Exopolysaccharides (EPSs) were isolated and purified from Lactobacillus rhamnosus strains RW-9595M, which has been shown to possess cytokine-stimulating activity, and R grown under various fermentation conditions (carbon source, incubation temperature and duration). Identical (1)H NMR spectra were obtained in all cases. Molecular masses were determined by gel permeation chromatography. The primary structure was elucidated using chemical and spectroscopic techniques. Organic acid, monosaccharide and absolute configuration analyses gave the following composition: pyruvate, 1; D-glucose, 2; D-galactose, 1; and l-rhamnose, 4. Methylation analysis indicated the presence of three residues of 3-linked rhamnose, and one residue each of 2,3-linked rhamnose, 2-linked glucose, 3-linked glucose and 4,6-linked galactose. The EPS was submitted to periodate oxidation followed by borohydride reduction. Monosaccharide analysis of the resulting polysaccharide gave the new composition: rhamnose, 4; and glucose, 1. Methylation analysis confirmed the loss of the 2-linked glucose and 4,6-linked galactose residues. On the basis of one- and two-dimensional (1)H and (13)C NMR data, the structure of the native EPS was consistent with the following heptasaccharide repeating unit: [3Rha alpha-3Glc beta-3[Gal4,6(R)Py alpha-2]Rha alpha-3Rha alpha-3Rha alpha-2Glc alpha-](n) where Rha corresponds to rhamnose (6-deoxymannose) and Py corresponds to pyruvate acetal. Complete (1)H and (13)C assignments are reported for the native and the corresponding pyruvate-hydrolysed polysaccharide. Electrospray MS and MS/MS data are given for the oligosaccharide produced by Smith degradation.

BACKGROUND

Premenstrual syndrome (PMS) is a cyclic mood disorder, widely believed, yet not conclusively shown, to be of endocrine etiology. This study examines basal levels of several hormones reported, albeit inconsistently, to differ in women with PMS compared with controls.

METHODS

Subjects (10 PMS patients and 10 controls) had their blood drawn for one full menstrual cycle. Subjects' mood and behavioral symptoms were assessed by daily self-ratings and objective ratings. Plasma was assayed for total and free testosterone (T), beta-endorphin (beta-EP), adrenocorticotropic hormone (ACTH), and cortisol.

RESULTS

No differences were observed between the PMS and control groups for beta-EP, ACTH, or cortisol. PMS subjects had significantly lower total and free T plasma levels with a blunting of the normal periovulatory peak, a finding that may be epiphenomenal to age.

CONCLUSIONS

This study does not confirm previous reports of abnormalities in plasma levels of either ACTH or beta-EP in women with PMS; it also fails to replicate a previous observation of high free T levels in women with PMS. These results are not supportive of a primary endocrine abnormality in PMS patients.

Ovarian steroids have previously been shown to regulate the hypothalamic content of beta-endorphin (beta EP) and its release into hypophyseal portal blood. Although the hypothalamic content of beta EP in cycling female rats was unchanged by ovariectomy, chronic treatment of ovariectomized rats with estradiol lowered hypothalamic beta EP levels. In this study, the hypothalamic content of beta EP was compared in male and cycling female rats, and the effects of orchiectomy and testosterone replacement on hypothalamic beta EP were examined. The beta EP content of the medial basal hypothalamus (MBH) was significantly higher in female rats compared to that in males of either the same weight (175-200 g) or the same age (65 days; P less than 0.025). When male rats were studied 4 weeks after castration, the beta-EP content of the MBH increased from a value of 2100 +/- 103 fmol in the controls to 2680 +/- 126 fmol (P less than 0.005). The hypothalamic beta EP content in the castrated males was similar to that in the intact females (2700 +/- 158 fmol). The increase in hypothalamic beta EP induced by castration was blocked by testosterone replacement. When orchiectomized animals were treated for 4 weeks with Silastic capsules filled with testosterone, there was a significant fall in the hypothalamic content of beta EP compared to that in the unreplaced animals. beta EP fell from 3180 +/- 115 to 2033 +/- 53 fmol in the MBH (P less than 0.001), from 1693 +/- 122 to 934 +/- 80 fmol in the anterior hypothalamus (P less than 0.001), and from 148 +/- 26 to 90.3 +/- 11 fmol in the median eminence (P less than 0.05). Testosterone replacement was also associated with a significant decline in the hypothalamic content of corticotropin-like intermediate lobe peptide and alpha MSH. Corticotropin-like intermediate lobe peptide fell from 2400 +/- 53 to 1560 +/- 84 fmol in the MBH (P less than 0.001) and from 1200 +/- 74 to 805 +/- 94 fmol in the anterior hypothalamus (P less than 0.01). alpha MSH fell from 1660 +/- 162 to 884 +/- 75 fmol in the MBH (P less than 0.001) and from 823 +/- 106 to 544 +/- 92 fmol in the anterior hypothalamus (P less than 0.05). Thus, testosterone, as well as estradiol, affects the hypothalamic content of several proopiomelanocortin-derived peptides. The effect on brain peptide content, however, depends on whether the steroids are secreted relatively constantly, as in the male, or fluctuate, as in the cycling female.

