Biomedical subjects

Y Yao

Publications and source records attributed to Y Yao.

At least 181 records · Page 10Linked to original sources

PubMed1997

Urinary and tissue levels of scatter factor in transitional cell carcinoma of bladder.

PURPOSE: We previously reported increased titers of scatter factor in urine of 20 patients with transitional cell carcinoma of the bladder compared to noncancer and cancer control groups. Scatter factor was also found in bladder tumor extracts but the number of samples examined was too small for detailed analysis. We report a followup study of larger numbers of patients with transitional cell carcinoma and controls. MATERIALS AND METHODS: The scatter factor content of urine samples and bladder tissue extracts was measured by enzyme-linked immunosorbent assay. Values were normalized per milligram creatinine or tissue protein. Statistical analysis was performed using the Mann-Whitney U test or, for multiple comparisons, the Kruskal-Wallis H test. RESULTS: Patients with transitional cell carcinoma of the bladder (52) had higher urinary scatter factor titers than did 24 normal subjects (p < 0.001) or 14 with benign prostatic hypertrophy (p < 0.001), 49 with prostate cancer (p < 0.001) and 13 with other genitourinary tract cancers (p < 0.01). Transitional cell carcinoma cases with clinicopathological evidence of disease had greater urinary scatter factor levels than those with no evidence of disease at urine sampling (p < 0.01). However, patients with transitional cell carcinomas in remission still had much greater urinary scatter factors than did normal subjects (p < 0.001). In contrast, patients with active prostate cancer had urinary scatter factor levels similar to those in remission. Patients with muscle invasive or high grade transitional cell carcinomas tended to have higher urinary scatter factor levels than patients with nonmuscle invasive or low grade tumors, respectively, but the differences were not significant. Greater levels of scatter factor were present in tissue extracts of muscle invasive transitional cell carcinomas than in nonmuscle invasive tumors (p < 0.001) or nontumor tissue (p < 0.02). Invasion was more closely related to tissue scatter factor content than tumor grade, since high grade noninvasive transitional cell carcinomas had a scatter factor content similar to that of low grade noninvasive transitional cell carcinomas. CONCLUSIONS: These studies suggest that scatter factor may be a marker of bladder cancer, urinary scatter factor titers tend to reflect disease activity and particularly high tissue titers of scatter factor are found in muscle invasive cancers. A larger prospective study will be necessary to determine the clinical significance of elevated scatter factor titers in transitional cell carcinoma of the bladder.

Carcinoma, Transitional Cell↗

PubMed1997

Scatter factor stimulates tumor growth and tumor angiogenesis in human breast cancers in the mammary fat pads of nude mice.

Scatter factor (SF) (also known as hepatocyte growth factor) is a plasminogen-related growth factor that induces tumor cell motility, invasion, and angiogenesis. Its receptor is a tyrosine kinase encoded by c-met, a protooncogene. Human breast cancer cells express SF and c-met in vivo; but human breast cancer cell lines do not produce SF in vitro. To determine whether SF can modulate the in vivo growth of human breast cancers within a natural mammary environment, we studied the orthotopic growth of SF-transfected (SF+) versus control (SF-) clones of MDAMB231 human mammary carcinoma cells in the mammary fat pads of athymic nude mice. SF+ clones expressed SF mRNA and produced very high titers of SF protein, whereas SF- clones did not express SF mRNA or produce detectable SF protein. Two SF+ clones (21 and 29) showed significantly increased tumor growth rates, reaching 3- to 4-fold larger primary tumor volumes and weights by time of killing (p < 0.001), as well as higher rates of axillary lymph node metastasis (p < 0.02), as compared with two SF- clones (32 and 34). In contrast, in vitro proliferation rates, two-dimensional colony formation, and soft agar colony formation were no greater in SF+ than in SF- clones. We performed further studies to investigate the discrepancy between the in vivo and in vitro growth results. Tumor extracts from SF+ clone (21 + 29) tumors had 50-fold higher SF content than did SF- clone (32 + 34) tumors, confirming high-level SF expression in vivo in SF+ tumors. Immunostaining of tumor sections for proliferating cell nuclear antigen revealed only a modest increase in the proportion of cycling cells in SF+ versus SF- tumors (70% versus 60%, respectively). The terminal deoxytransferase-labeling index was equally low (approximately 1%) in SF+ and SF- tumors, suggesting that apoptosis was not responsible for the slower growth of SF- tumors. However, SF+ tumors had significantly higher tumor microvessel densities than SF- tumors (p < 0.001). Moreover, there were much higher titers of chemotactic activity for microvascular endothelial cells in cell-conditioned media and primary tumor extracts from SF+ clones as compared with SF- clones. As demonstrated using the rat cornea assay, there was more angiogenic activity in SF+ tumor extracts than in SF- extracts. The increased chemotactic and angiogenic activities in SF+ tumor extracts were not explained by secondary alterations in the content of the angiogenic mediator, vascular endothelial growth factor, or the antiangiogenic glycoprotein, thrombospondin-1; and those activities were neutralized using an anti-SF monoclonal antibody. These findings suggest that SF promotes the orthotopic growth of human breast cancers, at least in part, by stimulating tumor angiogenesis.

