The cytoplasmic membrane is a primary target for the staphylocidal action of thrombin-induced platelet microbicidal protein

Thrombin-induced platelet microbicidal protein (tPMP-1) is a small, cationic peptide released from rabbit platelets exposed to thrombin in vitro. tPMP-1 is microbicidal against a broad spectrum of bloodstream pathogens, including Staphylococcus aureus. Preliminary evidence suggests that tPMP-1 targets and disrupts the staphylococcal cytoplasmic membrane. However, it is not clear if the cytoplasmic membrane is a direct or indirect target of tPMP-1. Therefore, we assessed the in vitro activity of tPMP-1 versus protoplasts prepared from logarithmic-phase (LOG) or stationary-phase (STAT) cells of the genetically related S. aureus strains 19S and 19R (tPMP-1 susceptible and resistant, respectively). Protoplasts exposed to tPMP-1 (2 microg/ml) for 2 h at 37 degrees C were monitored for lysis (decrease in optical density at 420 nm) and ultrastructural alterations (by transmission electron microscopy [TEM]). Exposure to tPMP-1 resulted in substantial lysis of LOG but not STAT protoplasts of 19S, coinciding with protoplast membrane disruption observed by TEM. Thus, it appears that tPMP-1-induced membrane damage is influenced by the bacterial growth phase but is independent of the staphylococcal cell wall. In contrast to 19S, neither LOG nor STAT protoplasts of 19R were lysed by tPMP-1. tPMP-1-induced membrane damage was further characterized with anionic planar lipid bilayers subjected to various trans-negative voltages. tPMP-1 increased conductance across bilayers at -90 mV but not at -30 mV. Once initiated, a reduction in voltage from -90 to -30 mV diminished conductance magnitude but did not eliminate tPMP-1-mediated membrane permeabilization. Therefore, tPMP-1 appears to directly target the staphylococcal cytoplasmic membrane as a primary event in its mechanism of action. Specifically, tPMP-1 likely leads to staphylococcal death, at least in part by permeabilizing the bacterial membrane in a voltage-dependent manner.     

Rapid shift in peak melatonin secretion associated with improved performance in short shift work schedule

OBJECTIVE: To describe the changes in the characteristics of human immunodeficiency virus (HIV)-related deaths in children with perinatally acquired infection. METHODS: A retrospective review of all deaths that occurred in HIV-infected children managed at The New York Hospital-Program for Children with AIDS during a 7-year period from January, 1990, to December, 1996. Differences in the characteristics at death between 15 children who died in 1990 and 10 children who died in 1996 were analyzed. RESULTS: Fifty-eight deaths in our cohort of HIV-infected children were identified during the 7-year period. The mean age at death was 4.43 years. Sixty-nine percent of children were black, 55% were male and 94% were receiving Medicaid. The mean weight/age Z score was -3.9 and the mean CD4 index was 0.067 with 65% having <50 CD4 cells/microl at the time of death (TOD). The most common organ/organ systems to be involved at the TOD were lung (78%) and central nervous system (61%). Mycobacterium avium complex (MAC) was the most common isolate at the TOD (26%) followed by Pneumocystis carinii (20%) and Pseudomonas aeruginosa (17%). The leading non-infectious cause of death was cardiac failure (9%). Comparison of the characteristics at the TOD between 1990 and 1996 revealed significant differences in mean age (2.1 vs. 9.2 years, P < 0.0001), mean CD4 count index (0.18 vs. 0.02, P < 0.03), mean number of organ/organ system involvement (3.9 vs. 5.9, P < 0.05), percent receiving antiretroviral therapy (33% vs. 70%, P < 0.02), mean number of years receiving antiretroviral therapy (0.88 vs. 3.86 years, P < 0.01), percent receiving P. carinii pneumonia prophylaxis (27% vs. 100%, P < 0.001), percent receiving MAC prophylaxis/therapy (0% vs. 100%, P < 0.0001), and cause of death from P. carinii pneumonia (53% vs. 0%, P < 0.01). CONCLUSIONS: Compared with children who died in 1990, HIV-infected children who died in 1996 were significantly older, more lymphopenic and more likely to have a greater number of organ system involvements and to have received antiviral therapy and antimicrobial prophylaxis. In 1996 no child died of P. carinii pneumonia. In 1996 MAC and P. aeruginosa were the two most important opportunistic infections causing death. These changes in the characteristics at death will warrant review of resources used in treating these children and may be critical in advising parents and care givers about the prognosis of this chronic infection.     

