Impairments in prospective and retrospective memory following stroke.

Kim, Hyun Jung,Craik, Fergus I M,Luo, Lin,Ween, Jon Erik Oxford University Press 2009 Neurocase Vol.15 No.2

<P>Prospective memory (PM) is the ability to carry out a planned intention at a future time. We studied PM deficits in a group of community-dwelling stroke survivors compared with normal controls. Twelve stroke patients and 12 matched controls performed a series of tests assessing executive function, prospective (PM) and retrospective memory (RM). Patients performed less well than controls on laboratory measures of PM and associative RM; they also showed deficits on standard tests of RM and executive control. The groups did not differ on more structured clinical measures of executive function, RM, PM or self-rated PM and RM. The results are discussed in terms of an impairment in the ability to 'self-initiate' effortful cognitive processes.</P>

Anti-proliferative Activities of Metallic Nanoparticles in an in Vitro Breast Cancer Model

Loutfy, Samah A,Al-Ansary, Nadia A,Abdel-Ghani, Nour T,Hamed, Ahmed R,Mohamed, Mona B,Craik, James D,Eldin, Taher A. Salah,Abdellah, Ahmed M,Hussein, Yassmein,Hasanin, MTM,Elbehairi, Serag Eldin I Asian Pacific Journal of Cancer Prevention 2015 Asian Pacific journal of cancer prevention Vol.16 No.14

Aims: To investigate effect of metallic nanoparticles, silver (AgNPs) and gold nanoparticles (AuNPs) as antitumor treatment in vitro against human breast cancer cells (MCF-7) and their associated mechanisms. This could provide new class of engineered nanoparticles with desired physicochemical properties and may present newer approaches for therapeutic modalities to breast cancer in women. Materials and Methods: A human breast cancer cell line (MCF-7) was used as a model of cells. Metallic nanoparticles were characterized using UV-visible spectra and transmission electron microscopy (TEM). Cytotoxic effects of metallic nanoparticles on MCF-7 cells were followed by colorimetric SRB cell viability assays, microscopy, and cellular uptake. Nature of cell death was further investigated by DNA analysis and flow cytometry. Results: Treatment of MCF-7 with different concentrations of 5-10nm diameter of AgNPs inhibited cell viability in a dose-dependent manner, with IC50 value of $6.28{\mu}M$, whereas treatment of MCF-7 with different concentrations of 13-15nm diameter of AuNPs inhibited cell viability in a dose-dependent manner, with IC50 value of $14.48{\mu}M$. Treatment of cells with a IC50 concentration of AgNPs generated progressive accumulation of cells in the S phase of the cell cycle and prevented entry into the M phase. The treatment of cells with IC50 concentrations of AuNPs similarly generated progressive accumulation of cells in sub-G1 and S phase, and inhibited the entrance of cells into the M phase of the cell cycle. DNA fragmentation, as demonstrated by electrophoresis, indicated induction of apoptosis. Conclusions: Our engineered silver nanoparticles effectively inhibit the proliferation of human breast carcinoma cell line MCF-7 in vitro at high concentration ($1000{\mu}M$) through apoptotic mechanisms, and may be a beneficial agent against human carcinoma but further detailed study is still needed.

Use of Recombinant Entamoeba histolytica Cysteine Proteinase 1 To Identify a Potent Inhibitor of Amebic Invasion in a Human Colonic Model

Melé,ndez-Ló,pez, Samuel G.,Herdman, Scott,Hirata, Ken,Choi, Min-Ho,Choe, Youngchool,Craik, Charles,Caffrey, Conor R.,Hansell, Elisabeth,Chá,vez-Munguí,a, Bibiana,Chen, Yen Tin American Society for Microbiology 2007 EUKARYOTIC CELL Vol.6 No.7

