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1.
Dev Cell ; 59(2): 187-198.e7, 2024 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-38198888

RESUMO

Chromatin organization is essential for maintaining cell-fate trajectories and developmental programs. Here, we find that disruption of H3K36 methylation dramatically impairs normal epithelial differentiation and development, which promotes increased cellular plasticity and enrichment of alternative cell fates. Specifically, we observe a striking increase in the aberrant generation of excessive epithelial glandular tissues, including hypertrophic salivary, sebaceous, and meibomian glands, as well as enhanced squamous tumorigenesis. These phenotypic and gene expression manifestations are associated with loss of H3K36me2 and rewiring of repressive H3K27me3, changes we also observe in human patients with glandular hyperplasia. Collectively, these results have identified a critical role for H3K36 methylation in both in vivo epithelial cell-fate decisions and the prevention of squamous carcinogenesis and suggest that H3K36 methylation modulation may offer new avenues for the treatment of numerous common disorders driven by altered glandular function, which collectively affect large segments of the human population.


Assuntos
Carcinoma de Células Escamosas , Histonas , Humanos , Histonas/metabolismo , Plasticidade Celular , Metilação , Carcinogênese/genética , Carcinoma de Células Escamosas/genética
3.
Sci Adv ; 9(35): eadg5234, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37656787

RESUMO

N6-methyladenosine (m6A) is the most abundant modification on messenger RNAs (mRNAs) and is catalyzed by methyltransferase-like protein 3 (Mettl3). To understand the role of m6A in a self-renewing somatic tissue, we deleted Mettl3 in epidermal progenitors in vivo. Mice lacking Mettl3 demonstrate marked features of dysfunctional development and self-renewal, including a loss of hair follicle morphogenesis and impaired cell adhesion and polarity associated with oral ulcerations. We show that Mettl3 promotes the m6A-mediated degradation of mRNAs encoding critical histone modifying enzymes. Depletion of Mettl3 results in the loss of m6A on these mRNAs and increases their expression and associated modifications, resulting in widespread gene expression abnormalities that mirror the gross phenotypic abnormalities. Collectively, these results have identified an additional layer of gene regulation within epithelial tissues, revealing an essential role for m6A in the regulation of chromatin modifiers, and underscoring a critical role for Mettl3-catalyzed m6A in proper epithelial development and self-renewal.


Assuntos
Histonas , Metiltransferases , Animais , Camundongos , Metiltransferases/genética , Adenosina , Adesão Celular , RNA Mensageiro , Catálise
4.
bioRxiv ; 2023 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-36993480

RESUMO

The versatility of somatosensation arises from heterogeneous dorsal root ganglion (DRG) neurons. However, soma transcriptomes of individual human DRG (hDRG) neurons-critical in-formation to decipher their functions-are lacking due to technical difficulties. Here, we developed a novel approach to isolate individual hDRG neuron somas for deep RNA sequencing (RNA-seq). On average, >9,000 unique genes per neuron were detected, and 16 neuronal types were identified. Cross-species analyses revealed remarkable divergence among pain-sensing neurons and the existence of human-specific nociceptor types. Our deep RNA-seq dataset was especially powerful for providing insight into the molecular mechanisms underlying human somatosensation and identifying high potential novel drug targets. Our dataset also guided the selection of molecular markers to visualize different types of human afferents and the discovery of novel functional properties using single-cell in vivo electrophysiological recordings. In summary, by employing a novel soma sequencing method, we generated an unprecedented hDRG neuron atlas, providing new insights into human somatosensation, establishing a critical foundation for translational work, and clarifying human species-species properties.

