Biomedical subjects
Publications and source records attributed to Y Gu.
Heteroaryl and cycloalkyl sulfonamide-hydroxamic acid MMP inhibitors were investigated. Of these, the pyridyl analogue 2 is the most potent and selective inhibitor of MMP-9 and MMP-13 in vitro.
Although the peptide C(alpha)H group has historically not been thought to form hydrogen bonds within proteins, ab initio quantum calculations show it to be a potent proton donor. Its binding energy to a water molecule lies in the range between 1.9 and 2.5 kcal/mol for nonpolar and polar amino acids; the hydrogen bond (H-bond) involving the charged lysine residue is even stronger than a conventional OH..O interaction. The preferred H-bond lengths are quite uniform, about 3.32 A. Formation of each interaction results in a downfield shift of the bridging hydrogen's chemical shift and a blue shift in the C(alpha)H stretching frequency, potential diagnostics of the presence of such an H-bond within a protein.
The modified base 7,8-dihydro-8-oxo-guanine (8-oxoG) is one of the most stable deleterious products of oxidative DNA damage because it mispairs with adenine during DNA replication. In the fission yeast Schizosaccharomyces pombe, the MutY homolog (SpMYH) is responsible for removing misincorporated adenines from A/8-oxoG or A/G mismatches and thus preventing G:C to T:A mutations. In order to study the functional role of SpMYH, an SpMYH knockout strain was constructed. The SpMYH knockout strain, which does not express SpMYH and has no A/8-oxoG glycosylase activity, displays a 36-fold higher frequency of spontaneous mutations than the wild type strain. Disruption of SpMYH causes increased sensitivity to H2O2 but not to UV-irradiation. Expression of SpMYH in the mutant cells restores the adenine glycosylase activity, reduces the mutation frequency, and elevates the resistance to H2O2. Asp172 of SpMYH is conserved in a helix-hairpin-helix superfamily of glycosylases. The SpMYHA strain expressing D172N SpMYH retained the mutator phenotype. Moreover, when D172N mutant SpMYH was expressed in the wild-type cells, the mutation frequency observed was even higher than that of the parental strains. Thus, a mutant SpMYH that retains substrate-binding activity but is defective in glycosylase activity exhibits a dominant negative effect. This is the first demonstration that a MutY homolog plays an important role in protecting cells against oxidative DNA damage in eukaryotes.
BACKGROUND: The use of video-assisted thoracic surgery (VATS) techniques to mobilize the phrenic nerve in the thoracic cavity for neurotization after brachial plexus injury was studied. METHODS: From August 1999 to January 2000, 10 men and 1 woman with brachial plexus injury (left side in 5 and right side in 6) joined the study group. Their ages ranged from 20 to 38 years (average, 28 years). Supine after general anesthesia, all the patients had double-lumen trachea cannulas to guarantee complete lung collapse on the operative side. Three port incisions were made to allow introduction of the following: a 10-mm Stryker endoscope through the sixth intercostal space 2 cm medial to the anterior axillary line, one instrument for manipulation in the anterior axillary line of the third intercostal space, and another in the second intercostal space about 2 cm lateral to the parasternal line. The nerve was mobilized with two common long Mixter clamps and some endoscopic instruments by blunt and sharp dissection. RESULTS: All patients were managed successfully without severe complications. The mean additional length of phrenic nerves by this technique was 16 cm. CONCLUSIONS: Mobilization of the phrenic nerve by VATS is a safe and minimally invasive method for elongating the nerve for neurotization after brachial plexus injury.
