; Farrall, Martin; Franks, Paul W; Freedman, Barry I; Froguel, Philippe; Gasparini, Paolo; Gieger, Christian; Jonas, Jost Bruno; Kamatani, Yoichiro; Kato, Norihiro; Kooner, Jaspal S; Kutalik, Zoltán; Laakso, Markku; Laurie, Cathy C; Leander, Karin; Lehtimäki, Terho; Study, Lifelines Cohort; Magnusson, Patrik K E; Oldehinkel, Albertine J; Penninx, Brenda W J H; Polasek, Ozren; Porteous, David J; Rauramaa, Rainer; Samani, Nilesh J; Scott, James; Shu, Xiao-Ou; van der Harst, Pim; Wagenknecht, Lynne E; Wareham, Nicholas J; Watkins, Hugh; Weir, David R; Wickremasinghe, Ananda R; Wu, Tangchun; Zheng, Wei; Bouchard, Claude; Christensen, Kaare; Evans, Michele K; Gudnason, Vilmundur; Horta, Bernardo L; Kardia, Sharon L R; Liu, Yongmei; Pereira, Alexandre C; Psaty, Bruce M; Ridker, Paul M; van Dam, Rob M; Gauderman, W James; Zhu, Xiaofeng; Mook-Kanamori, Dennis O; Fornage, Myriam; Rotimi, Charles N; Cupples, L Adrienne; Kelly, Tanika N; Fox, Ervin R; Hayward, Caroline; van Duijn, Cornelia M; Tai, E Shyong; Wong, Tien Yin; Kooperberg, Charles; Palmas, Walter; Rice, Kenneth; Morrison, Alanna C; Elliott, Paul; Caulfield, Mark J; Munroe, Patricia B; Rao, Dabeeru C; Province, Michael A; Levy, Daniel

2018-01-01

Heavy alcohol consumption is an established risk factor for hypertension; the mechanism by which alcohol consumption impact blood pressure (BP) regulation remains unknown. We hypothesized that a genome-wide association study accounting for gene-alcohol consumption interaction for BP might identify additional BP loci and contribute to the understanding of alcohol-related BP regulation. We conducted a large two-stage investigation incorporating joint testing of main genetic effects and single nucleotide variant (SNV)-alcohol consumption interactions. In Stage 1, genome-wide discovery meta-analyses in ≈131K individuals across several ancestry groups yielded 3,514 SNVs (245 loci) with suggestive evidence of association (P < 1.0 x 10-5). In Stage 2, these SNVs were tested for independent external replication in ≈440K individuals across multiple ancestries. We identified and replicated (at Bonferroni correction threshold) five novel BP loci (380 SNVs in 21 genes) and 49 previously reported BP loci (2,159 SNVs in 109 genes) in European ancestry, and in multi-ancestry meta-analyses (P < 5.0 x 10-8). For African ancestry samples, we detected 18 potentially novel BP loci (P < 5.0 x 10-8) in Stage 1 that warrant further replication. Additionally, correlated meta-analysis identified eight novel BP loci (11 genes). Several genes in these loci (e.g., PINX1, GATA4, BLK, FTO and GABBR2) have been previously reported to be associated with alcohol consumption. These findings provide insights into the role of alcohol consumption in the genetic architecture of hypertension.

A novel harmony search-K means hybrid algorithm for clustering gene expression data

PubMed Central

Nazeer, KA Abdul; Sebastian, MP; Kumar, SD Madhu

2013-01-01

Recent progress in bioinformatics research has led to the accumulation of huge quantities of biological data at various data sources. The DNA microarray technology makes it possible to simultaneously analyze large number of genes across different samples. Clustering of microarray data can reveal the hidden gene expression patterns from large quantities of expression data that in turn offers tremendous possibilities in functional genomics, comparative genomics, disease diagnosis and drug development. The k- ¬means clustering algorithm is widely used for many practical applications. But the original k-¬means algorithm has several drawbacks. It is computationally expensive and generates locally optimal solutions based on the random choice of the initial centroids. Several methods have been proposed in the literature for improving the performance of the k-¬means algorithm. A meta-heuristic optimization algorithm named harmony search helps find out near-global optimal solutions by searching the entire solution space. Low clustering accuracy of the existing algorithms limits their use in many crucial applications of life sciences. In this paper we propose a novel Harmony Search-K means Hybrid (HSKH) algorithm for clustering the gene expression data. Experimental results show that the proposed algorithm produces clusters with better accuracy in comparison with the existing algorithms. PMID:23390351

A novel harmony search-K means hybrid algorithm for clustering gene expression data.