The purpose of this study was to evaluate the clinical performance of two "packable" posterior composites: Prodigy Condensable/Optibond Solo-Kerr (PC-OS) and Definite/Etch & Prime-Degussa (D-EP). Thirty-six patients participated in this study. A total of 78 restorations (40 with D-EP and 38 with PC-OS) were made. Each patient received at least two restorations (one of each studied material). The materials were handled according to the manufacturer's instructions. The occlusal adjustments were made at the placement visit. The restorations were finished and polished after 1 week. They were evaluated at baseline, and after 1 year and 2 years by two independent evaluators using the USPHS criteria. Colored slides were made of all the restorations. After 2 years, 34 patients and 74 restorations (38 with D-EP and 36 with PC-OS) were available for evaluation. A total of 50% of PC-OS restorations received A criterion and 50% received B criterion (2.8% color, 11.1% marginal staining, 27.8% superficial staining, 2.8% anatomic form and 5.6% marginal adaptation). For D-EP, 60.5% of restorations received A criterion and 39.5% received B criterion (2.6% color, 5.3% marginal staining, 10.5% superficial staining, 7.9% anatomic form and 13.2% marginal adaptation). The C criterion was observed only for marginal adaptation with D-EP (2 restorations-5.3%). The obtained data were tabulated and statistically analyzed using the Fisher, Chi-square and McNemar tests. After 2 years, PC-OS showed a significant increase in superficial and marginal staining. For D-EP the marginal adaptation and superficial staining became significantly worse than baseline.

Cells immunoreactive with anti-alpha-(17-39) ACTH, beta-(1-24) corticotropin, beta-LPH, alpha- and beta-EP were identified in the human fetal anterior pituitary at the ultrastructural level using the peroxidase-antiperoxidase complex method on ultrathin sections. Only one definite cell type was revealed by all these antisera. All granules of each individual immunostained cell reacted regardless of the antiserum used. The immunostained cells occurred in groups and were sometimes located in the wall of the follicle-like structures commonly observed in the fetal anterior pituitary. The cells revealed two main aspects: 1) The largest elements were rich in organelles, and their numerous secretory granules showed significantly variations in size (250-500 nm in diameter), electron density of their content and stain-deposit intensity. The ergastoplasm, consisting of irregular tubules, was poorly developed. In the vicinity of the conspicuous Golgi apparatus, organelles related to the GERL complex were commonly observed. Multivesicular bodies were frequent. Some of these cells showed bundles of microfilaments (60 nm in thickness). 2) The smaller cells had an electron-lucent hyaloplasm with sparse organelles; they contained fewer granules and never showed microfilaments. The immunocytological results are consistent with the synthesis of a molecule similar to pro-opiocortin by this type of endocrine cell in human fetuses. Morphological evidence for the maturation process of this precursor and for the secretory activity of these cells and its possible regulation is presented and discussed.

The N-type voltage-gated calcium channel (Ca(v)2.2) functions in neurons to regulate neurotransmitter release. It comprises a clinically relevant target for chronic pain. We have validated a calcium mobilization approach to assessing Ca(v)2.2 pharmacology in two stable Ca(v)2.2 cell lines: alpha1(B), alpha2delta, beta(3)-HEK-293 and alpha1(B), beta(3)-HEK-293. Ca(v)2.2 channels were opened by addition of KCl and Ca(2+) mobilization was measured by Fluo-4 fluorescence on a fluorescence imaging plate reader (FLIPR(96)). Ca(v)2.2 expression and biophysics were confirmed by patch-clamp electrophysiology (EP). Both cell lines responded to KCl with adequate signal-to-background. Signals from both cell lines were inhibited by omega-conotoxin (ctx)-MVIIa and omega-conotoxin (ctx)-GVIa with IC(50) values of 1.8 and 1nM, respectively, for the three-subunit stable, and 0.9 and 0.6nM, respectively, for the two-subunit stable. Other known Ca(v)2.2 blockers were characterized including cadmium, flunarizine, fluspirilene, and mibefradil. IC(50) values correlated with literature EP-derived values. Novel Ca(v)2.2 pharmacology was identified in classes of compounds with other primary pharmacological activities, including Na(+) channel inhibitors and antidepressants. Novel Na(+) channel compounds with high potency at Ca(v)2.2 were identified in the phenoxyphenyl pyridine, phenoxyphenyl pyrazole, and other classes. The highest potency at Ca(v)2.2 tricyclic antidepressant identified was desipramine.