5.
Sci Transl Med ; 14(630): eabj0324, 2022 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-35108061

RESUMO

Skin is composed of diverse cell populations that cooperatively maintain homeostasis. Up-regulation of the nuclear factor κB (NF-κB) pathway may lead to the development of chronic inflammatory disorders of the skin, but its role during the early events remains unclear. Through analysis of single-cell RNA sequencing data via iterative random forest leave one out prediction, an explainable artificial intelligence method, we identified an immunoregulatory role for a unique paired related homeobox-1 (Prx1)+ fibroblast subpopulation. Disruption of Ikkb-NF-κB under homeostatic conditions in these fibroblasts paradoxically induced skin inflammation due to the overexpression of C-C motif chemokine ligand 11 (CCL11; or eotaxin-1) characterized by eosinophil infiltration and a subsequent TH2 immune response. Because the inflammatory phenotype resembled that seen in human atopic dermatitis (AD), we examined human AD skin samples and found that human AD fibroblasts also overexpressed CCL11 and that perturbation of Ikkb-NF-κB in primary human dermal fibroblasts up-regulated CCL11. Monoclonal antibody treatment against CCL11 was effective in reducing the eosinophilia and TH2 inflammation in a mouse model. Together, the murine model and human AD specimens point to dysregulated Prx1+ fibroblasts as a previously unrecognized etiologic factor that may contribute to the pathogenesis of AD and suggest that targeting CCL11 may be a way to treat AD-like skin lesions.


Assuntos
Dermatite Atópica , Animais , Inteligência Artificial , Dermatite Atópica/patologia , Fibroblastos/patologia , Imunidade , Camundongos , NF-kappa B/metabolismo , Pele/patologia
6.
Sci Adv ; 7(50): eabj9141, 2021 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-34890228

RESUMO

The epigenetic regulator, MLL4 (KMT2D), has been described as an essential gene in both humans and mice. In addition, it is one of the most commonly mutated genes in all of cancer biology. Here, we identify a critical role for Mll4 in the promotion of epidermal differentiation and ferroptosis, a key mechanism of tumor suppression. Mice lacking epidermal Mll4, but not the related enzyme Mll3 (Kmt2c), display features of impaired differentiation and human precancerous neoplasms, all of which progress with age. Mll4 deficiency profoundly alters epidermal gene expression and uniquely rewires the expression of key genes and markers of ferroptosis (Alox12, Alox12b, and Aloxe3). Beyond revealing a new mechanistic basis for Mll4-mediated tumor suppression, our data uncover a potentially much broader and general role for ferroptosis in the process of differentiation and skin homeostasis.

7.
Cell Stem Cell ; 28(9): 1582-1596.e6, 2021 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-34102139

RESUMO

Stem cells support lifelong maintenance of adult organs, but their specific roles during injury are poorly understood. Here we demonstrate that Lgr6 marks a regionally restricted population of epidermal stem cells that interact with nerves and specialize in wound re-epithelialization. Diphtheria toxin-mediated ablation of Lgr6 stem cells delays wound healing, and skin denervation phenocopies this effect. Using intravital imaging to capture stem cell dynamics after injury, we show that wound re-epithelialization by Lgr6 stem cells is diminished following loss of nerves. This induces recruitment of other stem cell populations, including hair follicle stem cells, which partially compensate to mediate wound closure. Single-cell lineage tracing and gene expression analysis reveal that the fate of Lgr6 stem cells is shifted toward differentiation following loss of their niche. We conclude that Lgr6 epidermal stem cells are primed for injury response and interact with nerves to regulate their fate.


Assuntos
Reepitelização , Receptores Acoplados a Proteínas G , Células Epidérmicas , Folículo Piloso , Células-Tronco
8.
Acad Radiol ; 28(6): 871-876, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-32828663