Adipose Tissue↗

PubMed1997

Oxidative metabolism of clarithromycin in the presence of human liver microsomes. Major role for the cytochrome P4503A (CYP3A) subfamily.

In vitro studies were conducted to identify the hepatic cytochrome P450 (CYP) protein(s) involved in the oxidative metabolism of [14C]clarithromycin (CLAR) in the presence of native human liver microsomes. The identity of the two major CLAR metabolites present in microsome incubates, 14-(R)-hydroxy-CLAR and N-desmethyl-CLAR, was confirmed by MS. Over the CLAR concentration range of 1.0-140 microM, the rate of CLAR 14-(R)-hydroxylation (KM = 48 +/- 17.7 microM; Vmax = 206 +/- 76 pmol/min/mg protein; Vmax/KM = 4.2 +/- 0.21 microliters/min/mg; mean +/- SD, N = 3 livers) and N-demethylation (KM = 59.1 +/- 24.0 microM; Vmax = 189 +/- 52.0 pmol/min/mg protein; Vmax/KM = 3.3 +/- 0.53 microliters/min/mg) conformed to monophasic (saturable) Michaelis-Menten kinetics and was highly correlated (r = 0.90-0.92; p < 0.001; N = 11) with CYP3A-selective erythromycin N-demethylase activity. Ketoconazole (< or = 2.0 microM) or troleadomycin, CYP3A-selective inhibitors, markedly decreased (> or = 99%) the formation of both metabolites, whereas inhibitors selective of other CYP forms were relatively ineffective (< or = 10% inhibition). In agreement with chemical inhibitor studies, CLAR metabolism was only detectable with human B-lymphoblastoid microsomes containing cDNA-expressed CYP3A4 (vs. CYP2C19, CYP2C9, CYP2D6, CYP1A2, CYP2E1, or CYP2A6). Furthermore, the apparent KM characterizing the 14-(R)-hydroxylation and N-demethylation of CLAR in the presence of insect cell microsomes containing cDNA-expressed CYP3A4 (KM = 18-63 microM) was similar to that obtained with native human liver microsomes. Based on the results of this study, it is concluded that the 14-(R)-hydroxylation and N-demethylation of CLAR is primarily mediated by one or more members of the human liver CYP3A subfamily.

Anti-Bacterial Agents↗

PubMed1997

[O-ethyl 14C]phenacetin O-deethylase activity in human liver microsomes.

