Significant Increase In Release Research Articles

In Vitro Evaluation of Mercury Release from Dental Amalgam After Magnetic Resonance Imaging

Background Dental amalgam is a widely used restorative material that is prepared by triturating mercury with a metal alloy. Mercury vapor is released during placement and removal of the restoration. It is also released during chewing and brushing. A few studies have reported that mercury is also released on exposure to MRI.Objectives This was an in vitro cross-sectional study. In this study we aimed to evaluate mercury release from dental amalgam after exposure to magnetic resonance imaging MRI.Methods Seven blocks of amalgam measuring about 5times5times5 mm were prepared. Each block was placed in 20 mL of artificial saliva for 48 hours and mercury analysis was performed using Inductive coupled plasma mass spectrometry before and after exposure to MRI. Students lsquotrsquo test was used for statistical analysis.Results Mean mercury levels before and after exposure to MRI were 4.74 ppb and 8.68 ppb respectively. Thus there was a significant increase in release of mercury after MRI. Conclusion It can be concluded that it is advisable to use non-mercury containing restorative materials in dentistry.

Protease-Responsive Hydrogel Microparticles for Intradermal Drug Delivery.

Protease-responsive multi-arm polyethylene glycol-based microparticles with biscysteine peptide crosslinkers (CGPGG↓LAGGC) were obtained for intradermal drug delivery through inverse suspension photopolymerization. The average size of the spherical hydrated microparticles was ∼40 μm after crosslinking, making them attractive as a skin depot and suitable for intradermal injections, as they are readily dispensable through 27G needles. The effects of exposure to matrix metalloproteinase 9 (MMP-9) on the microparticles were evaluated by scanning electron microscopy and atomic force microscopy, demonstrating partial network destruction and decrease in elastic moduli. Given the recurring course of many skin diseases, the microparticles were exposed to MMP-9 in a flare-up mimicking fashion (multiple-time exposure), showing a significant increase in release of tofacitinib citrate (TC) from the MMP-responsive microparticles, which was not seen for the non-responsive microparticles (polyethylene glycol dithiol crosslinker). It was found that the degree of multi-arm complexity of the polyethylene glycol building blocks can be utilized to tune not only the release profile of TC but also the elastic moduli of the hydrogel microparticles, with Young's moduli ranging from 14 to 140 kPa going from 4-arm to 8-arm MMP-responsive microparticles. Finally, cytotoxicity studies conducted with skin fibroblasts showed no reduction in metabolic activity after 24 h exposure to the microparticles. Overall, these findings demonstrate that protease-responsive microparticles exhibit the properties of interest for intradermal drug delivery.

Formulation and Evaluation of Microspheres as an effective Colon Targeting Drug Delivery System

The present work comprised of the preparation of coated HPMC capsules of meloxicam microspheres in order to target the drug release in colon resulting in increased absorption and subsequent bioavailability. The meloxicam microspheres were developed in nineteen different batches using HPMC, combination of EC with HPMC, Eudragits (S100, RS100 and E100) in different drug and polymer ratios by solvent evaporation method. The formulation of different batches were examined by various studies i.e. percentage yield, surface morphology (SEM), particle size analysis, entrapment efficiency, drug compatibility with polymers using FTIR and in-vitro drug release determinations. The complex of meloxicam with β-cyclodextrin increased the solubility of the drug accompanying its in-vitro release. The microspheres filled HPMC capsules were coated with eudragit S100 which proved to be an effective method for drug targeting to the colon. A 22 factorial design showed that a very significant increase in release of the drug using polymers i.e. HPMC, E.C with HPMC, Eudragit S100, Eudragit RS100, Eudragit E100 could be obtained by the exclusive manipulation of two variables i.e. surfactant and polymer concentrations. The drug loaded microspheres showed percentage drug entrapment as 35.09-62.50% in A1-A4, 76.29-87.78% in B1-B4, 81.36-92.70% in S1-S4, 65.47-69.4% in RS1-RS4 and 68.91-78.08% in E1-E3. The pH 7.4 phosphate buffer and simulated colonic fluid were used for the in-vitro release tests. The best drug release profiles were seen with formulations A3, B2, RS1, S1 and E2 (containing different ratios of drug: polymer) coated with 15% (w/v) Eudragit S-100 solution.

Effects of cyclic compression on intervertebral disc metabolism using a whole-organ rat tail model.

