Interleukin-1 Receptor antagonist Production by Human Keratinocytes

Human keratinocytes produce biologically active pro–IL-Iα and inactive pro-IL-1β with most protein remaining intracellular. IL-1 receptor antagonist (IL-1ra) is a newly described member of the IL-1 family that is secreted by stimulated monocytes and binds competitively to IL-1 receptors without stimulating target cells. We examined the characteristics of IL-1ra production by cultured human keratinocytes. By ELISA, keratinocyte lysates contained 390 ng IL-1ra/mg total protein with little IL-1ra detected in supernatants. In contrast, monocytes produced 297 ng IL- 1ra/mg total protein during 24 h of culture on adherent IgG with about half of the IL-1ra detected in supernatants. By Western blot analysis, keratinocyte IL-1ra was ≈ 20 kD in size and was slightly larger than recombinant monocyte IL- 1ra. In contrast to monocytes, human keratinocyte IL-1ra was not secreted in 22- 25-kD molecular weight glycosylated forms. Affinity-purified keratinocyte IL-1ra exhibited identical biologic activity to recombinant monocyte IL-1ra, each inhibiting IL-1 -dependent augmentation of murint thymocyte proliferation to the same degree per amount protein. An IL-1ra mRNA of 1.8 kb was detected by Northern blot analysis in RNA extracted from keratinocytes. in order to determine the effect of differentiation on IL-1 and IL-1ra production, human keratinocytes were cultured for 72 h in low (0.03 mM), medium (0.15 mM), or high (1.0 mM) -calcium concentrations. The absolute amounts of IL-1ra increased twofold and the ratio of IL-1ra to IL-1ra in keratinocyte lysates increased from ≈ 12: 1 to 25: 1 during differentiation. These results indicate that keratinocytes constitutively produce large amounts of a biologically active intracellular variant of IL-1ra that increase with differentiation. IL-1ra released during keratinocyte damage may be important in modifying the inflammatory effects of IL-1α in human skin

Enhancement of Specific Immunofluorescent Finding with Use of a Para-Phenylenediamine Mounting Buffer

A recently described immunofluorescence mounting buffer containing para-phenylenediamine prevents fading of specific staining in skin sections during microscopic examination, and allows better appreciation of morphological detail. Examination of slides at high powers with intense illumination, as well as improved photomicrographs, are possible with this reagent

Serum-Free Serial Culture of Adult Human Keratinocytes From Suction-Blister Roof Epidermis

Coating cell culture flasks with natural extracellular matrix (ECM) enhanced the culture of adult human keratinocytes from suction-blister roof epidermis in an environment without fetal calf serum (FCS), bovine pituitary extracts or cellular feeder layers. A higher incidence of cell attachment on natural ECM was observed than on collagen and human fibronectins(HFN)-coated plastic dishes, and natural ECM was necessary for growth and proliferation of attached cells under the culture conditions used. Cells in primary culture grew to confluency on natural ECM-coated surfaces within about 14 days, and subsequent serial passage could be made up to fourth passage in collagen- and HFN-coated plastic flasks. Cultured keratinocytes in this serum-free environment formed colonies of small cuboidal, healthy cells with little keratinization or stratification and demonstrated antigenic characteristics of human basal cells

Solstice: An Electronic Journal of Geography and Mathematics: Vol. 31, No. 2

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/163731/1/SolsticeVolumeXXXINumber2.pdfd0a18e86-7d9e-4669-812b-ead353cc4899b2c2c9fd-ee9d-4e83-89fe-dc520ef84368Description of SolsticeVolumeXXXINumber2.pdf : Solstice, Vol. XXXI, No. 2.SEL

Human Monocyte Chemotaxis: A Quantitative in Vivo Technique

This report describes a new quantitative technique for evaluating monocyte chemotaxis to a site of superficial epidermal abrasion. Micro-acrylic chambers containing 50% Zymosan activated autologous serum were separated from a 5-mm diameter epidermal abrasion by 2 Nucleopore filters which entrapped migrating monocytes but allowed free neutrophil migration. Monocytes were specifically identified by alpha napthyl acetate esterase activity. Monocytes accumulated within the filters by 4 hr and maximized at 16 and 20 hr.This technique is superior to previous skin chamber techniques in the high yield of monocytes and in specific histochemical identification of monocytes. In contrast to the Rebuck window, it does not generate attractants and has greater reproducibility.This technique will be useful in the study of diseases characterized by monocytic infiltrates, in contrasting the function of peripheral blood monocytes to those available in the skin, and in testing the effects of drugs, immunodeficiency and infection on monocyte function in vivo

Advancing Academic Pathways for Building Capacity in the Youth Development Profession

The growing literature on the theory and practice of youth development provides a framework for youth-serving practitioners to design, implement, and grow youth programs in their communities. Yet such a framework is not widely known in many youth-serving organizations where professional development is variable. The youth development field would benefit from academically trained professionals who could apply the youth development literature to serve youth more effectively in organizations or communities. This article describes a graduate level academic degree program in youth development to: (a) increase awareness of the importance of youth programs and (b) bring attention to the fundamental learning structures that can be deployed to build human capacity in the youth development field. The history, theoretical frameworks, and structure of the program are briefly described. Details are provided on 4 effective features of this academic degree program rated by students and graduates as being valuable components of the program. Applications to youth development practice are explored

Detecting Remote Evolutionary Relationships among Proteins by Large-Scale Semantic Embedding

Virtually every molecular biologist has searched a protein or DNA sequence database to find sequences that are evolutionarily related to a given query. Pairwise sequence comparison methods—i.e., measures of similarity between query and target sequences—provide the engine for sequence database search and have been the subject of 30 years of computational research. For the difficult problem of detecting remote evolutionary relationships between protein sequences, the most successful pairwise comparison methods involve building local models (e.g., profile hidden Markov models) of protein sequences. However, recent work in massive data domains like web search and natural language processing demonstrate the advantage of exploiting the global structure of the data space. Motivated by this work, we present a large-scale algorithm called ProtEmbed, which learns an embedding of protein sequences into a low-dimensional “semantic space.” Evolutionarily related proteins are embedded in close proximity, and additional pieces of evidence, such as 3D structural similarity or class labels, can be incorporated into the learning process. We find that ProtEmbed achieves superior accuracy to widely used pairwise sequence methods like PSI-BLAST and HHSearch for remote homology detection; it also outperforms our previous RankProp algorithm, which incorporates global structure in the form of a protein similarity network. Finally, the ProtEmbed embedding space can be visualized, both at the global level and local to a given query, yielding intuition about the structure of protein sequence space