RESUMO

RATIONALE AND OBJECTIVES: Three-dimensional (3D) visualization has been shown to benefit new generations of medical students and physicians-in-training in a variety of contexts. However, there is limited research directly comparing student performance after using 3D tools to those using two-dimensional (2D) screens. MATERIALS AND METHODS: A CT was performed on a donated cadaver and a 3D CT hologram was created. A total of 30 first-year medical students were randomly assigned into two groups to review head and neck anatomy in a teaching session that incorporated CT. The first group used an augmented reality headset, while the second group used a laptop screen. The students were administered a five-question anatomy test before and after the session. Two-tailed t-tests were used for statistical comparison of pretest and posttest performance within and between groups. A feedback survey was distributed for qualitative data. RESULTS: Pretest vs. posttest comparison of average percentage of questions answered correctly demonstrated both groups showing significant in-group improvement (p < 0.05), from 59% to 95% in the augmented reality group, and from 57% to 80% in the screen group. Between-group analysis indicated that posttest performance was significantly better in the augmented reality group (p = 0.022, effect size = 0.73). CONCLUSION: Immersive 3D visualization has the potential to improve short-term anatomic recall in the head and neck compared to traditional 2D screen-based review, as well as engage millennial learners to learn better in anatomy laboratory. Our findings may reflect additional benefit gained from the stereoscopic depth cues present in augmented reality-based visualization.


Assuntos
Anatomia , Realidade Aumentada , Estudantes de Medicina , Cabeça/diagnóstico por imagem , Humanos , Imageamento Tridimensional , Pescoço/diagnóstico por imagem , Tomografia Computadorizada por Raios X
9.
J Invest Dermatol ; 141(2): 295-307.e13, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32649944

RESUMO

Squamous cell carcinoma in situ (SCCIS) is a prevalent precancerous lesion that can progress to cutaneous squamous cell carcinoma. Although SCCIS is common, its pathogenesis remains poorly understood. To better understand SCCIS development, we performed laser captured microdissection of human SCCIS and the adjacent epidermis to isolate genomic DNA and RNA for next-generation sequencing. Whole-exome sequencing identified UV-signature mutations in multiple genes, including NOTCH1-3 in the epidermis and SCCIS and oncogenic TP53 mutations in SCCIS. Gene families, including SLFN genes, contained UV/oxidative-signature disruptive epidermal mutations that manifested positive selection in SCCIS. The frequency and distribution of NOTCH and TP53 mutations indicate that NOTCH mutations may precede TP53 mutations. RNA sequencing identified 1,166 differentially expressed genes; the top five enriched gene ontology biological processes included (i) immune response, (ii) epidermal development, (iii) protein phosphorylation, (iv) regulation of catalytic activity, and (v) cytoskeletal regulation. The NEURL1 ubiquitin ligase, which targets Notch ligands for degradation, was upregulated in SCCIS. NEURL1 protein was found to be elevated in SCCIS suggesting that increased levels could represent a mechanism for downregulating Notch during UV-induced carcinogenesis. The data from DNA and RNA sequencing of epidermis and SCCIS provide insights regarding SCCIS formation.


Assuntos
Carcinoma in Situ/etiologia , Carcinoma de Células Escamosas/etiologia , Epiderme/efeitos da radiação , Exoma , Perfilação da Expressão Gênica , Neoplasias Induzidas por Radiação/etiologia , Neoplasias Cutâneas/etiologia , Carcinogênese/genética , Carcinoma in Situ/genética , Carcinoma de Células Escamosas/genética , Genes p53 , Humanos , Mutação , Neoplasias Induzidas por Radiação/genética , Receptores Notch/genética , Análise de Sequência de RNA , Neoplasias Cutâneas/genética , Raios Ultravioleta
10.
J Investig Dermatol Symp Proc ; 19(2): S103-S105, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30471750

RESUMO

Cutaneous squamous cell carcinoma (cSCC) is the second most common cancer in humans, with an incidence of approximately 700,000 cases per year in the United States (Rogers et al., 2010). It is known that cSCC is strongly associated with sun exposure, specifically UVB and UVA, as well as other risk factors, such as human papillomavirus infection, immunodeficiency, and specific medications (Ratushny et al., 2012). However, the precise sequence of biological events leading to tumor development remains unknown. With projected higher incidence of patients with cSCCs in the future, there is a strong need to elucidate the molecular pathways that regulate formation of cSCCs.