The exopolysaccharide (EPS) structure from Lactobacillus johnsonii strain 151 isolated from the intestinal tract of mice was investigated. Sugar and methylation analyses together with (1)H and (13)C NMR spectroscopy, including two-dimensional (1)H,(1)H COSY, TOCSY, NOESY, and (1)H,(13)C HSQC, HMBC experiments, revealed that the repeating unit of the EPS is the linear pentasaccharide: →6)-α-d-Galp-(1→6)-α-d-Glcp-(1→3)-β-d-Galf-(1→3)-α-d-Glcp-(1→2)-β-d-Galf-(1→ The immunoreactivity of two structurally different exopolysaccharides isolated from L. johnsonii, 151 and 142 (Carbohydr. Res. 2010, 345, 108-114), was compared. Both EPSs differed in their reactivity with antisera. EPS from L. johnsonii 151 reacted with anti-Lactobacillus polyclonal sera against cells of five different strains, while EPS from L. johnsonii 142 was found to react only with its own antiserum. The broader specificity and higher reactivity of EPS from 151 strain than EPS from 142 strain were also observed with human sera. The physiological antibodies recognizing polysaccharide antigens were present in both adults and umbilical cord blood sera. A highly specific EPS 142 bearing strain was isolated from experimentally induced inflammatory bowel disease (IBD) mice, while a strain with EPS 151 isolated from the intestinal tract of healthy mice is characterized by a broad immune reactivity common structure.

BACKGROUND

Encapsulating peritoneal sclerosis (EPS) is a rare complication of peritoneal dialysis. The causes of EPS are not well defined and are likely multifactorial. A suitable animal model would facilitate research into the pathophysiology and treatment of EPS.

METHODS

We developed a helper-dependent adenovirus that expresses both green fluorescent protein (GFP) and active transforming growth factor-beta (TGF-beta1; HDAdTGF-beta1). Mice were administered HDAdTGF-beta1 via intraperitoneal injection and the response was compared with mice administered either first-generation adenovirus expressing TGF-beta1 (AdTGF-beta1) or control adenovirus (AdGFP).

RESULTS

HDAdTGF-beta1-treated mice continued to express the GFP reporter transgene to day 74, the end of the observation period. Transgene expression lasted less than 28 days in the animals treated with first-generation adenoviruses. Animals treated with first-generation AdTGF-beta1 demonstrated submesothelial thickening and angiogenesis at day 7, with almost complete resolution by day 28. The HDAdTGF-beta1-treated mice demonstrated progressive peritoneal fibrosis with adhesion formation and encapsulation of bowels. Weight gain was significantly reduced in animals treated with HDAdTGF-beta1 compared to both the control-treated animals and the AdTGF-beta1-treated animals. Inflammation was not a major component of the fibroproliferative response.

CONCLUSIONS

Peritoneal administration of a first-generation AdTGF-beta1 leads to transient gene expression, resulting in a resolving fibrotic response and histology similar to that seen in simple peritoneal sclerosis. Prolonged TGF-beta1 expression induced by the helper-dependent HDAdTGF-beta1 led to changes in peritoneal morphology resembling EPS. This suggests that TGF-beta1 may be a contributing factor in both simple peritoneal sclerosis and EPS. This model will be useful for elucidation of the mechanism of EPS and evaluation of potential treatment.

The current study deals with the evaluation of two coral-associated bacterial (CAB) extracts to inhibit the biofilm synthesis in vitro as well as the virulence production like hemolysin and exopolysaccharide (EPS), and also to assess their ability to modify the adhesion properties, that is cell surface hydrophobicity (CSH) of methicillin-resistant (MRSA) and -susceptible Staphylococcus aureus (MSSA). Out of nine CAB screened, the ethyl acetate extract of CAB-E2 (Bacillus firmus) and CAB-E4 (Vibrio parahemolyticus) have shown excellent antibiofilm activity against S. aureus. CAB-E2 reduced the production of EPS (57-79%) and hemolysin (43-70%), which ultimately resulted in the significant inhibition of biofilms (80-87%) formed by both MRSA and MSSA. Similarly, CAB-E4 was also found to decrease the production of EPS (43-57%), hemolysin (43-57%) and biofilms (80-85%) of test pathogens. CLSM analysis also proved the antibiofilm efficacy of CAB extracts. Furthermore, the CAB extracts strongly decreased the CSH of S. aureus. Additionally, FT-IR analysis of S. aureus treated with CAB extracts evidenced the reduction in cellular components compared to their respective controls. Thus, the present study reports for the first time, B. firmus-a coral-associated bacterium, as a promising source of antibiofilm agent against the recalcitrant biofilms formed by multidrug resistant S. aureus.