The activity of human liver microsomal cytochrome P450 1A2 (CYP1A2) is readily estimated by following the O-deethylation of [O-ethyl 14C]phenacetin (PODase). The basis of the assay is the quantitative measurement of [14C]acetaldehyde, remaining in the supernatant of assay incubates, after extraction of unmetabolized [O-ethyl 14C]phenacetin with charcoal. In the presence of native human liver microsomes (K(m) = 54 +/- 27 microM; V(max) = 14 +/- 2.3 nmol/hr/mg; mean +/- SD; N = 3 different livers) and human B-lymphoblastoid cell microsomes containing cDNA-expressed CYP1A2 (K(m) = 46 microM; V(max) = 55 nmol/hr/nmol CYP), PODase activity conformed to monophasic Michaelis-Menten kinetics. Furthermore, PODase activity in a panel of microsomes prepared from a series of human livers was significantly correlated (r = 0.91; p < 0.001; N = 11) with CYP1A2-selective 7-ethoxyresorufin O-deethylase activity, and was markedly inhibited (> or = 92%) by furafylline (FURA, IC50 = 0.4 microM) and 7,8-benzoflavone (ANF, IC50 = 0.1 microM), two well known CYP1A2 inhibitors. Inhibitors selective for other forms of CYP (e.g. CYP3A, CYP2C, CYP2D6, CYP2E1) elicited a marginal effect (< or = 17% inhibition) at relatively high concentrations (> or = 10.K(i)). It is concluded that the inhibition of human liver microsomal CYP1A2 activity can be readily determined by using a charcoal-based radiometric method employing [O-ethyl 14C]phenacetin as substrate.

Carbon Radioisotopes↗

PubMed1997

The relationship between gut-derived endotoxemia and tumor necrosis factor, neopterin: experimental and clinical studies.

OBJECTIVE: To determine the relationship between gut-derived endotoxemia and tumor necrosis factor (TNF), neopterin/biopterin formation following hemorrhage, trauma and burns. METHODS: Rats were subjected to hemorrhagic shock (30 mmHg, 90-180 min) and 40% III degrees thermal injury. Circulating endotoxin, TNF, biopterin levels and liver TNF mRNA expression were measured in animals following acute insults. Also, the subjects of this study included 35 patients with burn size greater than 30%, and 25 patients with multiple injuries (n = 18) and major surgery (n = 7). RESULTS: It was found that significant portal and systemic endotoxemia took place in the control animal after hemorrhagic shock and thermal injury, but almost not in the animals that treated by measures aiming at controlling endotoxin/bacteria translocation, including polymyxin B, monoclonal antibody against core lipopolysaccharide, and selective decontamination of the digestive tract (SDD). Concomitantly, hemorrhage and thermal injury resulted in significant increases in systemic plasma TNF level together with tissue TNF mRNA expression, which were associated with the initial appearance of endotoxin in portal vein. However, anti-endotoxin treatment markedly decreased circulating TNF level as well as peak TNF mRNA expression caused by acute insults. There were also lower serum biopterin values in the SDD-treated group as compared with the control group on day 5 postburn. On the other hand, the results showed that the amounts of plasma endotoxin in patients increased during the early stages following major burns, which was significantly correlated with plasma TNF levels, particularly in patients who developed sepsis and multiple organ failure. Although the presence of early endotoxemia did not influence the alterations in serum neopterin, patients with endotoxemia had much higher neopterin values than those who showed no endotoxemia from the second week onward. CONCLUSION: These results suggest that gut-derived endotoxemia could account, at least in part, for the inflammatory mediators formation and release, which might be involved in the pathogenesis of sepsis and multiple organ dysfunction following severe hemorrhage, trauma and burns.

Adult↗

PubMed1997

[Multifactor analysis on proliferative retinopathy in elderly patients with type II diabetes].