The nucleus lentiformis mesencephali and the nucleus of the basal optic root in birds, homologous to the nucleus of the optic tract and the terminal nuclei of the accessory optic tract in mammals, are involved in optokinetic nystagmus. The present study provides the first electrophysiological evidence that reversible blockade of the pigeon nucleus of the basal optic root by lidocaine can change visual responsiveness of pretectal neurons in a direction-dependent manner. Thirty pretectal cells examined were classified as unidirectional (80%), bidirectional (10%) and omnidirectional (10%) cells according to their directional selectivity. Among the unidirectional cells, seven cells changed firing rates in all directions of motion, 11 changed visual responses only in the preferred directions and six others did not change their responsiveness during lidocaine. Most of the bidirectional cells changed firing rates in the temporonasal direction, and two-thirds of the omnidirectional cells showed these changes in all directions. Thirteen lidocaine administration sites were marked within the nucleus of the basal optic root and 19 recording sites were marked within the nucleus lentiformis mesencephali. This histological verification indicates that the effects of lidocaine blockade in the accessory optic nucleus on the directional selectivity and visual responsiveness of pretectal cells appear to be related, to some extent, to the location of drug injections in the nucleus of the basal optic root. This study has found that visual neurons in the nucleus of the basal optic root, which predominantly prefer vertical and backward motion, could modulate the directional selectivity and visual responsiveness of neurons in the nucleus lentiformis mesencephali, which mainly prefer horizontal motion. It is conceivable that both nuclei work together in coordination and in competition during optokinetic nystagmus.
Lactic acid bacteria (LAB) are characterised by their conversion of a large proportion of their carbon feed, fermentable sugars, to lactic acid. However, in addition to lactic acid production, the LAB are able to divert a small proportion of fermentable sugars towards the biosynthesis of exopolysaccharides (EPSs) that are independent of the cell surface and cell wall material. These microbial EPSs when suspended or dissolved in aqueous solution provide thickening and gelling properties, and, as such, there is great interest in using EPSs from food grade microorganisms (such as the LAB that are traditionally used for food fermentations) for use as thickening agents. The current review includes a brief summary of the recent literature describing features of the biosynthetic pathways leading to EPS production. Many aspects of EPS biosynthesis in LAB are still not fully understood and a number of inferences are made regarding the similarity of the pathway to those involved in the synthesis of other cell polysaccharides, e.g., cell wall components. The main body of the review will cover practical aspects concerned with the isolation and characterisation of EPS structures. In the last couple of years, a substantial number of structures have been published and a summary of the common elements of these structures is included as is a suggestion for a system for representing structures. A brief highlight of the attempts that are being made to design 'tailor'-made polysaccharides using genetic modification and control of metabolic flux is presented.
The Fourier-transform Raman (FT-Raman), infrared (FT-IR), and UV-visible absorption spectra of four dipyrrinones and two mesobilirubins have been investigated in the solid state and in CH2Cl2 solutions. A detailed spectral analysis, assignment and discussion of these spectra are presented. The bands at 1735-1738, 1691-1707 and 1359-1377 cm(-1) which were assigned to the stretching vibrations of the C-O-C and C-O-H and symmetric deformation of C-H bonds, respectively, can act as a marker to distinguish the compounds of this class. The striking differences between the spectra of the compounds suggest that mesobilirubin XIIIalpha is tending to adopt as ridge-tile conformation, rather than linear conformation.