11.
Cell Cycle ; 13(10): 1551-9, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24626198

RESUMO

Fibroblast growth factor receptor 3 (FGFR3) activating mutations are drivers of malignancy in several human tissues, including bladder, lung, cervix, and blood. However, in skin, these mutations are associated predominantly with benign, common epidermal growths called seborrheic keratoses (SKs). How epidermis resists FGFR3 mediated transformation is unclear, but previous studies have suggested that FGFR3 activation in skin keratinocytes may serve a tumor-suppressive role by driving differentiation and antagonizing Ras signaling. To define the role of FGFR3 in human normal and neoplastic epidermis, and to directly test the hypothesis that FGFR3 antagonizes Ras, we engineered human skin grafts in vivo with mutant active FGFR3 or shRNA FGFR3 knockdown. We show that FGFR3 active mutants drive mild hyperproliferation, but are insufficient to support benign or malignant tumorigenesis, either alone, or in combination with G 1-S checkpoint release. This suggests that additional cell-intrinsic or stromal cues are required for formation of benign SKs with FGFR3 mutations. Further, FGFR3 activation does not alter the growth kinetics or differentiation status of engineered human epidermal SCCs driven by Ras, and FGFR3 protein itself is dispensable for Ras-driven SCC. To extend these findings to patients, we examined a uniquely informative human tumor in which SCC developed in continuity with a SK, raising the hypothesis that one of the tumors evolved from the other. However, mutational analysis from each tumor indicates that the overlapping SK and SCC evolved independently and supports our conclusion that FGFR3 activation is insufficient to drive SCC.


Assuntos
Carcinoma de Células Escamosas/patologia , Epiderme/patologia , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Neoplasias Cutâneas/patologia , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Células Cultivadas , Aberrações Cromossômicas , Epiderme/metabolismo , Xenoenxertos , Humanos , Hiperplasia , Recém-Nascido , Queratinócitos/metabolismo , Ceratose Seborreica/genética , Ceratose Seborreica/metabolismo , Ceratose Seborreica/patologia , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteína Oncogênica p21(ras)/metabolismo , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/metabolismo , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo
12.
Skin Res Technol ; 17(4): 387-97, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21492240

RESUMO

BACKGROUND: In vivo confocal scanning laser microscopy (CSLM) is a recently developed non-invasive technique for visualizing microscopic structures with the skin. CSLM has been used to characterize proliferative and inflammatory skin diseases, neoplastic skin lesions and pigmented lesions. OBJECTIVE: Here, we assessed the ability of CSLM to evaluate the formation of neogenic hair follicles after a full-thickness wound in mice. METHODS: Full-thickness wounds were made on the dorsal skin of 3-week-old mice. After scab detachment (SD), the number, width, length, space and volume of neogenic hair follicles were analyzed using CSLM. The results were compared with those from conventional methods, including staining for alkaline phosphatase (AP) and keratin 17 (K17) as well as histology. RESULTS: Quantification of neogenic hair follicles using CSLM compared favorably with the results from direct measurements on isolated epidermal tissue after immunostaining for K17, a marker for the epithelial portion of new hair follicles. CSLM detected 89% of K17-stained follicles. CSLM more accurately quantified the number of new follicles compared with AP staining, which detects the dermal portion of the new follicle. The width and length measurement from CSLM and histology were very close and correlated with each other. The minimum length of a neogenic hair follicle that could be detected by CSLM was 21 µm. The space between neogenic hair follicles was decreased in histological sections compared with CSLM. CONCLUSION: CSLM is an accurate and valuable method for counting and measuring neogenic hair follicles non-invasively. CSLM produces images similar to histology in mice. Measurements of microstructures using CSLM more accurately reflect actual sizes as this technique avoids fixation artifacts. In vivo visualization of developing follicles with CSLM allows the detection of serial changes in hair follicle formation, thus conserving the numbers of mice required for studies and improving the detection of temporal changes in developing hair follicles.