A cohort study of pregnancy outcome after amniocentesis in twin pregnancy

We conducted a retrospective cohort study to assess the risk of amniocentesis in twin pregnancy for adverse outcomes. The study base consisted of women who had an amniocentesis performed during twin pregnancy and a comparison representative sample of women who carried a twin pregnancy, but did not have invasive prenatal diagnosis. The 227 women in each of the exposed and non-exposed groups were residents of the state of New South Wales, Australia, over the period 1980-92, and were matched on maternal age and period of the infant's birth. Nearly 10% of twin pregnancies among the women having an amniocentesis were affected by a stillbirth, and the stillbirth rate among exposed fetuses (5.3%) was nearly twice as high as among non-exposed fetuses (3.1%). After adjustment for confounding and excluding abnormalities, there was a non-significant elevated relative risk of stillbirth after exposure to amniocentesis. The analysis by type of amniocentesis (with and without methylene blue dye) was limited by small numbers, but the burden of risk was primarily among women who had dye exposure during amniocentesis (relative risk = 3.64, 95% confidence interval = 1.15, 11.48). This increase remained after adjusting for confounding, although the confidence interval was wide. In conclusion, we were unable to establish with certainty whether an increased risk of stillbirth could be ruled out among women who had any type of amniocentesis in twin pregnancy.     

The NH2 terminus of titin spans the Z-disc: its interaction with a novel 19-kD ligand (T-cap) is required for sarcomeric integrity

Titin is a giant elastic protein in vertebrate striated muscles with an unprecedented molecular mass of 3-4 megadaltons. Single molecules of titin extend from the Z-line to the M-line. Here, we define the molecular layout of titin within the Z-line; the most NH2-terminal 30 kD of titin is located at the periphery of the Z-line at the border of the adjacent sarcomere, whereas the subsequent 60 kD of titin spans the entire width of the Z-line. In vitro binding studies reveal that mammalian titins have at least four potential binding sites for alpha-actinin within their Z-line spanning region. Titin filaments may specify Z-line width and internal structure by varying the length of their NH2-terminal overlap and number of alpha-actinin binding sites that serve to cross-link the titin and thin filaments. Furthermore, we demonstrate that the NH2-terminal titin Ig repeats Z1 and Z2 in the periphery of the Z-line bind to a novel 19-kD protein, referred to as titin-cap. Using dominant-negative approaches in cardiac myocytes, both the titin Z1-Z2 domains and titin-cap are shown to be required for the structural integrity of sarcomeres, suggesting that their interaction is critical in titin filament-regulated sarcomeric assembly.     

Self-care behaviors, self-efficacy, and social support effect on the glycemic control of patients newly diagnosed with non-insulin-dependent diabetes mellitus

The purpose of this study was to explore the glycemic control and influencing factors in outpatients newly diagnosed with non-insulin-dependent diabetes mellitus (NIDDM). By purposeful sampling, data were collected from 130 outpatients with NIDDM at one medical center in Kaohsiung. The results indicated: (1) the mean value for HbA1C was 7.12%; and 63.1% of the patients belonged to moderate to well controlled group; (2) male patient's HbA1C value was significantly lower than female patient's; patients with no religious belief also had a lower HbA1C value than patients with a religious background; (3) there were strongly negative correlations between self-care behaviors, social support, and self-efficacy and HbA1C; (4) using a multiple stepwise regression analysis, religious belief and self-care behaviors were found to explain 10.9% variance of HbA1C level. The results of this study could be used as a reference for diabetes health education program.     

Autonomy and its implications for palliative care: a northern European perspective

The aim of this work was to analyze the use of some renographic parameters derived from the 99mTc-MAG3 renogram to classify post-transplantion studies in two groups: Normal functioning grafts (NFG) and delayed grafts function (DGF) which included Acute Tubular Necrosis (ATN) and Acute Rejection (AR). The analysis included data from 38 NFG, 33 ATN and 23 AR. The parameters calculated were: mean transit time (MTT), the Initial Uptake (IU) and the maximum activity (MA) of the renogram. The results obtained from this analysis show that ln(IU) is the best parameter to classify individuals in either group and that the use of MA or the MTT does not improve the results. Using a cutoff point of ln(IU) = 0.28 and a prevalence of 25% for DGF, the estimated predictive values were 92% for DGF and 99% for NFG.     