UNLABELLED: To evaluate the importance of different risk factors and their mutual interactions in the occurrence of proliferative diabetic retinopathy (PDR) in the elderly patients with type II diabetes, an investigation was carried out on the relationship between PDR and various risk factors in 98 elderly cases of type II diabetes by way of stepwise regression analysis method. Results showed: (1) the average course of PDR was longer than that of diabetic patients without diabetic retinopathy (P < 0.01). Patients with PDR had higher values of BUN (P < 0.01), Cr(P < 0.05), GHbA1c(P < 0.01) and TC(P < 0.01) than those without retinopathy. Patients with PDR had higher coincidences of hypertension, coronary artery disease, proteinuria and urine microalbumin than those without retionpathy (P < 0.01) respectively, (2) multifactor analysis showed that BUN was one of the most important variables positively correlated with PDR, second to it in orders were the course of diabetes militus, GHbA1c, hypertention, proteinuria and urine microalbumin and TC. CONCLUSION: To prevent the occurrence and progress of PDR in the elderly patients with type II diabetes, it is important not only to control blood sugar, but to control blood pressure and relieve renal damage.

Aged↗

PubMed1996

Mouse RNA polymerase I 16-kDa subunit able to associate with 40-kDa subunit is a homolog of yeast AC19 subunit of RNA polymerases I and III.

We have previously isolated a mouse RPA40 (mRPA40) cDNA encoding the 40-kDa subunit of mouse RNA polymerase I and demonstrated that mRPA40 is a mouse homolog of the yeast subunit AC40, which is a subunit of RNA polymerases I and III, having a limited homology to bacterial RNA polymerase subunit alpha (Song, C. Z., Hanada, K., Yano, K., Maeda, Y., Yamamoto, K., and Muramatsu, M. (1994) J. Biol. Chem. 269, 26976-26981). In an extension of the study we have now cloned mouse RPA16 (mRPA16) cDNA encoding the 16-kDa subunit of mouse RNA polymerase I by a yeast two-hybrid system using mRPA40 as a bait. The deduced amino acid sequence shows 45% identity to the yeast subunit AC19 of RNA polymerases I and III, known to associate with AC40, and a local similarity to bacterial alpha subunit. We have shown that mRPA40 mutants failed to interact with mRPA16 and that neither mRPA16 nor mRPA40 can interact by itself in the yeast two-hybrid system. These results suggest that higher eukaryotic RNA polymerase I conserves two distinct alpha-related subunits that function to associate with each other in an early stage of RNA polymerase I assembly.

Amino Acid Sequence↗

PubMed1996

Dynamic structure of the calmodulin-binding domain of the plasma membrane Ca-ATPase in native erythrocyte ghost membranes.

We have used frequency-domain fluorescence resonance energy transfer (FRET) and anisotropy measurements to identify the structural properties of wheat germ calmodulin (CaM) bound to either the plasma membrane Ca-ATPase (PM-Ca-ATPase) in native erythrocyte ghost membranes or a peptide (C25W) that has an identical sequence to the CaM-binding domain on the PM-Ca-ATPase. Cross-linking experiments using benzophenone labeled CaM in conjunction with immunoblots using antibodies specific for either CaM or the PM-Ca-ATPase indicate that one molecule of CaM selectively binds one PM-Ca-ATPase polypeptide chain in native erythrocyte ghost membranes. There are no other proteins in the erythrocyte membrane that bind CaM with high affinity, permitting the measurement of the structural properties of CaM bound to the PM-Ca-ATPase in native erythrocyte ghost membranes. FRET measurements between the fluorophore pyrene maleimide (PMal) located at Cys27 in calcium binding loop I and nitrotyrosine139 in calcium binding loop IV on wheat germ CaM indicate that the average spatial separation and conformational heterogeneity associated with the two opposing globular domains of CaM are virtually identical upon CaM binding to either the PM-Ca-ATPase or C25W. Measurements of the solvent accessibility and segmental rotational dynamics of PMal-CaM bound to either the PM-Ca-ATPase or C25W further indicate that the local environment around the pyrene label located at Cys27 is very similar. However, the overall rotational dynamics of CaM bound to the PM-Ca-ATPase is much slower (phi 2 = 83 +/- 14 ns) than observed when CaM binds C25W (phi 2 = 10.3 +/- 0.5 ns). This implies that CaM is tightly associated with the CaM-binding domain of the PM-Ca-ATPase and that the observed rotational motion of pyrenylmaleimide labeled CaM is characteristic of the global motion of the CaM-binding domain on the PM-Ca-ATPase. The similar conformational heterogeneity and local environment of CaM bound to either the PM-Ca-ATPase or C25W indicates that CaM binds to a contiguous sequence of amino acids on the Ca-ATPase that are analogous to C25W and that there are no significant interactions with other structural elements within the PM-Ca-ATPase. The rate of rotational motion associated with CaM bound to the PM-Ca-ATPase is consistent with hydrodynamic calculation in which the calmodulin-binding domain located at the carboxyl-terminus of the PM-Ca-ATPase has a stable and defined tertiary structure that is independent of the other cytoplasmic domains of the PM-Ca-ATPase.