Many factors contribute to the development and progression of intervertebral disc (IVD) degeneration. This study was designed to assess the effects of compressive load magnitude on IVD metabolism. It was hypothesized that as load magnitude increased, there would be a significant increase in release of proinflammatory and degradative biomarkers, and a significant decrease in tissue proteoglycan (GAG) and collagen contents compared with unloaded controls. IVD whole organ functional spinal units (FSU) consisting of cranial and caudal body halves, cartilage endplates, and IVD (n = 36) were harvested from the tails of six Sprague Dawley rats, and FSUs were cultured at 0.0 MPa, 0.5 MPa, or 1.0 MPa at 0.5 Hz for 3 days. After culture, media were collected for biomarker analysis and FSUs were analyzed for extracellular matrix composition. Significant differences were determined using a one-way analysis of variance or Kruskal-Wallis test and post hoc analyses. Media concentrations of IFN-γ, IL-6, IL-1β, and MMP-8 were significantly higher in the 0.5 MPa compared with the 0.0 MPa group. Media concentrations of PGE2 and TIMP-1 were significantly higher in the 1.0 MPa group compared with the 0.0 MPa group, and media PGE2 was significantly higher in the 1.0 MPa group compared with the 0.5 MPa group. Media GAG content was significantly higher in the 1.0 MPa group compared with the 0.0 MPa group, and percent GAG in the tissue was significantly lower in 0.5 MPa and 1.0 MPa groups compared with the 0.0 MPa group. Clinical Significance: These data suggest that there are magnitude-dependent inflammatory and degradative IVD responses to cyclic loading, which may contribute to IVD degeneration.

Exposure to particle debris generated from passenger and truck tires induces different genotoxicity and inflammatory responses in the RAW 264.7 cell line.

A number of studies have shown variable grades of cytotoxicity and genotoxicity in in vitro cell cultures, laboratory animals and humans when directly exposed to particle debris generated from tires. However, no study has compared the effects of particles generated from passenger tires with the effects of particles from truck tires. The aim of this study was to investigate and relate the cyto- and genotoxic effects of different types of particles (PP, passenger tire particles vs. TP, truck tire particles) in vitro using the phagocytic cell line RAW 264.7 (mouse leukaemic monocyte macrophage cell line). The viability of RAW 264.7 cells was determined by the 3- (4,5-dimethylthiazol-2-yl) -5- (3-carboxymethoxyphenyl) -2- (4-sulfophenyl) -2H-tetrazolium (MTS) assay following exposure for 4, 24 and 48 hours to different particle concentrations (10 μg / ml, 25 μg / ml, 50 μg / ml, 100 μg / ml). The effects of particles of passenger and truck tires on cell proliferation and genotoxicity were evaluated by means of the cytokinesis-block micronucleus (CBMN) assay following exposure for 24 hours to different particle concentrations (10 μg / ml, 25 μg / ml, 50 μg / ml, 100 μg / ml). In MTS assay, after 24 hours, it was found that PP induced a 30% decrease in metabolic activity at a concentration of 10 μg/ml, while TP caused reductions of 20% and 10% at concentrations of 10 μg/ml and 50 μg/ml, respectively. At 48 hours after the treatments, we observed increased metabolic activity at 50 μg/ml and 100 μg/ml for the PP while only at 50 μg/ml for the TP. The CBMN assay showed a significant increase in the number of micronuclei in the cells incubated with PP in all experimental conditions, while the cells treated with TP showed a meaningful increase only at 10 μg /ml. We utilized the TNF-α ELISA mouse test to detect the production of tumour necrosis factor-alpha (TNF-α) in RAW 264.7 cells. The effect of passenger and truck particles on TNF-α release was evaluated following exposure for 4 and 24 hours.After 4 hours of incubation, the cells treated with PP and TP at 100 μg / ml showed a slight but significant increase in TNF-α release, while there was a significant increase in the release of TNF-α after 24 hours of incubation with both tire samples in the cells treated with 50 and 100 μg / ml PP. The data obtained show a higher cytotoxic, clastogenic/genotoxic and inflammatory effects of passenger compared to the truck tire particles.