Bacillus cereus ATCC 14579 can respond to nutrient changes by adopting different forms of surface translocation. The B. cereus ATCC 14579 DeltaplcR mutant, but not the wild type, formed dendritic (branched) patterns on EPS [a low-nutrient medium that contains 7.0 g K(2)HPO(4), 3.0 g KH(2)PO(4), 0.1 g MgSO(4).7H(2)O, 0.1 g (NH(4))(2)SO(4), 0.01 g CaCl(2), 0.001 g FeSO(4), 0.1 g NaCl, 1.0 g glucose, and 125 mg yeast extract per liter] containing 0.7% agar. The dendritic patterns formed by sliding translocation of nonflagellated cells are enhanced under low-nutrient conditions and require sufficient production of a biosurfactant, which appears to be repressed by PlcR. The wild-type and complemented strains failed to slide on the surface of EPS agar because of the production of low levels of biosurfactant. Precoating EPS agar surfaces with surfactin (a biosurfactant produced by Bacillus subtilis) or biosurfactant purified from the DeltaplcR mutant rescued the ability of the wild-type and complemented strains to slide. When grown on a nutrient-rich medium like Luria-Bertani agar, both the wild-type and DeltaplcR mutant strains produced flagella. The wild type was hyperflagellated and elongated and exhibited swarming behavior, while the DeltaplcR mutant was multiflagellated and the cells often formed long chains but did not swarm. Thin-layer chromatography and mass spectrometry analyses suggested that the biosurfactant purified from the DeltaplcR mutant was a lipopeptide and had a mass of 1,278.1722 (m/z). This biosurfactant has hemolytic activity and inhibited the growth of several gram-positive bacteria.

The validation of a coupled 1D-0D model of the lower-limb arterial hemodynamics is presented. This study focuses on pathological subjects (6 patients, 72.7±11.1 years) suffering from atherosclerosis who underwent a femoro-popliteal bypass surgery. The 1D model comprises four vessels from the upper-leg, peripheral networks are modeled with three-element windkessels and in vivo velocity is prescribed at the inlet. The model is patient-specific: its parameters reflect the physiological condition of the subjects. In vivo data are acquired invasively during bypass surgery using B-mode ultrasonography and catheter. Simulations from the model compare well with measured velocity (u) and pressure (p) waveforms: average relative root-mean-square error between numerical and experimental waveforms are limited to εp=9.6%, εu=16.0%. The model is able to reproduce the intensity and shape of waveforms observed in different clinical cases. This work also details the introduction of blood leakages along the pathological arterial network, and the sensitivity of the model to its parameters. This study constitutes a first validation of a patient-specific numerical model of a pathological arterial network. It presents an efficient tool for engineers and clinicians to help them improve their understanding of the hemodynamics in diseased arteries.

BACKGROUND

Management of symptomatic atrial arrhythmia in pregnancy remains a challenge. In this case report, a pregnant woman with incessant tachycardia underwent successful left atrial ablation. The entire procedure was performed without fluoroscopy.

RESULTS

A 20-year-old woman, 27 weeks pregnant, was admitted with congestive cardiac failure and incessant atrial tachycardia. She had an elevated brain natriuretic peptide (BNP) and chest X-ray demonstrating heart failure. The 12-lead electrocardiogram (ECG) showed atrial tachycardia with a cycle length of 310 ms, inverted P waves in lead I and the inferior leads, and a ventricular rate of 84 bpm during 2:1 block. Echocardiogram showed a global reduction in left ventricular function with a left ventricular ejection fraction (LVEF) of 0.40. Electrical cardioversion failed. Rate control could not be achieved with beta-blockers and calcium antagonists. Amiodarone with repeat cardioversion was also unsuccessful. The patient then underwent catheter ablation. The entire procedure was performed using intracardiac echocardiography (ICE) and electroanatomical mapping with no fluoroscopy. Electrophysiology (EP) study and an activation map of the left atrium confirmed a focal left atrial tachycardia which was successfully ablated. Six weeks postablation, the left ventricular function had normalized and the patient delivered a healthy child at term, without complication.

CONCLUSIONS

Ablation of left atrial tachycardia using ICE and electroanatomical guidance is feasible in pregnant women.

BACKGROUND

This phase Ib study aimed to establish the feasible everolimus dose given with standard-dose etoposide plus cisplatin (EP) for extensive-stage small-cell lung cancer (SCLC).