Osteocytes play an important role in signaling within bone. Communication of osteocytes with each other and with bone lining cells may have a function in mineral homeostasis and mechanotransduction. However, very little is known of the expression of ion channels in these cells. Using the whole-cell patch-clamp technique, we have detected three types of K(+) currents in the mouse osteocyte-like cell line MLO-Y4. The most commonly observed current (48% of cells) activated rapidly (20 msec) in response to depolarizing steps from -40 mV and exhibited voltage-dependent inactivation. The current was inhibited by 20 mmol/L tetraethyl ammonium (TEA) and abolished by intracellular 2 mmol/L 4-aminopyridine (4-AP). Biophysical and pharmacological characteristics of the current differed from those of inactivating K(+) currents in osteoblastic cells. In 22% of cells, a slowly activating, voltage-activated current was observed (threshold at 20-30 mV). This current was TEA insensitive, was abolished by intracellular application of 2 mmol/L 4-AP, and was strongly inhibited by apamin, a selective inhibitor of small conductance (SK) Ca(2+)-activated K(+) channels. A third current developed during whole-cell dialysis (37% of cells). This current showed little voltage sensitivity. It was abolished by intracellular application of 2 mmol/L 4-AP, high-extracellular Ba(2+) (108 mmol/L), or by inclusion of ATP in the intracellular solution, but was insensitive to TEA, apamin, Cs(+), and glibenclamide. None of these currents was affected by replacement of chloride with acetate in the bath or pipette salines. Reverse-transcription polymerase chain reaction confirmed the presence of mRNA for the types 1 and 2 SK channels, but message for the large conductance (BK) Ca(2+)-activated K(+) channel was not detected in these cells. Message for the sulphonylurea receptor SUR2, a subunit of glibenclamide-insensitive ATP-dependent K(+) channels (K(ATP)), was also detected, but the glibenclamide-sensitive SUR1 subunit was not. These data are the first descriptions of SK- and ATP-sensitive, glibenclamide-insensitive channels in cells of osteoblastic lineage. Our findings are consistent with a change in K(+) channel expression during differentiation from osteoblast to osteocyte. K(+) channels of osteocytes will contribute to maintenance of the cell membrane potential and thus may participate in mechanosensitivity and osteocyte intercellular communication. In addition, they may be involved in homeostatic maintenance of the extracellular fluid occupying the periosteocytic space.
Growth hormone (GH) has been reported to be useful to treat heart failure. To elucidate whether GH has direct beneficial effects on the heart, we examined effects of GH on oxidative stress-induced apoptosis in cardiac myocytes. TUNEL staining and DNA ladder analysis revealed that hydrogen peroxide (H2O2)-induced apoptosis of cardiomyocytes was significantly suppressed by the pretreatment with GH. GH strongly activated extracellular signal-regulated kinases (ERKs) in cardiac myocytes and the cardioprotective effect of GH was abolished by inhibition of ERKs. Overexpression of dominant negative mutant Ras suppressed GH-stimulated ERK activation. Overexpression of Csk that inactivates Src family tyrosine kinases also inhibited ERK activation evoked by GH. A broad-spectrum inhibitor of protein tyrosine kinases (PTKs), genistein, strongly suppressed GH-induced ERK activation and the cardioprotective effect of GH against apoptotic cell death. GH induced tyrosine phosphorylation of EGF receptor and JAK2 in cardiac myocytes, and an EGF receptor inhibitor tyrphostin AG1478 and a JAK2 inhibitor tyrphostin B42 completely inhibited GH-induced ERK activation. Tyrphostin B42 also suppressed the phosphorylation of EGF receptor stimulated by GH. These findings suggest that GH has a direct protective effect on cardiac myocytes against apoptosis and that the effect of GH is attributed at least in part to the activation of ERKs through Ras and PTKs including JAK2, Src, and EGF receptor tyrosine kinase.
AIMS: To compare galactose-negative strains of Streptococcus thermophilus and Lactobacillus delbrueckii subspecies bulgaricus isolated from fermented milk products and known to produce exopolysaccharides (EPSs). METHODS AND RESULTS: The structures of the EPSs were determined using nuclear magnetic resonance (NMR) and their genetic relationships determined using restriction endonuclease analysis (REA) and random amplification of polymorphic DNA (RAPD). Similar groupings were apparent by REA and RAPD, and each group produced an EPS with a particular subunit structure. CONCLUSION: Although none of the strains assimilated galactose, all inserted a high proportion of galactose into their EPS when grown in skimmed milk, and fell into three distinct groups. SIGNIFICANCE AND IMPACT OF THE STUDY: This information should help in an understanding of genetic exchanges in lactic acid bacteria.
