Poly(3-hydroxybutyrate) biosynthesis in the biofilm of Alcaligenes eutrophus, using glucose enzymatically released from pulp fiber sludge

Glucose, enzymatically released from pulp fiber sludge, was combined with inorganic salts and used as a growth medium for Alcaligenes eutrophus, a gram-negative strain producing poly(3-hydroxybutyrate) (PHB). By controlling the concentrations of the inorganic salts in the growth medium, almost 78% of the cell mass was converted to pure PHB. Efforts were made to find conditions for bacterial growth in the form of a biofilm on a cheap and reusable carrier. A number of positively charged carriers were tested, and the anion exchanger DEAE-Sephadex A-25 was chosen as a microcarrier for packed-bed biofilm cultures of A. eutrophus. Conditions for attachment, growth, and detachment were established. Biofilm formation on the microcarrier is strongly dependent on the ionic strength of the attachment medium. In order to achieve formation of the biofilm and its recovery from the microcarrier, the ionic strengths of the attachment and the detachment media were varied. Low ionic strength was tested for attachment, and high ionic strength was tested for detachment. Although biofilm formation in the packed-bed reactor is limited, the volumetric yield of cells based on the void volume of the packed bed is comparable with the batch culture yield.     

Bile acid amides derived from chiral amino alcohols: novel antimicrobials and antifungals

Cholic and deoxycholic acid amides 10-17 have been synthesised from (1R,2R)-1-phenyl-2-amino-1,3-propanediol 2, (1S,2S)-1-phenyl-2-amino-1,3-propanediol 4, (1R,2R)-1-para-nitrophenyl-2-amino-1,3-propanediol 3, (1S,2S)-1-para-nitrophenyl-2-amino-1,3-propanediol 5. Amide 12 derived from N-succinimidyl ester 9 of deoxycholic acid and (1R,2R)-1-phenyl-2-amino-1,3-propanediol 2, found to be active against Cryptococcus neoformans and the amide 17 obtained from N-succinimidyl ester 9 of deoxycholic acid and (1S,2S)-1-para-nitrophenyl-2-amino-1,3-propanediol 5, is found to be potent against various gram-positive bacteria.     

Rafts: scale-dependent, active lipid organization at the cell surface

Rafts have been conceptualized as lateral heterogeneities in the organization of cholesterol and sphingolipids, endowed with sorting and signaling functions. In this review we critically examine evidence for the main tenet of the 'raft hypothesis', namely lipid-dependent segregation of specific membrane components in the plasma membrane. We suggest that conventional approaches to studying raft organization wherein membranes are treated as passive, thermally equilibrated systems are unlikely to provide an adequate framework to understand the mechanisms of raft-organization in vivo. An emerging view of raft organization is that it is spatio-temporally regulated at different scales by the cell. This argues that rafts must be defined by simultaneous observation of components involved in particular functions. Recent evidence from the study of glycosylphosphatidyl inositol-anchored proteins, a common raft-marker, supports this picture in which larger scale, more stable rafts are induced from preexisting small-scale lipid-dependent structures actively maintained by cellular processes.     

Mouse zinc transporter 1 gene provides an essential function during early embryonic development

The SLC30 family of cation diffusion transporters includes at least nine members in mammals, most of which have been documented to play a role in zinc transport. The founding member of this family, Znt1, was discovered by virtue of its ability to efflux zinc from cells and to protect them from zinc toxicity. However, its physiological functions remain unknown. To address this issue, mice with targeted knockout of the Znt1 gene were generated by homologous recombination in embryonic stem cells. Heterozygous Znt1 mice were viable. In contrast, homozygous Znt1 mice died in utero soon after implantation due to a catastrophic failure of embryonic development. Although extraembryonic membranes formed around these embryos, the embryo proper failed to undergo morphogenesis past the egg cylinder stage and was amorphous by d9 of pregnancy. Expression of the Znt1 gene was detected predominantly in trophoblasts and in the maternal deciduum during the postimplantation period (d5 to d8). The failure of homozygous Znt1 embryos to develop could not be rescued by manipulating maternal dietary zinc (either excess or deficiency) during pregnancy. However, embryos in Znt1 heterozygous females were approximately 3 times more likely to develop abnormally when exposed to maternal dietary zinc deficiency during later pregnancy than were those in wildtype females. These studies suggest that Znt1 serves an essential function of transporting maternal zinc into the embryonic environment during the egg cylinder stage of development, and further suggest that Znt1 plays a role in zinc homeostasis in adult mice.     

Synthesis and structure-activity relationship studies of cinnamic acid-based novel thiazolidinedione antihyperglycemic agents

A number of 2,4-thiazolidinedione derivatives of -phenyl substituted cinnamic acid were synthesized and studied for their PPAR agonist activity. The E-isomer of cinnamic acid, 11, showed moderate PPAR transactivation. The corresponding Z-isomer, 23, and double bond reduced derivative, 15, were found to be much less potent. Although the E-isomer showed a moderate PPAR gamma transactivation, it demonstrated a strong glucose-lowering effect in a genetic rodent model of diabetes. Results of pharmacokinetic, metabolism and permeability studies are consistent with 11 being an active prodrug with an active metabolite, 14, that has similar glucose lowering and PPAR gamma agonist properties.     