METHODS

An adaptive Bayesian dose-escalation model and investigator opinion were used to identify feasible daily or weekly everolimus doses given with EP in adults with treatment-naive extensive-stage SCLC. A protocol amendment mandated prophylactic granulocyte colony-stimulating factor (G-CSF). Primary end point was cycle 1 dose-limiting toxicity (DLT) rate. Secondary end points included safety, relative EP dose intensity, pharmacokinetics, and tumor response.

RESULTS

Patients received everolimus 2.5 or 5 mg/day without G-CSF (n=10; cohort A), 20 or 30 mg/week without G-CSF (n=18; cohort B), or 2.5 or 5 mg/day with G-CSF (n=12; cohort C); all received EP. Cycle 1 DLT rates were 50.0%, 22.2%, and 16.7% in cohorts A, B, and C, respectively. Cycle 1 DLTs were neutropenia (cohorts A and B), febrile neutropenia (all cohorts), and thrombocytopenia (cohorts A and C). The most common grade 3/4 adverse events were hematologic. Best overall response was partial response (40.0%, 61.1%, and 58.3% in cohorts A, B, and C, respectively).

CONCLUSIONS

Everolimus 2.5 mg/day plus G-CSF was the only feasible dose given with standard-dose EP in untreated extensive-stage SCLC.

Histamine, prostaglandin E(2), and catecholamines have been demonstrated to regulate the innate and acquired immune responses. In this review, we describe one of the mechanisms common to the action of these agonists; the regulation of the expression of costimulatory adhesion molecules such as ICAM-1 and B7 antigens on monocytes/macrophages. The specific receptor subtypes involved in the action of each agonist were H(2) for histamine, EP(2)/EP(4) for prostaglandin E(2), and beta(2) for catecholamines, all of which are coupled with adenylate cyclase via Gs protein. The regulation of the expression of adhesion molecules by these agonists in turn leads to the modulation of subsequent cytokine production mediated by cell-cell interaction under different stimuli. Histamine is synthesized in monocytes and T cells by the induction of histidine decarboxylase. The inducible histamine has different dynamics from that in storage granules of mast cells and basophils. Also, noradrenaline appears to be synthesized in lymphocytes. Thus, immune cells can produce histamine, prostaglandins, and noradrenaline by themselves and modulate the cell-cell interaction between monocytes and other cells. Some of the inhibitors of HMG-CoA reductase were shown to bind to the ICAM-1-binding domain of LFA-1, reducing the interaction mediated by ICAM-1/LFA-1. The regulation of interaction mediated by adhesion molecules may provide a new target for controlling inflammatory and immune responses.

BACKGROUND

Though moxibustion is frequently used to treat primary dysmenorrhea in China, relevant evidence supporting its effectiveness is still scanty.

METHODS

This study was a pragmatic randomized, conventional drug controlled, open-labeled clinical trial. After initial screen, 152 eligible participants were averagely randomized to receive two different treatment strategies: Moxibustion and conventional drugs. Participants and practitioners were not blinded in this study. The duration of each treatment was 3 months. The primary outcome was pain relief measured by the Visual Analogue Scale. The menstrual pain severity was recorded in a menstrual pain diary.

RESULTS

152 eligible patients were included but only 133 of them eventually completed the whole treatment course. The results showed that the menstrual pain intensity in experimental group and control group was reduced from 6.38±1.28 and 6.41±1.29, respectively, at baseline, to 2.54±1.41 and 2.47±1.29 after treatment. The pain reduction was not significantly different between these two groups (P = 0.76), however; the pain intensity was significantly reduced relative to baseline for each group (P<0.01). Three months after treatment, the effectiveness of moxibustion sustained and started to be superior to the drug's effect (-0.87, 95%CI -1.32 to -0.42, P<0.01). Secondary outcome analyses showed that moxibustion was as effective as drugs in alleviating menstrual pain-related symptoms. The serum levels of pain mediators, such as PGF2α, OT, vWF, β-EP, PGE2, were significantly improved after treatment in both groups (P<0.05). No adverse events were reported in this trial.

CONCLUSIONS

Both moxibustion and conventional drug showed desirable merits in managing menstrual pain, given their treatment effects and economic costs. This study as a pragmatic trial only demonstrates the effectiveness, not the efficacy, of moxibustion for menstrual pain. It can't rule out the effect of psychological factors during treatment process, because no blind procedure or sham control was used due to availability. In clinical practice, moxibustion should be used at the discretion of patients and their physicians.

BACKGROUND

ClinialTrials.gov NCT01972906.