Animals↗

PubMed1996

Variable conformation and dynamics of calmodulin complexed with peptides derived from the autoinhibitory domains of target proteins.

Calcium-saturated calmodulin (CaM) can bind and activate many target proteins through the direct association with the respective autoinhibitory domains. The CaM binding sequences within the autoinhibitory domains of these proteins have little sequence homology, and the mechanisms associated with CaM's ability to recognize and productively bind with these variable sequences is unclear. Common structural features of CaM bound to five peptides that are homologous to the autoinhibitory domains of smooth muscle myosin light chain kinase, CaM-dependent protein kinase II alpha, the plasma membrane Ca-ATPase, a MARCKS homolog, and glycogen phosphorylase kinase were assessed using frequency-domain fluorescence spectroscopy. In addition, the structural features of CaM complexed with the peptide melittin was also considered. We observe similar decreases in the average fluorescence lifetime and similar increases in the solvent accessibility of N-(1-pyrenyl)maleimide (PM) bound at Cys27 in calcium binding loop I in the amino terminal domain of CaM upon association with all six target peptides. Likewise, using fluorescence resonance energy transfer to measure the spatial separation between the opposing globular domains in CaM, we observe a similar spatial separation between the opposing globular domains of CaM bound to all six peptides. This indicates that CaM undergoes comparable structural changes upon association with all six target peptides. However, there are significant differences in the observed lifetime, solvent accessibility, correlation time associated with the segmented rotational motion of PM-CaM, and in the spatial separation between the opposing globular domains in CaM upon association with the individual target peptides, which indicates that CaM adopts a different tertiary structure that is dependent on the structural features of the bound target peptide. The correlation times associated with the overall hydrodynamic properties of CaM complexed with all six peptides are nearly identical (phi 2 approximately 10.6 +/- 0.4 ns) and are consistent with the known dimensions of CaM complexed to a peptide homologous to the CaM binding sequence of CaM-dependent protein kinase II alpha. Therefore, while these results are consistent with a common binding mechanism between CaM and all six target peptides, they indicate that the binding domains of CaM adopt different tertiary structures that allow them to bind with the variable sequences found in the autoinhibitory domains of target proteins with high affinity.

Amino Acid Sequence↗

PubMed1996

Ca2+ transients associated with openings of inositol trisphosphate-gated channels in Xenopus oocytes.

1. The mechanisms underlying inositol 1,4,5-trisphosphate (InsP3)-induced Ca2+ liberation were studied in Xenopus oocytes by using scanning and stationary-point confocal fluorescence microscopy to record Ca2+ signals evoked by photorelease of InsP3 from a caged precursor. 2. Fluorescence measurements from confocal images showed that increasing [InsP3] evoked three distinct modes of Ca2+ liberation: a diffuse 'pacemaker' signal, localized transient puffs, and propagating waves. Peak free Ca2+ concentrations during waves and puffs (respectively, 2-5 microM and 100-200 nM) varied only slightly with [InsP3], whereas the pacemaker amplitude varied over a wider range (at least 1-30 nM Ca2+). 3. The improved resolution provided by confocal point recording revealed discontinuous Ca2+ 'blips' during pacemaker release. These events were resolved only at particular locations and had time courses similar to the puffs (rise, approximately 50 ms; decay, a few hundred milliseconds) but with amplitudes one-fifth or less of puff amplitudes. 4. We conclude that blips may arise through opening of single InsP3-gated channels, whereas puffs reflect the concerted opening of several clustered channels due to local regenerative feedback by Ca2+.