Ebselen inhibits NO-induced apoptosis of differentiated PC12 cells via inhibition of ASK1-p38 MAPK-p53 and JNK signaling and activation of p44/42 MAPK and Bcl-2

Ebselen, a selenium-containing heterocyclic compound, prevents ischemia-induced cell death. However, the molecular mechanism through which ebselen exerts its cytoprotective effect remains to be elucidated. Using sodium nitroprusside (SNP) as a nitric oxide (NO) donor, we show here that ebselen potently inhibits NO-induced apoptosis of differentiated PC12 cells. This was associated with inhibition of NO-induced phosphatidyl Serine exposure, cytochrome c release, and caspase-3 activation by ebselen. Analysis of key apoptotic regulators during NO-induced apoptosis of differentiated PC12 cells showed that ebselen blocks the activation of the apoptosis signaling-regulating kinase 1 (ASK1), and inhibits phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-jun N-terminal protein kinase (JNK). Moreover, ebselen inhibits NO-induced p53 phosphorylation at Ser15 and c-Jun phosphorylation at Ser63 and Ser73. It appears that inhibition of p38 MAPK and p53 phosphorylation by ebselen occurs via a thiol-redox-dependent mechanism. Interestingly, ebselen also activates p44/42 MAPK, and inhibits the downregulation of the antiapoptotic protein Bcl-2 in SNP-treated PC12 cells. Together, these findings suggest that ebselen protects neuronal cells from NO cytotoxicity by reciprocally regulating the apoptotic and antiapoptotic signaling cascades.     

Activation of protein kinase C inhibits TRAIL-induced caspases activation, mitochondrial events and apoptosis in a human leukemic T cell line

TRAIL causes apoptosis in numerous types of tumor cells. However, the mechanisms regulating TRAIL-induced apoptosis remain to be elucidated. We have investigated the role of PKC in regulating TRAIL-induced mitochondrial events and apoptosis in the Jurkat T cell line. We found a caspase-dependent decline in mitochondrial membrane potential and translocation of cytochrome c from mitochondria into the cytosol in response to TRAIL. Both these events were prevented by PKC activation. Moreover, PKC activation considerably reduced the activation of caspases, PARP cleavage and apoptosis when induced upon TRAIL treatment. MAPK activation was involved in the mechanism of PKC-mediated inhibition of TRAIL-induced cytochrome c release from mitochondria. Furthermore, inhibition of the MAPK pathway partially reversed the PKC-mediated inhibition of TRAIL-induced apoptosis. Besides, PKC activation may also inhibit the TRAIL-induced apoptosis through a MAPK-independent mechanism. Altogether, these results indicate a negative role of PKC in the regulation of apoptotic signals generated upon TRAIL receptor activation.     

Terminal uridine nick end labelling and mitosis in breast carcinoma. Correlation with tumor grade and p53 overexpression

OBJECTIVE: To study the relationship of cell death and proliferation to histologic grade and p53 expression in invasive carcinoma of the breast. STUDY DESIGN: A total of 31 cases of infiltrating duct carcinoma of the breast were randomly selected. The terminal deoxynucleotidyl-transferase-mediated dUTP nick end labelling (TUNEL) reaction and p53 immunostaining were performed on representative paraffin-embedded tissue sections. Mitotic and apoptotic indices (MI and AI) were also measured on hematoxylin-eosin-stained sections. Histologic grade of infiltrating duct carcinoma was performed with the help of the Nottingham modification of the Bloom-Richardson system. Tumor grade and p53 overexpression were correlated with MI, AI and AI detected by TUNEL. RESULTS: There were a total of 31 infiltrating duct carcinomas of the breast, of which 13 cases were grade 1 and nine cases each were grade 2 and 3. Cells with positive TUNEL showed a strong brown nuclear positivity. TUNEL showed positivity from the periphery of the nuclear margin to the central portion. AI detected by TUNEL did not correlate with tumor grade (ANOVA, P > .05). MI was significant only in grade 1 versus grade 3 and 2 versus grade 3 carcinomas (ANOVA, P < .01). The morphologic apoptotic index was significant only in grade 1 versus grade 3 carcinomas. Nine cases showed p53 overexpression, and the rest of the cases were negative for p53 immunostaining. MI, AI and TUNEL were not significantly different in p53-negative and -positive groups. Pearson's correlation coefficient showed that AI and MI were significantly related, but there was no significant relation between AI detected by TUNEL and MI. CONCLUSION: MI is still more useful than AI or AI detected by TUNEL in differentiating various grades of carcinoma of the breast.     

Cytochrome P450 2E1 dependent catalytic activity and lipid peroxidation in rat blood lymphocytes

To investigate the similarities in the catalytic activity of blood lymphocyte P450 2E1 in blood lymphocyte with the liver isoenzyme, NADPH dependent lipid peroxidation and activity of N-nitrosodimethyamine demethylase (NDMA-d) was studied in rat blood lymphocytes. Blood lymphocytes were found to catalyse NADPH dependent (basal) lipid peroxidation and demethylation of N-nitrosodimethylamine (NDMA). Pretreatment with ethanol or pyrazole or acetone resulted in significant increase in the NADPH dependent lipid peroxidation and the activity of NDMA-d in blood lymphocytes and liver microsomes. In vitro addition of CCl(4) to the blood lymphocytes isolated from control or ethanol pretreated rats resulted in an increase in the NADPH dependent lipid peroxidation. Significant inhibition of the basal and CCl(4) supported NADPH dependent lipid peroxidation and NDMA-d activity in blood lymphocytes isolated from control or ethanol pretreated rats by dimethyl formamide or dimethyl sulfoxide or hexane, solvents known to inhibit P450 2E1 catalysed reactions in liver and anti- P450 2E1, have indicated the role of P450 2E1 in the NADPH dependent lipid peroxidation in rat blood lymphocytes. The data indicating similarities in the NADPH dependent lipid peroxidation and NDMA-d activity in blood lymphocyte with the liver microsome have provided evidence that blood lymphocyte P450 2E1 could be used as a surrogate to monitor and predict hepatic levels of the enzyme.