BACKGROUND

The clinical and neurophysiological effects of neurofeedback (NF) as treatment for children with ADHD are still unclear. This randomized controlled trial (RCT) examined electroencephalogram (EEG) power spectra before and after NF compared to methylphenidate (MPH) treatment and physical activity (PA) - as semi-active control group - during resting and active (effortful) task conditions to determine whether NF can induce sustained alterations in brain function.

METHODS

Using a multicentre three-way parallel group RCT design, 112 children with a DSM-IV diagnosis of ADHD, aged between 7 and 13 years, were initially included. NF training consisted of 30 sessions of theta/beta training at Cz over a 10-week period. PA training was a semi-active control group, matched in frequency and duration. Methylphenidate was titrated using a double-blind placebo controlled procedure in 6 weeks, followed by a stable dose for 4 weeks. EEG power spectra measures during eyes open (EO), eyes closed (EC) and task (effortful) conditions were available for 81 children at pre- and postintervention (n = 29 NF, n = 25 MPH, n = 27 PA).

BACKGROUND

Train Your Brain? Exercise and Neurofeedback Intervention for ADHD, https://clinicaltrials.gov/show/;NCT01363544, Ref. No. NCT01363544.

RESULTS

Both NF and MPH resulted in comparable reductions in theta power from pre- to postintervention during the EO condition compared to PA (ηp (2) = .08 and .12). For NF, greater reductions in theta were related to greater reductions in ADHD symptoms. During the task condition, only MPH showed reductions in theta and alpha power compared to PA (ηp (2) = .10 and .12).

CONCLUSIONS

This study provides evidence for specific neurophysiological effects after theta/beta NF and MPH treatment in children with ADHD. However, for NF these effects did not generalize to an active task condition, potentially explaining reduced behavioural effects of NF in the classroom.

Many conventional chemotherapeutic drugs, as well as DNA for cancer gene therapy, require efficient access to the cell interior to be effective. The cell membrane is a formidable barrier to many of these drugs, including therapeutic DNA constructs. Electropermeabilization (EP, often used synonymously with "electroporation") has become a useful method to temporarily increase the permeability of the cell membrane, allowing a broad variety of molecules efficient access to the cell interior. EP is achieved by the application of short electrical pulses of relatively high local field strength to the target tissue of choice. In cancer therapy, EP can be applied in vivo directly to the tumor to be treated, in order to enhance intracellular uptake of drugs or DNA. Alternatively, EP can be used to deliver DNA into cells of healthy tissue to achieve longer-lasting expression of cancer-suppressing genes. In addition, EP has been used in ex vivo therapeutic approaches for the transfection of a variety of cells in suspension. In this paper, we communicate results related to the development of a treatment for squamous cell carcinomas of the head and neck, using electropermeabilization to deliver the drug bleomycin in vivo directly into the tumor cells. This drug, which is not particularly effective as a conventional therapeutic, becomes highly potent when the intracellular concentration is enhanced by EP treatment. In animal model experiments we found a drug dose of 1 U/cm(3) tumor tissue (delivered in 0.25 mL of an aqueous solution/cm3 tumor tissue) and an electrical field strength of 750 V/cm or higher to be optimal for the treatment of human squamous cell tumors grown subcutaneously in mice. Within 24-48 hours, the majority of tumor cells are rapidly destroyed by this bleomycin-electroporation therapy (B-EPT). This raises the concern that healthy tissue may be similarly affected. In studies with large animals we showed that normal muscle and skin tissue, normal tissue surrounding major blood vessels and nerves, as well as healthy blood vessels and nerves themselves, are much less affected than tumor tissue. Normal tissues did show acute, focal, and transitory effects after treatment, but these effects are relatively minor under standard treatment conditions. The severity of these effects increases with the number of electric pulse cycles and applied voltage. The observed histological changes resolved 20 to 40 days after treatment or sooner, even after excessive EP treatment. Thus, B-EPT is distinct from other ablative therapies, such as thermal, cryo, or photodynamic ablation, which equally affect healthy and tumor tissue. In comparison to surgical or radiation therapy, B-EPT also has potential as a tissue-sparing and function-preserving therapy. In clinical studies with over 50 late stage head and neck cancer patients, objective tumor response rates of 55-58%, and complete tumor response rates of 19-30% have been achieved.