Assuntos
Folículo Piloso/lesões , Folículo Piloso/fisiologia , Microscopia Confocal/métodos , Regeneração/fisiologia , Cicatrização/fisiologia , Fosfatase Alcalina/metabolismo , Animais , Biomarcadores/metabolismo , Divisão Celular/fisiologia , Sobrevivência Celular/fisiologia , Derme/citologia , Derme/lesões , Derme/fisiologia , Células Epidérmicas , Epiderme/lesões , Epiderme/fisiologia , Folículo Piloso/citologia , Queratinas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL
13.
J Med Chem ; 49(11): 3402-11, 2006 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-16722660

RESUMO

A novel series of pyridazinone-functionalized phenylalanine analogues was prepared and evaluated for inhibition of cellular adhesion mediated by alpha4beta1/VCAM-1 and alpha4beta7/MAdCAM-1 interactions. Concise syntheses were developed and applied for exploration of structure-activity relationships pertaining to the pyridazinone ring as well as the N-acyl phenylalanine scaffold. Potent dual antagonists of alpha4beta1 and alpha4beta7 were generated from an amide subseries; antagonists selective for alpha4beta7 were identified from urea and carbamate-based subseries. The pharmacokinetic properties of selected members of the series have been determined in rats and demonstrate that the use of ester prodrugs and alterations to the amide linkage can lead to improved oral bioavailability in this series. An alpha4beta7-selective member of the carbamate subseries (36c), upon oral administration, demonstrated in vivo efficacy in the mouse DSS colitis model.


Assuntos
Integrina alfa4beta1/antagonistas & inibidores , Integrinas/antagonistas & inibidores , Fenilalanina/análogos & derivados , Piridazinas/síntese química , Animais , Disponibilidade Biológica , Adesão Celular/efeitos dos fármacos , Moléculas de Adesão Celular , Colite/induzido quimicamente , Colite/tratamento farmacológico , Sulfato de Dextrana , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/fisiologia , Ésteres/síntese química , Ésteres/química , Ésteres/farmacologia , Granulócitos/efeitos dos fármacos , Granulócitos/fisiologia , Humanos , Imunoglobulinas/metabolismo , Técnicas In Vitro , Integrina alfa4beta1/metabolismo , Integrinas/metabolismo , Células K562 , Linfócitos/efeitos dos fármacos , Linfócitos/fisiologia , Camundongos , Monócitos/efeitos dos fármacos , Monócitos/fisiologia , Mucoproteínas/metabolismo , Fenilalanina/síntese química , Fenilalanina/química , Fenilalanina/farmacologia , Pró-Fármacos/síntese química , Pró-Fármacos/química , Pró-Fármacos/farmacologia , Piridazinas/química , Piridazinas/farmacologia , Ratos , Relação Estrutura-Atividade , Veias Umbilicais/citologia , Molécula 1 de Adesão de Célula Vascular/metabolismo
14.
Bioorg Med Chem ; 13(24): 6693-702, 2005 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-16112583

RESUMO

A series of N-carboxy, N-alkyl, and N-carboxamido azabicyclo[2.2.2]octane carboxamides were prepared and assayed for inhibition of alpha4beta1-VCAM-1 and alpha4beta7-MAdCAM-1 interactions. Potency and alpha4beta1/alpha4beta7 selectivity were sensitive to the substituent R1-R3 in the structures 6, 7, and 8. Several compounds demonstrated low nanomolar balanced alpha4beta1/alpha4beta7 in vitro activity. Two compounds were selected for in vivo leukocytosis studies and demonstrated increases in circulating lymphocytes up to 250% over control.