Animals↗

PubMed1996

Oxidative modification of a carboxyl-terminal vicinal methionine in calmodulin by hydrogen peroxide inhibits calmodulin-dependent activation of the plasma membrane Ca-ATPase.

In order to investigate the possibility that calmodulin (CaM) may be a principal target of reactive oxygen species (ROS) produced under conditions of oxidative stress, we have examined wheat germ CaM for the presence of highly reactive sites that correlate with the loss of function. Using reversed-phase HPLC and FAB mass spectrometry after proteolytic digestion, we have identified the sites of modification by hydrogen peroxide. We find that one of the vicinal methionines (i.e., Met146 or Met147) near the C-terminus of CaM is selectively oxidized. The ability of CaM to bind and to activate the plasma membrane (PM)-Ca-ATPase from erythrocytes was measured. There is a 30-fold decrease in the calcium affinity of oxidatively modified CaM. While there is a little change in the binding constant between the carboxyl-terminal domain of calcium-saturated CaM and a peptide homologous to the autoinhibitory sequence of the PM-Ca-ATPase, we find that there is a 9-fold reduction in the affinity of the amino-terminal domain of CaM with respect to the ability to bind target peptides. The extent of oxidative modification to one of the vicinal methionines near the carboxyl-terminal domain correlates with the loss of CaM-dependent activation of the PM-Ca-ATPase. The presence of oxidatively modified CaM prevents native CaM from activating the PM-Ca-ATPase, indicating that the oxidatively modified CaM binds to the autoinhibitory sequence on the Ca-ATPase in an altered nonproductive conformation. We suggest that the functional sensitivity of CaM to the oxidation of one of the C-terminal vicinal methionines permits CAM to serve a regulatory role in modulating cellular metabolism under conditions of oxidative stress. The predominant oxidation of a methionine near the carboxyl terminal of CaM is rationalized in terms of the enhanced solvent accessibility of these vicinal methionines.

Amino Acid Sequence↗

PubMed1996

Decreased plasma membrane calcium transport activity in aging brain.

We have assessed the functional properties of both calmodulin (CaM) and the plasma membrane Ca(2+)-ATPase in brains of young, middle aged, and old Fisher 344 rats. Under optimal conditions of saturating Ca2+ and ATP, the CaM-activated Ca(2+)-ATPase activity was decreased with increasing age, particularly when CaM isolated from the brains of aged rats was used to stimulate the enzyme. In the case of CaM, structural modifications within the primary sequence of the protein from aged brains were identified. We found that during normal biological aging approximately 6 methionine residues were modified to their corresonding sulfoxide per CaM, and no other amino acids were modified. Some aspects of the age-related decline in the effectiveness of CaM as an activator of Ca(2+)-ATPase could be simulated using a range of reactive oxygen species (including hydrogen peroxide and oxoperoxynitrite) and, in the latter case, the extent of oxidative modification of specific methionine residues was directly related to their surface accessibility. The pattern of oxidative modification of the methionines in the aged CaM was less straightforward, though both in vitro oxidation of CaM and aging within the brain markedly decreased the functional properties of this important Ca(2+)-regulating protein.

Aging↗

PubMed1996

Fast kinetics of calcium liberation induced in Xenopus oocytes by photoreleased inositol trisphosphate.