Emerging evidence indicates the pronounced role of inflammasome activation linked to reactive oxygen species (ROS) in the sterile inflammatory response triggered by ischemia/reperfusion (I/R) injury. Ethyl pyruvate (EP) is an antioxidant and conveys myocardial protection against I/R injury, while the exact mechanisms remain elusive. We aimed to investigate the effect of EP on myocardial I/R injury through mechanisms related to ROS and inflammasome regulation. The rats were randomly assigned to four groups: (1) sham, (2) I/R-control (IRC), (3) EP-pretreatment + I/R, and (4) I/R + EP-posttreatment. I/R was induced by a 30 min ligation of the left anterior descending artery followed by 4 h of reperfusion. EP (50 mg/kg) was administered intraperitoneally at 1 h before ischemia (pretreatment) or upon reperfusion (posttreatment). Both pre- and post-EP treatment resulted in significant reductions in myocardial infarct size (by 34% and 31%, respectively) and neutrophil infiltration. I/R-induced myocardial expressions of NADPH oxidase-4, carnitine palmitoyltransferase 1A, and thioredoxin-interacting protein (TXNIP) were mitigated by EP. EP treatment was associated with diminished inflammasome activation (NOD-like receptor 3 (NLRP3), apoptosis-associated speck-like protein, and caspase-1) and interleukin-1β induced by I/R. I/R-induced phosphorylation of ERK and p38 were also mitigated with EP treatments. In H9c2 cells, hypoxia-induced TXNIP and NLRP3 expressions were inhibited by EP and to a lesser degree by U0126 (MEK inhibitor) and SB203580 (p38 inhibitor) as well. EP's downstream protective mechanisms in myocardial I/R injury would include mitigation of ROS-mediated NLRP3 inflammasome upregulation and its associated pathways, partly via inhibition of hypoxia-induced phosphorylation of ERK and p38.

Nanoparticle-based systems for concurrent delivery of multiple drugs can improve outcomes of cancer treatments, but face challenges because of differential solubility and fairly low threshold for incorporation of many drugs. Here we demonstrate that this approach can be used to greatly improve the treatment outcomes of etoposide (ETO) and platinum drug combination ("EP/PE") therapy that is the backbone for treatment of prevalent and deadly small cell lung cancer (SCLC). A polymeric micelle system based on amphiphilic block copolymer poly(2-oxazoline)s (POx) poly(2-methyl-2-oxazoline- block-2-butyl-2-oxazoline- block-2-methyl-2-oxazoline) (P(MeOx- b-BuOx- b-MeOx) is used along with an alkylated cisplatin prodrug to enable co-formulation of EP/PE in a single high-capacity vehicle. A broad range of drug mixing ratios and exceptionally high two-drug loading of over 50% wt. drug in dispersed phase is demonstrated. The highly loaded POx micelles have worm-like morphology, unprecedented for drug loaded polymeric micelles reported so far, which usually form spheres upon drug loading. The drugs co-loading in the micelles result in a slowed-down release, improved pharmacokinetics, and increased tumor distribution of both drugs. A superior antitumor activity of co-loaded EP/PE drug micelles compared to single drug micelles or their combination as well as free drug combination was demonstrated using several animal models of SCLC and non-small cell lung cancer.

A screening method based on differential staining of the wild type and exopolysaccharide-deficient mutants of Rhizobium (Sinorhizobium) meliloti by the lipophilic dye Sudan Black B is described. Mutants defective in the production of either succinoglycan or EPS II (galactoglucan) were isolated by using this method, which might also prove useful for isolating exopolysaccharide-defective derivatives of other bacteria.

Although glucocorticoids clearly inhibit proopiomelanocortin (POMC) gene transcription and peptide synthesis in the anterior pituitary, the effects of glucocorticoids on POMC in the hypothalamus are still unclear, even though most POMC neurons in the arcuate nucleus are known to have glucocorticoid receptors. In this study, we have therefore examined the effect of adrenalectomy (ADX) and glucocorticoid replacement on POMC mRNA and peptide (beta-EP and alpha-MSH) levels in the medial basal hypothalamus (MBH) of the rat. POMC mRNA was measured by a sensitive solution hybridization S1 nuclease protection assay, and beta-EP and alpha-MSH were measured by radioimmunoassay. In a first experiment, animals were studied 7 days after ADX or sham surgery. The mean POMC mRNA concentration was 1.01+/-0.14 pg/microg RNA (means+/-SE) in the intact animals and decreased to 0.55+/-0.07 pg/microg RNA in the MBH of the ADX animals (p < 0.005). Beta-EP levels decreased in parallel from 4.30+/-0.18 to 3.36+/-0.11 ng/mg protein (p < 0.001); alpha-MSH levels decreased from 3.25+/-0.21 to 2.41+/-0.16 ng/mg protein (p < 0.005). In a second experiment, animals were studied 2 weeks after ADX. POMC mRNA levels again fell significantly from 1.15+/-0.19 pg/microg RNA in the intact animals to 0.51+/-0.06 pg/microg in the ADX animals (p < 0.01). Beta-EP levels fell also, but this was not significant. In a third experiment, all animals underwent ADX, and half of them received daily subcutaneous injections of dexamethasone (20 microg). Nine days after ADX, the mean POMC mRNA level was 0.66+/-0.04 pg/microg RNA in the saline-treated animals and increased to 0.98+/-0.08 pg/microg RNA in the dexamethasone-treated animals (p < 0.005). A parallel increase in beta-EP levels from 5.03+/-0.41 to 6.01+/-0.53 ng/mg protein was also noted, but this was not statistically significant. We conclude that POMC gene expression is significantly inhibited in the MBH at 1 and 2 weeks after ADX. This effect was reversed by glucocorticoid replacement with doses close to the physiological range. The parallel changes in POMC mRNA and peptide levels strongly suggest that, in contrast to the anterior pituitary, low doses of glucocorticoids stimulate the biosynthesis of POMC in the MBH of ADX rats.