Assuntos
Aminoácidos Cíclicos/química , Compostos Aza/síntese química , Compostos Aza/farmacologia , Integrina alfa4beta1/antagonistas & inibidores , Integrinas/antagonistas & inibidores , Animais , Compostos Aza/química , Feminino , Integrina alfa4beta1/metabolismo , Integrinas/metabolismo , Leucocitose , Linfócitos/efeitos dos fármacos , Masculino , Camundongos , Estrutura Molecular , Ratos , Relação Estrutura-Atividade
15.
Biochem Biophys Res Commun ; 327(1): 64-9, 2005 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-15629430

RESUMO

Plucked (pk) is an autosomal recessive mouse mutation with a hair phenotype that arose spontaneously in the DBA/2J strain. Histological studies indicate that adult pk mutant mice lose truncal hair because of the scarring of follicles due to an apparent obstruction of the outward movement of the hair shaft within the follicular canal. We mapped the pk mutant phenotype to a 1.1cM region of chromosome 18 (between 6.6 and 7.7 cM from the centromere) using 370 backcross progeny. Within this region, among others, are genes for desmosome cadherins. Desmosome cadherins are interesting candidates because of their critical roles for cell-cell adhesion in epidermal function. Northern Blot analysis of wild-type and pk mutant mice indicates that expression of both desmoglein 1 (Dsg1) and desmoglein 3 (Dsg3) is up-regulated in the skin of mutant pk mice.


Assuntos
Caderinas/genética , Proteínas do Citoesqueleto/genética , Mutação/genética , Regulação para Cima/genética , Animais , Mapeamento Cromossômico , Desmogleína 1 , Desmogleína 3 , Desmogleínas , Desmoplaquinas , Feminino , Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Folículo Piloso/metabolismo , Humanos , Masculino , Camundongos , Camundongos Mutantes , Morfogênese , Fenótipo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Pele/metabolismo
16.
Br J Pharmacol ; 141(8): 1335-45, 2004 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15100166

RESUMO

From a historical perspective to the present day, all the evidence suggests that activation of cannabinoid receptors (CBRs) is beneficial for gut discomfort and pain, which are symptoms related to dysmotility and visceral perception. CBRs comprise G-protein coupled receptors that are predominantly in enteric and central neurones (CB1R) and immune cells (CB2R). In the last decade, evidence obtained from the use of selective agonists and inverse agonists/antagonists indicates that manipulation of CB1R can alter (1) sensory processing from the gut, (2) brain integration of brain-gut axis, (3) extrinsic control of the gut and (4) intrinsic control by the enteric nervous system. The extent to which activation of CB1R is most critical at these different levels is related to the region of the GI tract. The upper GI tract is strongly influenced by CB1R activation on central vagal pathways, whereas intestinal peristalsis can be modified by CB1R activation in the absence of extrinsic input. Actions at multiple levels make the CB1R a target for the treatment of functional bowel disorders, such as IBS. Since low-grade inflammation may act as a trigger for occurrence of IBS, CB2R modulation could be beneficial, but there is little supporting evidence for this yet. The challenge is to accomplish CBR activation while minimizing adverse effects and abuse liabilities. Potential therapeutic strategies involve increasing signaling by endocannabinoids (EC). The pathways involved in the biosynthesis, uptake and degradation of EC provide opportunities for modulation of CB1R and some recent evidence with inhibitors of EC uptake and metabolism suggest that these could be exploited for therapeutic gain.


Assuntos
Motilidade Gastrointestinal/fisiologia , Limiar da Dor/fisiologia , Receptores de Canabinoides/fisiologia , Fibras Aferentes Viscerais/fisiologia , Animais , Agonistas de Receptores de Canabinoides , Moduladores de Receptores de Canabinoides/farmacologia , Moduladores de Receptores de Canabinoides/uso terapêutico , Humanos , Limiar da Dor/efeitos dos fármacos , Fibras Aferentes Viscerais/efeitos dos fármacos , Fibras Aferentes Viscerais/metabolismo
17.
Bioorg Med Chem Lett ; 14(3): 591-6, 2004 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-14741249

RESUMO

The design, synthesis, and biological activity of novel alpha(4)beta(1) and alpha(4)beta(7) integrin antagonists, containing a bridged azabicyclic nucleus, are reported. Conformational analysis of targets containing an azabicyclo[2.2.2]octane carboxylic acid and known integrin antagonists indicated that this azabicycle would be a suitable molecular scaffold. Variation of substituents on the pendant arylsulfonamide and phenylalanine groups resulted in potent alpha(4)beta(1)-selective and dual alpha(4)beta(1)/alpha(4)beta(7) antagonists. Potent compounds 11i, 11h, and 14 were effective in the antigen-sensitized sheep model of asthma.