Inositol 1,4,5-trisphosphate (InsP3) acts on intracellular receptors to cause liberation of Ca2+ ions into the cytosol as repetitive spikes and propagating waves. We studied the processes underlying this regenerative release of Ca2+ by monitoring with high resolution the kinetics of Ca2+ flux evoked in Xenopus oocytes by flash photolysis of caged InsP3. Confocal microfluorimetry was used to monitor intracellular free [Ca2+] from femtoliter volumes within the cell, and the underlying Ca2+ flux was then derived from the rate of increase of the fluorescence signals. A threshold amount of InsP3 had to be photoreleased to evoke any appreciable Ca2+ signal, and the amount of liberated Ca2+ then increased only approximately fourfold with maximal stimulation, whereas the peak rate of increase of Ca2+ varied over a range of nearly 20-fold, reaching a maximum of approximately 150 microMs-1. Ca2+ flux increased as a first-order function of [InsP3]. Indicating a lack of cooperativity in channel opening, and was half-maximal with stimuli approximately 10 times threshold. After a brief photolysis flash, Ca2+ efflux began after a quiescent latent period that shortened from several hundred milliseconds with near-threshold stimuli to 25 ms with maximal flashes. This delay could not be explained by an initial "foot" of Ca2+ increasing toward a threshold at which regenerative release was triggered, and the onset of release seemed too abrupt to be accounted for by multiple sequential steps involved in channel opening. Ca2+ efflux increased to a maximum after the latent period in a time that reduced from > 100 ms to approximately 8 ms with increasing [InsP3] and subsequently declined along a two-exponential time course: a rapid fall with a time constant shortening from > 100 ms to approximately 25 ms with increasing [InsP3], followed by a much smaller fail persisting for several seconds. The results are discussed in terms of a model in which InsP3 receptors must undergo a slow transition after binding InsP3 before they can be activated by cytosolic Ca2+ acting as a co-agonist. Positive feedback by liberated Ca2+ ions then leads to a rapid increase in efflux to a maximal rate set by the proportion of receptors binding InsP3. Subsequently, Ca2+ efflux terminates because of a slower inhibitory action of cytosolic Ca2+ on gating of InsP3 receptor-channels.

Animals↗

PubMed1996

Elementary events of InsP3-induced Ca2+ liberation in Xenopus oocytes: hot spots, puffs and blips.

Liberation of sequestered Ca2+ ions in Xenopus oocytes by the second messenger inositol 1,4,5-trisphosphate (InP3) occurs from functionally discrete sites, which are spaced at intervals of several microns and probably represent clusterings of InsP3 receptor/channels (InsP3R) in the endoplasmic reticulum. As well as requiring InsP3, opening of release channels is regulated by dual positive and negative feedback by cytosolic Ca2+, leading to regenerative Ca2+ transients. Because the sensitivity of this process is determined by [InsP3], the ability of Ca2+ ions diffusing from one location to activate increasingly distant InsP3R is enhanced by increasing [InsP3]. Together with the spatial distribution of receptors, this results in generation of a hierarchy of Ca2+ release events, which may involve individual InsP3R (Ca2+ 'blips'), concerted activation of several receptors within a single release site (Ca2+ 'puffs'), and recruitment of successive sites by Ca2+ diffusing over micron distances to produce propagating Ca2+ waves. Thus, Ca2+ signalling in the oocyte is organized as at least two sizes of elemental 'building blocks'; highly localized Ca2+ transients that arise autonomously and stochastically from discrete sites at low [InsP3], but which become coordinated at higher [InsP3] to produce global Ca2+ responses.

Animals↗

PubMed1996

Treatment of Mycobacterium avium complex infection: do the results of in vitro susceptibility tests predict therapeutic outcome in humans?