<B>ABSTRACT</B><P>Cysteine proteinases are key virulence factors of the protozoan parasite <I>Entamoeba histolytica</I>. We have shown that cysteine proteinases play a central role in tissue invasion and disruption of host defenses by digesting components of the extracellular matrix, immunoglobulins, complement, and cytokines. Analysis of the <I>E. histolytica</I> genome project has revealed more than 40 genes encoding cysteine proteinases. We have focused on <I>E. histolytica</I> cysteine proteinase 1 (EhCP1) because it is one of two cysteine proteinases unique to invasive <I>E. histolytica</I> and is highly expressed and released. Recombinant EhCP1 was expressed in <I>Escherichia coli</I> and refolded to an active enzyme with a pH optimum of 6.0. We used positional-scanning synthetic tetrapeptide combinatorial libraries to map the specificity of the P1 to P4 subsites of the active site cleft. Arginine was strongly preferred at P2, an unusual specificity among clan CA proteinases. A new vinyl sulfone inhibitor, WRR483, was synthesized based on this specificity to target EhCP1. Recombinant EhCP1 cleaved key components of the host immune system, C3, immunoglobulin G, and pro-interleukin-18, in a time- and dose-dependent manner. EhCP1 localized to large cytoplasmic vesicles, distinct from the sites of other proteinases. To gain insight into the role of secreted cysteine proteinases in amebic invasion, we tested the effect of the vinyl sulfone cysteine proteinase inhibitors K11777 and WRR483 on invasion of human colonic xenografts. The resultant dramatic inhibition of invasion by both inhibitors in this human colonic model of amebiasis strongly suggests a significant role of secreted amebic proteinases, such as EhCP1, in the pathogenesis of amebiasis.</P>

Transport function and subcellular distribution of purified human erythrocyte glucose transporter reconstituted into rat adipocytes

Jo, Inho,Hah, Jong S.,Rampal, Amrit L.,Chakrabarti, Ranjan,Paterson, Alan R.P.,Craik, James D.,Cass, Carol E.,Zobel, C. Richard,Jung, Chan Y. 이화여자대학교 생명과학연구소 1992 생명과학연구논문집 Vol.3 No.-

In order to delineate the insulin-independent(constitutive)and insulin-dependent regulations of the plasma membrane glucose transporter concentrations in rat adipocytes, we introduced purified human erythrocyte GLUT-1(HEGT) into rat adipocytes by poly(ethylene glycol)-induced vesicle-cell fusion and its transport function and subcellular distribution in the host cell were measured. HEGT in adipocytes catalysed 3-O-methylglucose equilibrium exchange with a turnover number that is indistinguishable from that of the basal adipocyte transporters. However, insulin did not stimulate significantly the HEGT funciton in adipocytes where it stimulated the native transporter function by 7-8-fold. The steady state distribution and the transmembrane orientation assays revealed that more than 85% of the HEGT that were inserted in the physiological, cytoplasmic side-in orientation at the adipocytes plasma membrane were moved into low-density microsomes(LDM), while 90% of the HEGT that were inserted in the wrong, cytoplasmic side-out orientation were retained in the plasma membrane. Furthermore, more than 70% of the LDM-associated HEGT were found in asmall subset of LDM that also contained 80% of the LDM-associated GLUT-4, the insulin-regulatable, native adipocyte glucose transporter. However, insulin did not cause redistribution of HEGT from LDM to the plasma membrane under the condition where it recruited GLUT-4 from LDM to increase the plasmamembrane GLUT-4 content 4-5-fold. These results demonstrate that the erythrocyte GLUT-1 introduced in adipocytes transports glucose with an intrinsic activity similar to that of the adipocyte GLUT-1 and / or GLUT-4, and enters the constitutive GLUT-4 recruitment pathway. We suggested that the adipocyte plasma membrane glucose transporter concentration is constivutively kept low by a mechanism where a cell-specific constitutent interacts with a cytoplasmic domain common to GLUT-1 and GLUT-4, while the insulin-dependent recruitment requires a cytoplasmic domain specific to GLUT-4.