Assuntos
Antígenos de Helmintos/imunologia , Compostos Aza/farmacologia , Integrina alfa4beta1/antagonistas & inibidores , Integrinas/antagonistas & inibidores , Sulfonamidas/farmacologia , Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Ascaris/imunologia , Asma/tratamento farmacológico , Asma/patologia , Compostos Aza/síntese química , Compostos Aza/química , Brônquios/imunologia , Brônquios/fisiologia , Modelos Animais de Doenças , Hipersensibilidade/imunologia , Hipersensibilidade/patologia , Conformação Molecular , Estrutura Molecular , Fenilalanina/química , Ovinos , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química
18.
J Invest Dermatol ; 120(5): 707-14, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12713571

RESUMO

Delta-6 desaturase, also known as fatty acid desaturase-2 (FADS2), is a component of a lipid metabolic pathway that converts the essential fatty acids linoleate and alpha-linolenate into long-chain polyunsaturated fatty acids. Isolation of Delta-6 desaturase/FADS2 cDNA from human skin predicts an identical protein to that expressed in human brain and Southern analysis indicates a single locus, together suggestive of a single Delta-6 desaturase/FADS2 gene. Within human skin, Delta-6 desaturase/FADS2 mRNA and protein expression is restricted to differentiating sebocytes located in the suprabasal layers of the sebaceous gland. Enzymatic analysis using CHO cells overexpressing human Delta-6 desaturase/FADS2 indicates catalysis of a "polyunsaturated fatty acid type" reaction, but also an unexpected "sebaceous-type" reaction, that of converting palmitate into the mono-unsaturated fatty acid sapienate, a 16-carbon fatty acid with a single cis double bond at the sixth carbon from the carboxyl end. Sapienate is the most abundant fatty acid in human sebum, and among hair-bearing animals is restricted to humans. This work identifies Delta-6 desaturase/FADS2 as the major fatty acid desaturase in human sebaceous glands and suggests that the environment of the sebaceous gland permits catalysis of the sebaceous-type reaction and restricts catalysis of the polyunsaturated fatty acid type reaction.


Assuntos
Ácidos Graxos Dessaturases/química , Glândulas Sebáceas/enzimologia , Animais , Northern Blotting , Southern Blotting , Encéfalo/metabolismo , Células CHO , Clonagem Molecular , Cricetinae , DNA Complementar/metabolismo , Ácidos Graxos Dessaturases/biossíntese , Ácidos Graxos Dessaturases/genética , Ácidos Graxos Insaturados/metabolismo , Vetores Genéticos , Humanos , Imuno-Histoquímica , Hibridização In Situ , Linoleoil-CoA Desaturase , Metabolismo dos Lipídeos , Modelos Químicos , Fases de Leitura Aberta , Reação em Cadeia da Polimerase , RNA Mensageiro/metabolismo , Sebo/metabolismo , Análise de Sequência de DNA , Pele/metabolismo , Distribuição Tecidual
19.
J Cutan Pathol ; 29(5): 268-78, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-12100626