To observe the effect of sensory components on muscle atrophy, 4 surgical procedures applying sensory nerves or neurons to the denervated muscle were conducted in a rat model: sensory nerve implantation (group B), sensory motor nerve crossover (group C), dorsal root ganglia implantation (group D), and implantation of preganglionically avulsed sensory nerve (group E). Rats with complete denervation served as controls (group A). The degree of muscle atrophy was evaluated after surgery by fibrillation potential amplitude, muscle wet weight, muscle fiber cross-sectional area, collagen content, and protein content. Compared with group A, the results in groups B, D, and E were superior 1 month after surgery and the results in groups B, C, and E were superior 3 months after surgery. Implantation of normal or preganglionically avulsed sensory nerve delayed atrophy. Dorsal root ganglia implantation only had a short-term trophic influence. In the animal model setting, sensory motor nerve crossover is effective only after it is maintained for at least 3 months.
Increased neutrophil activation has been demonstrated in women with pre-eclampsia. Activated neutrophils may play a significant role in the vascular endothelial pathophysiology in this disorder of pregnancy. How neutrophils become activated in pre-eclampsia is unknown. It has been proposed that activating factors could be produced and released by the placenta. To test if placental factors could stimulate neutrophil activation and what mechanism might be involved, neutrophils isolated from healthy female volunteers were exposed to the conditioned medium (CM) derived from either normal (Nor) or pre-eclamptic (PE) placental villous culture. Neutrophil-endothelial adhesion, neutrophil superoxide generation, elastase activity and integrin expression were measured. The data were analysed by ANOVA. A P value less than 0.05 was considered statistically significant. All values are expressed as a mean+/-s.e. We found: (1) neutrophil-endothelial adhesion was significantly increased in neutrophils exposed PE-CM than those exposed to Nor-CM and non-CM, P< 0.01; (2) both Nor-CM and PE-CM could stimulate neutrophils to generate more superoxide radicals; (3) there was no difference in elastase activity after neutrophil exposure to Nor-CM compared to PE-CM, P> 0.1; (4) significant changes in CD62L and CD11b expression were found in neutrophils exposed to PE-CM. We conclude that factors produced by the placenta can activate neutrophils by an increase in superoxide generation and modulation of adhesion molecule expression. Upregulation of surface adhesion molecule CD11 expression may be responsible for the increased neutrophil-endothelial adhesion induced by factors derived from pre-eclamptic placentae.
This study was performed to compare the effects of recombinant human growth hormone (rhGH), glutamine (Gln) and simultaneous treatment with rhGH and Gln in rats subjected to 75% intestinal resection and maintained with parenteral nutrition (PN) for 6 days. Morphological changes including mucosal thickness, villus height, crypt depths and villus surface area of the residue jejunum were measured under a light microscope; expression of PCNA as an index of cell proliferation and apoptotic cells were observed using immunohistochemical staining; Ileal IGF-1 mRNA was determined by Northern blot analysis. The morphological parameters of the jejunal mucosa in rats treated with PN alone were only about 52-62% of those in reference group (P<0.01), this atrophy of the jejunal mucosa was accompanied by a 2.5-fold decrease in absolute counts of PCNA and a 10-fold increase in apoptotic index (P<0.01), IGF-1 mRNA transcript in residue ileum was decreased significantly (P<0.01). However, with rhGH or Gln, the mucosal architecture was improved significantly and was further improved when rhGH and Gln were given together, the morphological values in rats treated with Gln+rhGH was 79% higher than those with PN alone, and was associated with a 2-fold increase in PCNA counts and a 4-fold decrease in apoptotic index (P<0.01), IGF-1 mRNA expression was 78% higher than those with PN alone (P<0.01). We conclude that rhGH and Gln have synergistic effects on adaptation of the intestinal remnant in parenterally fed, short-bowel rats. The underlying mechanisms are associated with increased proliferation and decreased apoptosis in the intestinal epithelial cells. Local intestinal production of IGF-1 plays an important role in adaptation of the small intestine. Our findings support the concept that specific gut-trophic nutrients and growth factors may be combined to enhance the intestinal adaptation.
