Corticosteroids effect on caspase 3 expression in an in-vitro model of hypoxic brain cells.

OBJECTIVE: Effects of corticosteroids (CS) in the brain of growth-restricted fetus remain largely unstudied. We investigated if dexamethasone (DXM) exposure contributes to neuronal injury in an in-vitro model of neuronal cells under hypoxic conditions (surrogate for fetal growth restriction). STUDY DESIGN: U87 glioblastoma cells exposed to hypoxic or normoxic conditions for 10 h were incubated in the absence or presence of DXM for 48 h. Apoptosis as possible indicator of neurotoxicity was determined using a caspase-3-specific activity assay and western blotting. Caspase-3 was calculated as percentage of mean caspase-3 cleavage. Each experiment was performed in triplicate (n = 48). Caspase 3 activity in cell culture media was also measured by ELISA. RESULTS: Pro-caspase-3 (32 kDa) was expressed in culture, but activated 17 Kd caspase 3 was not expressed in cell lysate. There was no difference in ratio of caspase 3 activation when U87 cells were exposed to 10 v of hypoxia as compared to normoxia (0.46 ± 0.44 versus 0.37 ± 0.37). The pro-apoptotic effects of DXM were not increased by pre-exposure to hypoxia: (0.37 ± 0.37 versus 0.47 ± 0.40). CONCLUSION: The addition of DXM to hypoxic U87 cells had no additive or synergistic effects on the activation of caspase 3. Therefore, we speculate that the administration of CS in the setting of fetal growth restriction would not lead to increased apoptosis with potential neuronal injury.

Comparison of bactericidal properties of alcohol-based chlorhexidine versus povidone-iodine prior to amniocentesis.

OBJECTIVE: To compare the bactericidal properties of povidone-iodine versus alcohol-based chlorhexidine solution for cleansing the gravid abdomen prior to amniocentesis. METHODS: Fifty study participants were recruited from the University of Texas Women's Clinic in Houston, Texas. Two baseline swabs of the patients' abdomens were obtained to assess bacterial flora prior to treatment. A 10% povidone-iodine solution and 2% chlorhexidine gluconate with 70% isopropyl alcohol solution in a 3-mL prefilled applicator (Chloraprep, Cardinal Health, Inc., Leawood, KS) were then applied on different sides of the abdomen. After 30 seconds, cultures were obtained and plated on Trypticase Soy (PML Microbiologicals, Durham, NC) with sheep's blood agar for aerobic flora. Plates were incubated at 37°C for 48 hours for aerobic flora. Colony-forming units were counted and recorded. RESULTS: No statistically significant difference was detected between baseline colony counts between the left and right side of each patient's abdomen (p = 0.33) prior to cleansing. Postcleansing colony counts were evaluated, and a statistically significant difference was identified, favoring chlorhexidine as a more efficacious abdominal cleanser (p <0.001). CONCLUSION: We demonstrated that 2% chlorhexidine with 70% isopropyl alcohol had excellent bactericidal efficacy and was superior to povidone-iodine for cleansing the maternal abdomen.

Impact of progesterone on cytokine-stimulated nuclear factor-kappaB signaling in HeLa cells.

OBJECTIVE: A key event in the pathways leading to preterm labor may be the activation of nuclear factor-kappaB (NF-kappaB) in the fetal membranes and the cervix. Anti-inflammatory agents, such as the corticosteroids, inhibit the activation of NF-kappaB. We proposed to investigate the effects of progesterone pretreatment on cytokine-stimulated activation of NF-kappaB in HeLa cells, a human cervical epithelial cell line. METHODS: HeLa cells were pretreated with 10(-7) M progesterone for 24 hours and exposed to 1 ng/mL interleukin-1beta (IL-1beta) for 1 hour. Nuclear and cytosolic extracts were subjected to Western blot analysis using anti-p65 and anti-inhibitory protein-kappaBalpha (anti-IkappaBalpha) antibodies. Densitometric data (n=5) were compared using Kruskal-Wallis test. RESULTS: Pretreatment with progesterone interfered with IL-1beta-induced IkappaBalpha degradation. However, progesterone pretreatment resulted in a significant decrease in NF-kappaB protein subunit p65 in the cytoplasm. Pretreatment with progesterone did not reduce the amount of nuclear p65 and did not interfere with nuclear translocation of p65. CONCLUSION: Our observations suggest that any possible role played by progesterone in preterm labor prevention is not exerted through anti-inflammatory mechanisms of NF-kappaB down-regulation.