Biochemical Properties of a Novel Cysteine Protease of <i>Plasmodium vivax</i> , Vivapain-4

Na, Byoung-Kuk,Bae, Young-An,Zo, Young-Gun,Choe, Youngchool,Kim, Seon-Hee,Desai, Prashant V.,Avery, Mitchell A.,Craik, Charles S.,Kim, Tong-Soo,Rosenthal, Philip J.,Kong, Yoon Public Library of Science 2010 PLoS neglected tropical diseases Vol.4 No.10

<▼1><P><B>Background</B></P><P>Multiple cysteine proteases of malaria parasites are required for maintenance of parasite metabolic homeostasis and egress from the host erythrocyte. In <I>Plasmodium falciparum</I> these proteases appear to mediate the processing of hemoglobin and aspartic proteases (plasmepsins) in the acidic food vacuole and the hydrolysis of erythrocyte structural proteins at neutral pH. Two cysteine proteases, vivapain (VX)-2 and VX-3 have been characterized in <I>P. vivax</I>, but comprehensive studies of <I>P. vivax</I> cysteine proteases remain elusive.</P><P><B>Findings</B></P><P>We characterized a novel cysteine protease of <I>P. vivax</I>, VX-4, of which orthologs appears to have evolved differentially in primate plasmodia with strong cladistic affinity toward those of rodent <I>Plasmodium</I>. Recombinant VX-4 demonstrated dual substrate specificity depending on the surrounding micro-environmental pH. Its hydrolyzing activity against benzyloxycarbonyl-Leu-Arg-4-methyl-coumaryl-7-amide (Z-Leu-Arg-MCA) and Z-Phe-Arg-MCA was highest at acidic pH (5.5), whereas that against Z-Arg-Arg-MCA was maximal at neutral pH (6.5–7.5). VX-4 preferred positively charged amino acids and Gln at the P1 position, with less strict specificity at P3 and P4. P2 preferences depended on pH (Leu at pH 5.5 and Arg at pH 7.5). Three amino acids that delineate the S2 pocket were substituted in VX-4 compared to VX-2 and VX-3 (Ala90, Gly157 and Glu180). Replacement of Glu180 abolished activity against Z-Arg-Arg-MCA at neutral pH, indicating the importance of this amino acid in the pH-dependent substrate preference. VX-4 was localized in the food vacuoles and cytoplasm of the erythrocytic stage of <I>P. vivax</I>. VX-4 showed maximal activity against actin at neutral pH, and that against <I>P. vivax</I> plasmepsin 4 and hemoglobin was detected at neutral/acidic and acidic pH, respectively.</P><P><B>Conclusion</B></P><P>VX-4 demonstrates pH-dependent substrate switching, which might offer an efficient mechanism for the specific cleavage of different substrates in different intracellular environments. VX-4 might function as a hemoglobinase in the acidic parasite food vacuole, a maturase of <I>P. vivax</I> plasmepsin 4 at neutral or acidic pH, and a cytoskeleton-degrading protease in the neutral erythrocyte cytosol.</P></▼1><▼2><P><B>Author Summary</B></P><P><I>Plasmodium vivax</I> affects hundreds of millions each year and results in severe morbidity and mortality. Plasmodial cysteine proteases (CPs) play crucial roles during the progression of malaria since inhibition of these molecules impairs parasite growth. These CPs might be targeted for new antimalarial drugs. We characterized a novel <I>P. vivax</I> CP, vivapain-4 (VX-4), which appeared to evolve differentially among primate <I>Plasmodium</I> species. VX-4 showed highly unique substrate preference depending on surrounding micro-environmental pH. It effectively hydrolyzed benzyloxycarbonyl-Leu-Arg-4-methyl-coumaryl-7-amide (Z-Leu-Arg-MCA) and Z-Phe-Arg-MCA at acidic pH and Z-Arg-Arg-MCA at neutral pH. Three amino acids (Ala90, Gly157 and Glu180) that delineate the S2 pocket were found to be substituted in VX-4. Alteration of Glu180 abolished hydrolytic activity against Z-Arg-Arg-MCA at neutral pH, indicating Glu180 is intimately involved in the pH-dependent substrate preference. VX-4 hydrolyzed actin at neutral pH and hemoglobin at acidic pH, and participated in plasmepsin 4 activation at neutral/acidic pH. VX-4 was localized in the food vacuoles and cytoplasm of the erythrocytic stage of <I>P. vivax</I>. The differential substrate preferences depending on pH suggested a highly efficient mechanism to enlarge biological implications of VX-4, including hemoglobin degradation, maturation of plasmepsin, and remodeling of the parasite architecture during growth and development of <I>P. vivax</I>.</P></▼2>