In areas of southern Africa, aqueous extracts from the roots of Pelargonium sidoides are employed to cure various disorders. Nowadays, a modern formulation (EPs 7630), elaborated from the traditional herbal medicine, is successfully used for the treatment of respiratory tract diseases. We previously observed that root extracts of P. sidoides and their representative constituents exhibit moderate antibacterial and significant immunomodulatory capabilities. The present study was performed to further assess the efficacy and mode of action for these pharmacological activities. The results indicate that P. sidoides extracts may well possess antimycobacterial activity as claimed in traditional uses. Furthermore, significant antibacterial properties against multi-resistant Staphylococcus aureus strains as well as TNF-inducing potencies and prominent interferon-like activities in supernatants of sample-activated bone marrow-derived macrophages were observed using several functional assays. In addition, EPs 7630 stimulated the synthesis of IFN-beta in MG 63 cells as demonstrated by a specific enzyme immunoassay. For gallic acid, a characteristic constituent, evidence for the expression of iNOS and TNF-alpha transcripts in stimulated RAW 264.7 cells and, hence, activation at the transcriptional level was revealed by RT-PCR. The present results, when taken together with the recently reported pharmacological activities, provide for a rationale basis of the utilization of EPs 7630 in the treatment of respiratory tract infections.

1. To examine individual hormonal responses to extreme physical stress, blood samples were taken from eight highly trained athletes 1 day before and within 15 min of finishing the 1986 1000 km Sydney to Melbourne Ultramarathon foot race. 2. The baseline hormonal state of these highly trained athletes was quite different from normal. Resting serum conjugated catecholamines--epinephrine (E), norepinephrine (NE), dopamine (D), free E and free D--were significantly elevated above the normal mean (P less than 0.01). ACTH levels were significantly elevated above the normal range. Immunoreactive beta-endorphin (IR-beta EP), growth hormone (GH), prolactin (PRL), testosterone, cortisol and cortisol-binding globulin (CBG) were within the normal range. 3. The effect of the race on serum catecholamine levels was to elevate further free and conjugated NE (P less than 0.01). Other catecholamines, free and conjugated, remained significantly elevated above the normal mean (P less than 0.01). Adrenocorticotrophic hormone (ACTH) remained elevated, and IR-beta EP within the normal range, without significant change. A significant increase in GH (P less than 0.05), PRL (P less than 0.01), and cortisol (P less than 0.01) was seen, with no change in CBG. 4. As a model of chronic physical stress, the ultramarathon runner demonstrates a significantly altered baseline hormonal state as reflected in the primary mediators of the stress response, the catecholamines and the hypothalamic-pituitary-adrenal axis. Their response to severe exercise is distinct from that of untrained individuals in whom conjugated catecholamines decrease and ACTH increase. This may represent hormonal adaptation to prolonged stress.

The objective of this study was to investigate the effect of Mg(2+) ions on biofilm formation by Bacillus species, which are considered as problematic microorganisms in the food industry. We found that magnesium ions are capable to inhibit significantly biofilm formation of Bacillus species at 50 mM concentration and higher. We further report that Mg(2+) ions don't inhibit bacterial growth at elevated concentrations; hence, the mode of action of Mg(2+) ions is apparently specific to inhibition of biofilm formation. Biofilm formation depends on the synthesis of extracellular matrix, whose production in Bacillus subtilis is specified by two major operons: the epsA-O and tapA operons. We analyzed the effect of Mg(2+) ions on matrix gene expression using transcriptional fusions of the promoters for eps and tapA to the gene encoding β galactosidase. The expression of the two matrix operons was reduced drastically in response to Mg(2+) ions suggesting about their inhibitory effect on expression of the matrix genes in B. subtilis. Since the matrix gene expression is tightly controlled by Spo0A dependent pathway, we conclude that Mg(2+) ions could affect the signal transduction for biofilm formation through this pathway.