The effects of dexamethasone and betamethasone on surfactant protein-B messenger RNA expression in human type II pneumocytes and human lung adenocarcinoma cells.

OBJECTIVE: The purpose of this study was to compare the effect of a single 48-hour exposure to betamethasone or dexamethasone in the NCI-H441 cell line and in human type II pneumocytes. STUDY DESIGN: NCI-H441 cells were exposed 48 hours to varying concentrations of betamethasone or dexamethasone (10(-10) to 10(-7) mol/L) alone or in combination with 1 mmol/L dibutyryl cyclic adenosine monophosphate. Likewise, human type II pneumocytes were exposed 48 hours to varying concentrations of betamethasone or dexamethasone (10(-9) to 10(-7) mol/L) alone or in combination with 1 mmol/L dibutyryl cyclic adenosine monophosphate. The measured outcome was the stimulatory effect on surfactant protein B gene transcription as expressed by surfactant protein B messenger RNA accumulation. The experiment was conducted 5 times in NCI-H441 cells and 6 times in type II cells, in parallel with control. Surfactant protein B messenger RNA was determined at control level and 48 hours after exposure by quantitative reverse transcription-polymerase chain reaction. RESULTS: A similar dose-dependent response in surfactant protein B messenger RNA expression was seen with both betamethasone and dexamethasone. In human type II pneumocytes, the inductive profile of surfactant protein B messenger RNA after 48-hour exposure to betamethasone or dexamethasone was similar to that seen in the NCI-H441 cells. CONCLUSION: Dexamethasone and betamethasone achieved similar dose-response patterns of surfactant protein-B expression in vitro.

The effect of temperature on bactericidal properties of 10% povidone-iodine solution.

OBJECTIVE: Ten percent povidone-iodine (PVI) is commonly used as a bactericidal solution before amniocentesis is performed. Warming PVI may increase patient comfort; however, the effect of warming on its bactericidal properties is not known. The objective of this study was to determine the effect of warming 10% PVI on its bactericidal properties. STUDY DESIGN: Room temperature of PVI was 25 degrees C, and temperature in an ultrasonic gel warmer was 32 degrees C. In vitro experiments were conducted in 25 degrees C and 32 degrees C water baths. Nine milliliters of PVI at each temperature was added to 1 mL of bacteria (10(7) organisms/mL Staphylococcus aureus, Enterococcus species, Escherichia coli, group B Streptococcus). After 0.25, 0.5, 1, 2, 4, and 8 minutes, 1-mL samples were removed and added to 4 mL of 0.5% sodium thiosulfate (to neutralize the iodine and interrupt bactericidal action). The number of viable organisms was determined by plating 0.1-mL samples on trypticase soy agar plates. Plates were incubated at 37 degrees C for 24 hours and colony-forming units (CFUs) were counted. For in vivo experiments in 40 volunteers, a 9-cm2 area of the dorsum of each hand was wiped with a sterile cotton swab to obtain a sample of bacteria for culture. The area was then wiped for 15 seconds with PVI at 25 degrees C or 32 degrees C and recultured. The bactericidal properties of PVI at each temperature were compared. The Mann-Whitney U test was used as appropriate, and P <.05 was considered significant. RESULTS: In vitro, PVI was bactericidal against E coli, S aureus, Enterococcus species, and group B Streptococcus within 0.25 minutes at both 25 degrees C and 32 degrees C, with median bacterial growth of 0 CFU/plate for each species of bacteria studied (3 replicates at each temperature). Median bacterial growth from skin was 4 CFU/plate (range, 0 to >100). After hands were wiped with PVI at 25 degrees C and 32 degrees C, median bacterial growth was 0 CFU/plate (range, 0-8) and 0 CFU/plate (range, 0-15), respectively (not significant). CONCLUSION: PVI is as effective at 32 degrees C as it is at 25 degrees C. Use of PVI at 32 degrees C should be considered to increase patient comfort in procedures performed without the use of anesthetics.