PubMed

Nazeer, Ka Abdul; Sebastian, Mp; Kumar, Sd Madhu

2013-01-01

Recent progress in bioinformatics research has led to the accumulation of huge quantities of biological data at various data sources. The DNA microarray technology makes it possible to simultaneously analyze large number of genes across different samples. Clustering of microarray data can reveal the hidden gene expression patterns from large quantities of expression data that in turn offers tremendous possibilities in functional genomics, comparative genomics, disease diagnosis and drug development. The k- ¬means clustering algorithm is widely used for many practical applications. But the original k-¬means algorithm has several drawbacks. It is computationally expensive and generates locally optimal solutions based on the random choice of the initial centroids. Several methods have been proposed in the literature for improving the performance of the k-¬means algorithm. A meta-heuristic optimization algorithm named harmony search helps find out near-global optimal solutions by searching the entire solution space. Low clustering accuracy of the existing algorithms limits their use in many crucial applications of life sciences. In this paper we propose a novel Harmony Search-K means Hybrid (HSKH) algorithm for clustering the gene expression data. Experimental results show that the proposed algorithm produces clusters with better accuracy in comparison with the existing algorithms.

Major genes and QTL influencing wool production and quality: a review.

PubMed

Purvis, Ian William; Franklin, Ian Robert

2005-01-01

The opportunity exists to utilise our knowledge of major genes that influence the economically important traits in wool sheep. Genes with Mendelian inheritance have been identified for many important traits in wool sheep. Of particular importance are genes influencing pigmentation, wool quality and the keratin proteins, the latter of which are important for the morphology of the wool fibre. Gene mapping studies have identified some chromosomal regions associated with variation in wool quality and production traits. The challenge now is to build on this knowledge base in a cost-effective way to deliver molecular tools that facilitate enhanced genetic improvement programs for wool sheep.

Complete cDNA sequence and amino acid analysis of a bovine ribonuclease K6 gene.

PubMed

Pietrowski, D; Förster, M

2000-01-01

The complete cDNA sequence of a ribonuclease k6 gene of Bos Taurus has been determined. It codes for a protein with 154 amino acids and contains the invariant cysteine, histidine and lysine residues as well as the characteristic motifs specific to ribonuclease active sites. The deduced protein sequence is 27 residues longer than other known ribonucleases k6 and shows amino acids exchanges which could reflect a strain specificity or polymorphism within the bovine genome. Based on sequence similarity we have termed the identified gene bovine ribonuclease k6 b (brk6b).

Genetic heterogeneity of the dnaK gene locus including transcription terminator region (TTR) in Campylobacter lari.

PubMed

Shitara, M; Tsuboi, Y; Sekizuka, T; Tazumi, A; Moorei, J E; Millar, B C; Taneike, I; Matsuda, M

2008-01-01

Nucleotide sequences of approximately 3.1 kbp consisting of the full-length open reading frame (ORF) for grpE, a non-coding (NC) region and a putative ORF for the full-length dnaK gene (1860 bp) were identified from a urease-positive thermophilic Campylobacter (UPTC) CF89-12 isolate. Then, following the construction of a new degenerate polymerase chain reaction (PCR) primer pair for amplification of the dnaK structural gene, including the transcription terminator region of C. lari isolates, the dnaK region was amplified successfully, TA-cloned and sequenced in nine C. lari isolates. The dnaK gene sequences commenced with an ATG and terminated with a TAA in all 10 isolates, including CF89-12. In addition, the putative ORFs for the dnaK gene locus from seven UPTC isolates consisted of 1860 bases, and the four urease-negative (UN) C. lari isolates included C. lari RM2100 reference strain 1866. Interestingly, different probable ribosome binding sites and hypothetically intrinsic p-independent terminator structures were identified between the seven UPTC and four UN C. lari isolates, respectively. Moreover, it is interesting to note that 20 out of a total of 28 polymorphic sites occurred among amino acid sequences of the dnaK ORF from 11 C. lari isolates, identified to be alternatively UPTC-specific or UN C. lari-specific. In the neighbour-joining tree based on the nucleotide sequence information of the dnaK gene, C. lari forms two major distinct clusters consisting of UPTC and UN C. lari isolates, respectively, with UN C. lari being more closely related to other thermophilic campylobacters than to UPTC.

The Human Papillomavirus Type 16 E6 Gene Alone Is Sufficient To Induce Carcinomas in Transgenic Animals

PubMed Central

Song, Shiyu; Pitot, Henry C.; Lambert, Paul F.