IDO, COX and iNOS have an important role in the proliferation of Neospora caninum in neuron/glia co-cultures

Central nervous system (CNS) is the main site for encystment of Neospora caninum in different animal species. In this tissue, glial cells (astrocytes and microglia) modulate responses to aggression in order to preserve homeostasis and neuronal function. Previous data showed that when primary cultures of glial cells are infected with N. caninum, they develop gliosis and the immune response is characterized by the release of TNF and IL-10, followed by the control of parasite proliferation. In order to elucidate this control, three enzymatic systems involved in parasite-versus-host interactions were observed on a model of neuron/glia co/cultures obtained from rat brains. Indoleamine 2,3-dioxygenase (IDO), induced nitric oxide synthase (iNOS) responsible for the catabolism of tryptophan and arginine, respectively, and cycloxigenase (COX) were studied comparing their modulation by respective inhibitors with the number of tachyzoites or the immune response measured by the release of IL-10 and TNF. Cells were treated with the inhibitors of iNOS (1.5 mM L-NAME), IDO (1 mM 1-methyl tryptophan), COX-1 (1 μM indomethacin) and COX-2 (1 μM nimesulide) before infection with tachyzoites of N. caninum (1:1 cell: parasite). After 72 h of infection, immunocytochemistry showed astrogliosis and a significant increase in the number and length of neurites, compared with uninfected co-cultures, while an increase of IL-10 and TNF was verified. N. caninum did not change iNOS activity, but the inhibition of the basal levels of this enzyme stimulated parasite proliferation. Additionally, a significant increase of about 40% was verified in the IDO activity, whose inhibition caused 1.2-fold increase in parasitic growth. For COX-2 activity, infection of cultures stimulated a significant increase in release of PGE2 and its inhibition by nimesulide allowed the parasitic growth. These data indicate that iNOS, IDO and COX-2 control the proliferation of N. caninum in this in vitro model. On the other hand, the release of IL-10 by glia besides modulating the inflammation also allow the continuity of parasitism.

Gemcitabine: Selective cytotoxicity, induction of inflammation and effects on urothelial function

Intravesical gemcitabine has recently been introduced for the treatment of superficial bladder cancer and has a favourable efficacy and toxicity profile in comparison to mitomycin c (MMC), the most commonly used chemotherapeutic agent. The aim of this study was to assess the cytotoxic potency of gemcitabine in comparison to MMC in urothelial cell lines derived from non-malignant (UROtsa) and malignant (RT4 and T24) tissues to assess selectivity. Cells were treated with gemcitabine or mitomycin C at concentrations up to the clinical doses for 1 or 2h respectively (clinical duration). Treatment combined with hyperthermia was also examined. Cell viability, ROS formation, urothelial function (ATP, acetylcholine and PGE2 release) and secretion of inflammatory cytokines were assessed. Gemcitabine displayed a high cytotoxic selectivity for the two malignant cell lines (RT4, T24) compared to the non-malignant urothelial cells (UROtsa, proliferative and non-proliferative). In contrast, the cytotoxic effects of MMC were non-selective with equivalent potency in each of the cell lines. The cytotoxic effect of gemcitabine in the malignant cell lines was associated with an elevation in free radical formation and was significantly decreased in the presence of an equilibrative nucleoside transporter inhibitor. Transient changes in urothelial ATP and PGE2 release were observed, with significant increase in release of interleukin-6, interleukin-8 and interleukin-1β from urothelial cells treated with gemcitabine. The selectivity of gemcitabine for malignant urothelial cells may account for the less frequent adverse urological effects with comparison to other commonly used chemotherapeutic agents.

The Release of Elements from the Base Metal Alloys in a Protein Containing Biologic Environments and Artificial Saliva - An Invitro Study.

It has been reported that protein containing solutions can accelerate the release of elements from the base metal alloys. This study aims to determine whether the solution in which an alloy is submerged and the exposure time have any effect on the amount of release of elements from the Ni-Cr and Co-Cr alloys. A total of 126 specimens were made from the Ni-Cr alloy and 42 specimens were made from Co-Cr alloy in the form of 5mm diameter discs, 2mm in thickness. Dissolution experiments were carried out in Bovine Serum Albumin (BSA) and artificial saliva for a period of seven weeks and atomic absorption spectrophotometer was used for elemental analysis. T-test was done to correlate the difference of elemental release from both BSA and artificial saliva. ANOVA test was done to compare the release at different time intervals and to compare the release of elements at different time intervals within a particular solution. TUKEY HSD test was done for comparison between the elements in a particular solution. The results showed that the elemental release was seen in both the solutions with a significant increase of release in BSA. The release of elements from the Ni-Cr alloy showed the predominant release of Cr. The protein containing solution showed maximum release of elements from Ni-Cr and Co-Cr alloys. The elements that released from the alloys never reached their threshold for toxic effects. Hence these alloys can be safely used in fabrication of metal restorations without any ill effects.

Mimicking exposures to acute and lifetime concentrations of inhaled silver nanoparticles by two different in vitro approaches.