RESUMO

BACKGROUND: An animal model for the study of basal cell carcinoma (BCC) is required to better understand its biology. Several attempts to grow BCC in immuno-incompetent animals have been only modestly successful. METHODS: To test the ability of BCC to grow in a mouse with complete and severe immuno-incompetence, 14 individual BCC were transplanted into the subcutaneous tissue of 18 SCID-beige mice (T, B and natural killer cell deficient). Light microscopy and immunophenotypic analyses were performed on primary BCC and first and seventh passage tumors. RESULTS: Transplantation of three BCC yielded rapidly growing anaplastic tumors for a tumor take of 18% (3/18). SCID-beige mice without tumor growth had mostly scars or epidermoid cysts at the transplant sites. The three patients whose BCC gave rise to the anaplastic tumors were significantly older than those without tumor growth (87 vs. 64, p = 0.001), but they did not differ with respect to BCC type or general health. These three anaplastic tumors were histologically and immunophenotypically similar, being composed of dyscohesive, pleomorphic cells that expressed vimentin and smooth muscle actin. In the first passage mice these tumors were locally invasive, tumor-forming nodules associated with an expansion of donor inflammatory cells (T and B lymphocytes and plasma cells), rare remnants of BCC epithelium and epidermoid cysts. By the seventh passage, the tumors were homogenous and metastasized widely throughout the mice. Changing transplantation location to the dermis to wound environment or supplementing the tumor with BCC-derived fibroblasts did not alter the phenotype or growth rate in SCID-beige mice. Anaplastic tumors also grew easily in SCID mice (T and B cell deficient). However, transplantation of the anaplastic tumors into normal mice (CB-17) or less severely immunodeficient mice (NCr and Balb/c: T and natural killer cell deficient) did not allow for growth. Furthermore, tumor growth could not be maintained in vitro. CONCLUSION: Empirically, these data suggest that BCC has the potential to become an aggressive metastatic neoplasm, given the right immune and stromal environment. Moreover, a functional B lymphocyte system appears to prevent this growth. As human lymphocytes also engraft in SCID-beige mice, the original host immune response could be responsible for the lack of tumor growth in the majority of xenografts. Furthermore, the anaplastic and metastatic phenotype of these BCC derived neoplasms may be the experimental equivalent of metastatic BCC and BCC associated with carcinosarcoma.


Assuntos
Carcinoma Basocelular/metabolismo , Carcinoma Basocelular/patologia , Modelos Animais , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Anaplasia/patologia , Animais , Carcinoma Basocelular/genética , Carcinoma Basocelular/imunologia , Feminino , Humanos , Masculino , Camundongos , Camundongos SCID , Transplante de Neoplasias , Pele/patologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Transplante Heterólogo
20.
J Biol Chem ; 277(1): 59-66, 2002 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-11677241

RESUMO

The regulation of stearoyl-CoA desaturase (SCD), a rate-limiting enzyme in the synthesis of unsaturated fatty acids, is physiologically important because the ratio of saturated to unsaturated fatty acids is thought to control cellular functions by modulating the structural integrity and fluidity of cell membranes. Transforming growth factor-beta (TGF-beta), a multifunctional cytokine, increased SCD mRNA expression in cultured human retinal pigment epithelial cells. This response was elicited by all three TGF-beta isoforms, beta1, beta2, and beta3. However, SCD mRNA expression was not increased either by other members of the TGF-beta family or by other growth factors or cytokines. TGF-beta also increased SCD mRNA expression in several other cell lines tested. The increase in SCD mRNA expression was preceded by a marked increase in Smad2 phosphorylation in TGF-beta-treated human retinal pigment epithelial cells. TGF-beta did not induce SCD mRNA expression in a Smad4-deficient cell line. However, Smad4 overexpression restored the TGF-beta effect in this cell line. Moreover, TGF-beta-induced SCD mRNA expression was effectively blocked by the overexpression of Smad7, an inhibitory Smad. Thus, a TGF-beta signal transduction pathway involving Smad proteins appears to regulate the cellular expression of the SCD gene, and this regulation may play an important role in lipid metabolism.


Assuntos
Proteínas de Ligação a DNA/fisiologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Estearoil-CoA Dessaturase/genética , Transativadores/fisiologia , Fator de Crescimento Transformador beta/farmacologia , Células Cultivadas , Dactinomicina/farmacologia , Humanos , Fosforilação , Epitélio Pigmentado Ocular/enzimologia , Proteína Smad2 , Proteína Smad4 , Proteína Smad7
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