Murine class Alpha glutathione (GSH) transferase A1-1 (mGSTA1-1) is unique among mammalian Alpha class GSTs due to its exceptionally high catalytic activity toward (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-anti-BPDE], which is the activated metabolite of an environmentally relevant carcinogen, benzo[a] pyrene (BP). However, the molecular basis for high catalytic activity of mGSTA1-1 toward (+)-anti-BPDE is not clear. In the present study, we demonstrate that an arginine residue at position 216, which is conserved in some but not all mammalian class Alpha GSTs, plays an important role in catalytic activity of mGSTA1-1 toward (+)-anti-BPDE and carcinogenic diol epoxides of other environmentally relevant polycyclic aromatic hydrocarbons (PAHs). The catalytic efficiency (k(cat)/K(m)) of mGSTA1-1 for the GSH conjugation of (+)-anti-BPDE (108/mM/s) was reduced by about 58% upon replacement of arginine 216 with alanine (R216A). This was mainly due to a significantly lower V(max) for the R216A mutant of mGSTA1-1 compared with wild-type mGSTA1-1. The R216A mutation also resulted in a statistically significant reduction (>70%) in specific activity of mGSTA1-1 toward racemic anti-diol epoxides of chrysene and benzo[c]phenanthrene (anti-CDE and anti-B[c]PDE, respectively). The catalytic activity of mGSTA2-2, which is a close structural homologue of mGSTA1-1, was also reduced upon R216A mutation. The results of the present study clearly indicate that an arginine residue at position 216 is critical for catalytic activity of mGSTA1-1 and mGSTA2-2 toward carcinogenic diol epoxide metabolites of various PAHs that are abundant in the environment and suspected human carcinogens.
BACKGROUND: Aetiological profiles of non-Hodgkin's lymphoma (NHL) may differ depending upon whether the disease is inheritance-related or sporadic. Because familial risk (a probable surrogate of inheritance-relatedness) of NHL is influenced by haematolymphoproliferative malignancies (HLPM), we evaluated whether non-familial risk factors differ between NHL with and without a family history of HLPM, using the Selected Cancers Study data. METHODS: Cases were 1511 men aged 31-59 and diagnosed with NHL during 1984-1988. Controls were men without NHL, frequency-matched to cases by age range and cancer registry (n = 1910). These groups were compared: cases with a family history of HLPM and without, and controls without such a family history. RESULTS: Polytomous logistic regression analyses showed that the odds ratio (OR) estimates of homosexual behaviour were 18.2 (95% confidence interval (CI) : 4.8-69.4) and 5.6 (95% CI : 3.3-9.5) for NHL with and without a family history of HLPM, respectively. The corresponding estimates were 3.9 (95% CI : 1.7-8.9) and 2.2 (95% CI : 1.5-3.1) for history of enlarged lymph nodes. Variables only related to NHL with a family history were use of heroin (OR = 15.6, 95% CI : 3.4-70.4), exposure to a chlorinated hydrocarbon pesticide (OR = 2.3, 95% CI : 1.0-5.0), occupational exposure to plywood, fibreboard or particleboard (OR = 2.0, 95% CI : 1.2-3.4) and history of liver diseases (other than hepatitis or cirrhosis) (OR = 6.5, 95% CI : 1.2-36.2). The association between homosexual behaviour and NHL among men with a family history was stronger for those aged 31-44, especially for B-cell type of the disease. CONCLUSIONS: This study suggests differences in the risk factor profiles between NHL with and without a family history of HLPM. The higher risks of NHL for homosexual behaviour and heroin use, surrogates of HIV infection, in men with a family history of HLPM imply that genetic susceptibility may be influential on the occurrence of HIV-related NHL.
