Biomedical subjects

Yanina Malykh

Publications and source records attributed to Yanina Malykh.

4 recordsLinked to original sources

PubMed2006

Analyzing proteome topology and function by automated multidimensional fluorescence microscopy.

Temporal and spatial regulation of proteins contributes to function. We describe a multidimensional microscopic robot technology for high-throughput protein colocalization studies that runs cycles of fluorescence tagging, imaging and bleaching in situ. This technology combines three advances: a fluorescence technique capable of mapping hundreds of different proteins in one tissue section or cell sample; a method selecting the most prominent combinatorial molecular patterns by representing the data as binary vectors; and a system for imaging the distribution of these protein clusters in a so-called toponome map. By analyzing many cell and tissue types, we show that this approach reveals rules of hierarchical protein network organization, in which the frequency distribution of different protein clusters obeys Zipf's law, and state-specific lead proteins appear to control protein network topology and function. The technology may facilitate the development of diagnostics and targeted therapies.

Dermatitis, Atopic↗

PubMed2003

Reduction of CMP-N-acetylneuraminic acid hydroxylase activity in engineered Chinese hamster ovary cells using an antisense-RNA strategy.

Rodent cells, widely used for the industrial production of recombinant human glycoproteins, possess CMP-N-acetylneuraminic acid hydroxylase (CMP-Neu5Ac hydroxylase; EC 1.14.13.45) which is the key enzyme in the formation of the sialic acid, N-glycolylneuraminic acid (Neu5Gc). This enzyme is not expressed in an active form in man and evidence suggests that the presence of Neu5Gc in recombinant therapeutic glycoproteins may elicit an immune response. The aim of this work was, therefore, to reduce CMP-Neu5Ac hydroxylase activity in a Chinese Hamster Ovary (CHO) cell line, and thus the Neu5Gc content of the resulting glycoconjugates, using a rational antisense RNA approach. For this purpose, the cDNA of the hamster hydroxylase was partially cloned and sequenced. Based on the sequence of the mouse and hamster cDNAs, optimal antisense RNA fragments were selected from preliminary in vitro translation tests. Compared to the parental cell line, the new strain (CHO-AsUH2), which was transfected with a 199-bp antisense fragment derived from the mouse CMP-Neu5Ac hydroxylase cDNA, showed an 80% reduction in hydroxylase activity. An analysis of the sialic acids present in the cells' own glycoconjugates revealed a decrease in the percentage of Neu5Gc residues from 4% in the parental cells to less than 1% in the CHO-AsUH2 cell line.

Animals↗

PubMed2002

Biological properties of N-acyl and N-haloacetyl neuraminic acids: processing by enzymes of sialic acid metabolism, and interaction with influenza virus.

Several unnatural N-acyl neuraminic acids (N-propionyl, N-hexanoyl, N-benzoyl, N-trifluoroacetyl, N-chloroacetyl, N-difluoroacetyl) were prepared enzymatically using immobilised sialic acid aldolase. N-Trifluoroacetyl-, N-chloroacetyl- and N-difluoroacetyl neuraminic acids were shown to enhance up to 10-fold the rate of association of influenza virus A to a sialoglycolipid neomembrane by surface plasmon resonance, and were found to act as weak inhibitors (K(iapp) 0.45-2.0 mM) of influenza virus neuraminidase. The N-propionyl, N-chloroacetyl- and N-difluoroacetyl neuraminic acids were found to be substrates for recombinant Escherichia coli CMP sialate synthase, to give the corresponding CMP-N-acyl-neuraminic acids. CMP-N-propionyl neuraminic acid was found not to be a substrate for CMP-N-acetyl neuraminic acid hydroxylase from pig submandibular gland.

Animals↗

PubMedDate not supplied

Profiling lymphocyte subpopulations in peripheral blood under efalizumab treatment of psoriasis by multi epitope ligand cartography (MELC) robot microscopy.

CD11a-blocking efalizumab has recently been approved as a systemic treatment of moderate to severe chronic plaque psoriasis. When treating 6 psoriasis patients with efalizumab over 12 weeks in the present study, we observed an overall good tolerability and 5 treatment responders characterized by a decrease of PASI from 21.3 +/- 5.4 to 3.9 +/- 0.6. The accompanying significant increase of peripheral blood lymphocytes from 1.9 +/- 0.7 to 4.3 +/- 1.0 x 10(9)/L (p < 0.05) was analyzed by multi epitope ligand cartography (MELC) robot microscopy. Thereby a high-dimension simultaneous multiplex immunophenotyping was pursued using 39 fluorophore-labeled antibodies including labeled efalizumab and 3 other affinity reagents such as lectins. Due to efalizumab treatment there was a substantial decrease of the cellular expression of CD11a (detected by mab clone 25.3.1) and efalizumab binding sites (EfaBSs). This was paralleled by an increase of the number of EfaBS- and EfaBS+ lymphocytes by a factor of 2.4x and 2.2x, respectively. The latter effect was mainly derived from a subpopulation showing a low degree of EfaBS expression. Efalizumab treatment led furthermore to an increase of the numbers of CD3+, CD4+, CD8+, CD44+, CD45+, CD45R0+, CD45 RA+, CD52+, CD58+, CD247+, HLA-DR+ and Sambucus nigra lectin-reactive lymphocytes (by factors from 2.0 to 3.3x). In terms of a combinatorial molecular phenotype we identified a CD3+/CD4+/CD44+/CD52+ lymphocyte subpopulation which accumulated most predominantly from 0.824 +/- 0.270 x 10(9)/L up to 1.616 +/- 0.152 x 10(9)/L under efalizumab treatment (p < 0.01). Thus, the current study extends the knowledge of efalizumab-dependent perturbations of recirculating blood lymphocyte subpopulations in psoriasis patients.

Adult↗