The ability of various in vitro methods of antibiotic susceptibility testing to predict therapeutic outcome in patients infected with Mycobacterium avium complex (MAC) was evaluated. Pretreatment bloodstream MAC isolates from 38 patients with AIDS, previously treated in a randomized fashion with either ethambutol, rifampin, or clofazimine, were tested by three conventional methods using broth or agar, as well as by cocultivation with macrophages. The results obtained with each method were compared with the quantitatively determined bacteriologic response to the administration of the single agent in humans. None of the conventional in vitro susceptibility methods was predictive of therapeutic outcome, while the results of cocultivation with macrophages were of moderate predictive value. The positive predictive value of a response in humans based on a response in macrophages (defined by > or = to 1.0 log reduction in baseline colony counts after 5 days of treatment) was 74%. The negative predictive value was 82%.

AIDS-Related Opportunistic Infections↗

PubMed1996

Treatment of Myocardium avium complex infection: does the beige mouse model predict therapeutic outcome in humans?

To determine the predictive value of a standard murine model in the treatment of disseminated Myocardium avium complex (MAC) infection, beige mice were infected with MAC strains isolated from human immunodeficiency virus-infected patients and treated with the same antibiotic (ethambutol, clofazimine, or rifampin) that had been administered to the subject from whom that strain had been recovered. While ethambutol had the greatest bacteriologic efficacy in humans (mean decrease +/-SD, 1.0+/-0.5 log 10 cfu/mL of blood), clofazimine had the greatest bacteriostatic efficacy in mice (mean decrease +/- SD, 2.8 +/- 0.7 log(10) cfu/g of tissue). A linear correlation was not observed between bacteriostatic activity in mouse liver or spleen and the degree of bacteriologic response in humans (P > or = to .1). Odds ratios for a response in humans based on a bacteriologic response in mice were not significant for each agent (P > or = to .1, all cases).

AIDS-Related Opportunistic Infections↗

PubMed1996

Multiple organ injuries and failures caused by shock and reperfusion after gunshot wounds.

Experiments were performed to observe the changes of multiple system organ failure (MSOF) and gut barrier function caused by shock and reperfusion after gunshot wounds. Eighteen dogs were divided randomly into two groups. In the experimental group, the dogs were subjected to 60 minutes of shock (40mm Hg), followed by reinfusion of shed blood after hindlimb gunshot wounds. In the control group, the dogs experienced pure gunshot wounds without shock and reperfusion. The results showed that dogs in the experimental group developed multiple system organ injuries or failures compared with the control group. The levels of malondialdehyde (MDA) values in plasma were significantly elevated in the experimental group when compared with preinjury and the control group. Gut flora disorder, bacillus intestinalis overgrowth, and bacterial translocation occurred in the experimental group. The pathological results support the gut barrier function injury. The results indicated that pure gunshot wounds do not easily injure gut barrier function and produce MSOF. Gunshot wounds with shock and reperfusion are capable of causing gut flora disorder, bacillus intestinalis overgrowth, and lead to bacterial translocation, furthermore causing MSOF. Although fluid resuscitation is a potential treatment modality, pathogenically, it can lead to MSOF.

Animals↗

PubMed1996

Expression of chicken troponin T isoforms in cultured muscle cells.

Cells prepared from chicken skeletal muscles of different developmental stages were cultured to study their troponin T isoform expression, using antisera specific to the fast- and slow-muscle-type isoforms. We found that the cultured myogenic cells from chickens and chick embryos were classified into two types, fast type and fast/slow type in which fast- and slow-muscle-type isoforms were coexpressed. Cells expressing only slow-muscle-type troponin T isoforms could not be found. Most cells prepared from pectoralis major (fast muscle) and gastrocnemius (mixed muscle) of 11-day old embryos belonged to the latter, with only a small fraction belonging to the former. The percentage of fast type cells in those cells prepared from pectoralis major increased along development to over 90% by the 17th day of incubation, while, in the cells prepared from gastrocnemius, it reached a plateau of 30-40% by the 13th day of incubation. All the cells from anterior latissimus dorsi (slow muscle) belonged to the fast/slow type. Ratios of these two types of muscle cells varied depending on their origins and stages. The in vitro expression of troponin T isoforms was different from the in vivo expression, and each muscle seems to be determined differently in the composition of cell types during the developmental course.

Animals↗