Inhibition of gastric emptying and intestinal transit by amphetamine through a mechanism involving an increased secretion of CCK in male rats

1. The effect of amphetamine on gastrointestinal (GI) transit and the plasma levels of cholecystokinin (CCK) were studied in male rats. 2. Gastric emptying was inhibited both acutely and chronically by the administration of amphetamine. GI transit was decreased by the acute administration of amphetamine but not affected by the chronic administration of amphetamine. 3. Plasma CCK levels were increased dose-dependently by amphetamine. 4. Proglumide, a CCK receptor antagonist, prevented amphetamine-induced inhibition of gastric emptying and the decrease in GI transit in male rats. 5. The selective CCK(A) receptor antagonist, lorglumide, dose-dependently attenuated the amphetamine-induced inhibition of gastric emptying in male rats. In contrast, the selective CCK(B) receptor antagonist, PD 135,158, did not reverse the effect of amphetamine on gastric emptying. 6. Both lorglumide and PD 135,158 reversed the inhibitory effect of amphetamine on GI transit in male rats. 7. These results suggest that amphetamine-induced inhibition of gastric emptying and intestinal transit is due in part to a mechanism associated with the hypersecretion of endogenous CCK.     

The function of adenosylcobalamin in the mechanism of ribonucleoside triphosphate reductase from Lactobacillus leichmannii

BACKGROUND: Tumour necrosis factor alpha (TNF-alpha) is a proinflammatory cytokine found in abundance in diseased intestine. AIMS: The T cell production of TNF-alpha and the impact of this cytokine on intestinal T cell proliferation, migration, and cytotoxicity were studied. METHODS: Intestinal lymphocytes from normal jejunum were used. TNF-alpha production in culture supernates was measured by enzyme linked immunosorbent assay (ELISA). Lymphocyte proliferation was measured using 3H thymidine uptake; migration, using transwell chambers; and cytotoxicity of HT-29 colon cancer cells, using the chromium-51 release assay. RESULTS: TNF-alpha was produced mainly by the CD8+ T cells in the intraepithelial lymphocytes (IEL) and the CD4+ T cells in the lamina propria lymphocytes in response to CD2 stimulation: 478 (94) and 782 (136) pg/ml, respectively. TNF-alpha (1 ng/ml or greater) augmented proliferation of IEL in response to interleukin 2 (IL-2), IL-7, or antibody to CD3 due to increased activation that did not involve IL-2 production or receptor generation. Conversely, antibody to TNF-alpha reduced IEL proliferation in response to IL-2 or IL-7. TNF-alpha also induced calcium mobilisation and chemokinesis (by 2.8 (0.5) fold over spontaneous migration). TNF-alpha had no effect on lymphokine activated killer cell activity. CONCLUSIONS: TNF-alpha increases the proliferation and migration of IEL, which may expand their number in the epithelium.     

Effect of mannitol crystallization on the stability and aerosol performance of a spray-dried pharmaceutical protein, recombinant humanized anti-IgE monoclonal antibody

We have examined the stability and aerosol performance of the pharmaceutical protein recombinant humanized anti-IgE monoclonal antibody (rhuMAbE25) spray dried with mannitol. The aerosol performance was measured by the fine particle fraction (FPF), and stability was assessed by the formation of soluble aggregates. When mannitol was added to the spray-dried rhuMAbE25 formulation, its ability to stabilize the protein leveled off above about 20% (w/w, dry basis). The FPF of the spray-dried formulations was stable during storage for rhuMAbE25 containing 10% and 20% mannitol, but the 30% formulation exhibited a dramatic decrease upon storage at both 5 degreesC and 30 degreesC, due to mannitol crystallization. We tested the addition of sodium phosphate to a 60:40 rhuMAbE25:mannitol (w:w) mixture, which otherwise crystallized upon spray drying and yielded a nonrespirable powder. The presence of sodium phosphate was successful in inhibiting mannitol crystallization upon spray drying and dramatically lowering the rate of solid-state aggregation. However, over long-term storage some crystallization was observed even for the phosphate-containing samples, concomitantly with increased particle size and decreased suitability for aerosol delivery. Therefore, the physical state of mannitol (i.e., amorphous or crystalline) plays a role both in maintaining protein stability and providing suitable aerosol performance when used as an excipient for spray-dried powders. Agents which retard mannitol crystallization, e.g., sodium phosphate, may be useful in extending the utility of mannitol as an excipient in spray-dried protein formulations.