1999-01-01

High-risk human papillomaviruses (HPVs) are the causative agents of certain human cancers. HPV type 16 (HPV16) is the papillomavirus most frequently associated with cervical cancer in women. The E6 and E7 genes of HPV are expressed in cells derived from these cancers and can transform cells in tissue culture. Animal experiments have demonstrated that E6 and E7 together cause tumors. We showed previously that E6 and E7 together or E7 alone could induce skin tumors in mice when these genes were expressed in the basal epithelia of the skin. In this study, we investigated the role that the E6 gene plays in carcinogenesis. We generated K14E6 transgenic mice, in which the HPV16 E6 gene was directed in its expression by the human keratin 14 promoter (hK14) to the basal layer of the epidermis. We found that E6 induced cellular hyperproliferation and epidermal hyperplasia and caused skin tumors in adult mice. Interestingly, the tumors derived from E6 were mostly malignant, as opposed to the tumors from E7 mice, which were mostly benign. This result leads us to hypothesize that E6 may contribute differently than E7 to HPV-associated carcinogenesis; whereas E7 primarily contributes to the early stages of carcinogenesis that lead to the formation of benign tumors, E6 primarily contributes to the late stages of carcinogenesis that lead to malignancy. PMID:10364340

Reexamining Transcriptional Regulation of the Bacillus subtilis htpX Gene and the ykrK Gene, Encoding a Novel Type of Transcriptional Regulator, and Redefining the YkrK Operator

PubMed Central

Lin, Ta-Hui; Huang, Shih-Chien

2012-01-01

HtpX is an integral cytoplasmic membrane metalloprotease well conserved in numerous bacteria. A recent study showed that expression of the Bacillus subtilis htpX gene is under dual negative control by Rok and a novel type of transcriptional regulator, YkrK. Here we report that expression of the B. subtilis htpX gene is strongly heat inducible. Contrary to the previous prediction, ykrK expression has been found to be not subject to autoregulation. We have identified the htpX promoter and the authentic ykrK promoter, which is also distinct from the previously predicted one. We have redefined a conserved inverted repeat sequence to be the YkrK operator, which is somewhat different from the previously proposed one. We provide evidence that YkrK is not a substrate of HtpX and that heat induction of htpX is not YkrK mediated. We have also found that the absence of FtsH or HtpX alone did not impair B. subtilis cell viability on LB agar plates at high temperature, whereas the absence of both FtsH and HtpX caused a severe growth defect under heat stress. This finding supports the notion that FtsH and HtpX may have partially overlapping functions in heat resistance. Finally, we show that htpX expression is subject to transient negative control by sigB under heat stress in a Rok- and YkrK-independent manner. Triple negative control of htpX expression at high temperature by rok, sigB, and ykrK may help cells to prevent uncontrolled and detrimental oversynthesis of the HtpX protease. PMID:23042994

Vitamins K2, K3 and K5 exert antitumor effects on established colorectal cancer in mice by inducing apoptotic death of tumor cells.

PubMed

Ogawa, Mutsumi; Nakai, Seiji; Deguchi, Akihiro; Nonomura, Takako; Masaki, Tsutomu; Uchida, Naohito; Yoshiji, Hitoshi; Kuriyama, Shigeki

2007-08-01

Although a number of studies have shown that vitamin K possesses antitumor activities on various neoplastic cell lines, there are few reports demonstrating in vivo antitumor effects of vitamin K, and the antitumor effect on colorectal cancer (CRC) remains to be examined. Therefore, antitumor effects of vitamin K on CRC were examined both in vitro and in vivo. Vitamins K2, K3 and K5 suppressed the proliferation of colon 26 cells in a dose-dependent manner, while vitamin K1 did not. On flow cytometry, induction of apoptosis by vitamins K2, K3 and K5 was suggested by population in sub-G1 phase of the cell cycle. Hoechst 33342 staining and a two-color flow cytometric assay using fluorescein isothiocyanate-conjugated annexin V and propidium iodide confirmed that vitamins K2, K3 and K5 induced apoptotic death of colon 26 cells. Enzymatic activity of caspase-3 in colon 26 cells was significantly up-regulated by vitamins K2, K3 and K5. The pan-caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone, substantially prevented vitamin K-mediated apoptosis. In vivo study using syngeneic mice with subcutaneously established colon 26 tumors demonstrated that intravenous administration of vitamins K2, K3 and K5 significantly suppressed the tumor growth. The number of apoptotic tumor cells was significantly larger in the vitamin K-treated groups than in the control group. These results suggest that vitamins K2, K3 and K5 exerted effective antitumor effects on CRC in vitro and in vivo by inducing caspase-dependent apoptotic death of tumor cells, suggesting that these K vitamins may be promising agents for the treatment of patients with CRC.