BACKGROUND: Skeletal muscle atrophy induced by denervation is associated with apoptosis. This study was undertaken to determine the role of apoptosis and the expression of apoptosis-associated genes in rat skeletal muscle made atrophic by brachial plexus injury, and to study the apoptotic signal transduction pathway. METHODS: An animal model of skeletal muscle atrophy was established in rats by severing the brachial plexus of one forelimb. Apoptosis of muscle cells was investigated with terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate end labeling, flow cytometry, deoxyribonucleic acid (DNA) electrophoresis, and electron microscopy. The apoptosis-associated genes Fas, FADD, Caspase 8, c-myc, p53, and Bcl-2 were detected by immunohistochemistry and Northern blot. RESULTS: By terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate end labeling and flow cytometry we found that the percentage of apoptotic muscle cells was higher in atrophic than in healthy skeletal muscle (p < 0.05). DNA laddering could be seen in gel electrophoresis of DNA from atrophic muscle. By electron microscopy, we observed morphologic change of early apoptosis, such as aggregation of chromosomes, expansion of nucleic cistern, and contraction of the nucleus. Using immunohistochemistry, we determined that in atrophic muscle Fas, FADD, and Caspase-8 genes were highly expressed, whereas Bcl-2 was poorly expressed (p < 0.01). However, we did not detect a change in the expression of p53 or c-myc genes. Northern blots indicated that Fas messenger ribonucleic acid was higher and Bcl-2 messenger ribonucleic acid was lower in atrophic than in healthy muscle (p < 0.01). CONCLUSION: There are many more apoptotic cells in muscle atrophied as a result of brachial plexus injury than in healthy muscle, and apoptosis plays an important role in the pathogenesis of atrophy. The apoptotic signal may be transmitted from Fas to FADD to Caspase-8, with a decrease in Bcl-2 expression aggravating the process.
The plant-specific Rop family GTPases are versatile molecular switches in many processes during plant growth, development, and responses to the environment. To understand how Rop achieves its functional versatility in signaling, we performed a genome-wide identification of putative Rop targets using a combination of the yeast two-hybrid method, bioinformatic tools, and a robust functional assay in pollen. In this study, we have identified 11 Arabidopsis genes encoding novel proteins, termed RICs (for Rop-interactive CRIB motif-containing proteins), that contain a CRIB (for Cdc42/Rac-interactive binding) motif required for their specific interaction with GTP-bound Rop1. RICs are divergent and classified into five groups that share little sequence homology outside of the conserved Rop-interactive domain. Overexpression in tobacco pollen tubes of the nine Ric genes that are expressed in Arabidopsis pollen causes distinct phenotypes, implying distinct functions for various RICs. RIC3 (group III) and RIC4 (group V) both cause depolarized growth like Rop1 and display Rop1-enhanced localization to the tip of pollen tubes, suggesting that these RICs may be two distinct targets of Rop1. In contrast, RIC10 (group I) promotes pollen tube elongation but does not affect pollen tube growth polarity and shows Rop1-independent localization to the cytoplasm, suggesting that RIC10 may participate in a Rop1-independent pathway probably controlled by a different Rop. Expression of all other RICs causes various degrees of growth inhibition in pollen tubes. Furthermore, these inhibitory RICs also exhibit distinct patterns of localization in pollen tubes. Our results suggest that various RICs have evolved to interact with Rops differentially and to perform distinct functions in pollen tubes. Reverse transcriptase-mediated polymerase chain reaction analysis showed that six of the nine RICs are expressed in various parts of Arabidopsis plants. On the basis of these observations, we propose that RICs function as Rop GTPase targets that control various Rop-dependent signaling pathways in plants.
Large-field patterns are effective stimuli for eliciting visual responses from neurons in the pretectal nucleus lentiformis mesencephali of nonmammals. The present study shows that stimulation beyond the receptive field does not contribute to the visual responses of neurons in the nucleus lentiformis mesencephali in two respects. First, changes in the direction and velocity of motion beyond the receptive field did not affect the visual responses of the pretectal cells to motion within the receptive field. Second, time differences in the onset of stimulation within and outside the receptive field did not influence the visual responses of the pretectal cells to motion in the receptive field, implying that there may be no long-range interaction between the receptive field and its surrounding field. The present study also indicates that the pretectal cells are not only sensitive to the direction and velocity of motion, but also to the size and density of dots in a random-dot pattern moving through the receptive field. Taken together with previous studies, these results suggest that the receptive field of the pretectal cells within the nucleus lentiformis mesencephali is large in size but well defined in boundaries, and that the pretectal cells respond to motion of visual stimuli within but not beyond their receptive fields.