In the emerging market of nano-sized products, silver nanoparticles (Ag NPs) are widely used due to their antimicrobial properties. Human interaction with Ag NPs can occur through the lung, skin, gastrointestinal tract, and bloodstream. However, the inhalation of Ag NP aerosols is a primary concern. To study the possible effects of inhaled Ag NPs, an in vitro triple cell co-culture model of the human alveolar/airway barrier (A549 epithelial cells, human peripheral blood monocyte derived dendritic and macrophage cells) together with an air–liquid interface cell exposure (ALICE) system was used in order to reflect a real-life exposure scenario. Cells were exposed at the air–liquid interface (ALI) to 0.03, 0.3, and 3 µg Ag/cm2 of Ag NPs (diameter 100 nm; coated with polyvinylpyrrolidone: PVP). Ag NPs were found to be highly aggregated within ALI exposed cells with no impairment of cell morphology. Furthermore, a significant increase in release of cytotoxic (LDH), oxidative stress (SOD-1, HMOX-1) or pro-inflammatory markers (TNF-α, IL-8) was absent. As a comparison, cells were exposed to Ag NPs in submerged conditions to 10, 20, and 30 µg Ag/mL. The deposited dose per surface area was estimated by using a dosimetry model (ISDD) to directly compare submerged vs ALI exposure concentrations after 4 and 24 h. Unlike ALI exposures, the two highest concentrations under submerged conditions promoted a cytotoxic and pro-inflammatory response after 24 h. Interestingly, when cell cultures were co-incubated with lipopolysaccharide (LPS), no synergistic inflammatory effects were observed. By using two different exposure scenarios it has been shown that the ALI as well as the suspension conditions for the lower concentrations after 4 h, reflecting real-life concentrations of an acute 24 h exposure, did not induce any adverse effects in a complex 3D model mimicking the human alveolar/airway barrier. However, the highest concentrations used in the ALI setup, as well as all concentrations under submerged conditions after 24 h, reflecting more of a chronic lifetime exposure concentration, showed cytotoxic as well as pro-inflammatory effects. In conclusion, more studies need to address long-term and chronic Ag NP exposure effects.

Hepatotoxicity of Dorema aucheri (Bilhar) in albino mice.

The geographic map of cancer prevalence differs due to environmental and dietary factors in various populations. High prevalence of a number of cancers in some regions is thought to be attributed to local dietary habits. Dorema aucheri (Bilhar) is used commonly as an herbal medicine in some regions including Iran. The aim of this study was to evaluate whether Dorema aucheri has carcinogenic effects in albino mice or not. The Dorema aucheri leaves were extracted by Soxhlet method and were injected intraperitoneally and randomly into 28 healthy albino mice which were divided into seven groups. One was put aside as the non-injected control group. The second control group was chosen to be injected by a known carcinogen. Another group was injected by carcinogen and then, Bilhar extract. The left four groups were injected the extracts in a dose- dependent manner, increasingly in the range of 0.4 - 3.2mL/kg. Extract injections were repeated every 48- hour intervals for three times. Then, liver and serum samples were analyzed biochemically and pathologically. The pathologic and biochemical studies showed that the injection of plant extracts caused necrosis, inflammation of the liver tissue, cell proliferation, cholestasis, and there were significant increases in release of liver enzymes [ALP, ALT (SGPT) and AST (SGOT)] and bilirubin compared to the non-injected control group. The level of liver damage was dose dependent. Dorema aucheri has potential hepatotoxic capacities and possibly this may be related to the high prevalence of cancer in some regions of Iran.

The Use of Natural Biopolymer of Chitosan as Biodegradable Beads for Local Antibiotic Delivery: Release Studies

Chitosan is a naturally occurring biopolymer which has been widely used in a variety of biomedical applications including local antibiotic delivery due to its excellent mechanical properties, biodegradability and biocompatibility. Beads are spherical, porous carriers which are prepared from various materials including chitosan. The current study aimed to fabricate a new controlled delivery system for local anti-infective treatment and to study its release behavior. Twenty beads were prepared from 1% or 2% chitosan solutions and immersed in vancomycin (VM) or teicoplanin (TN) solutions. The antibiotic release kinetics was determined by linear regression analysis supposing first order kinetics. Immersion for 3 h resulted in significant increase in the total TN release that differed from 0.5 h of immersion, except for the 1% beads immersed in VM. Increasing the chitosan concentration significantly increased the total release and antibiotic load of beads. The release of TN was more delayed compared to that of VM, which allowed a gradual release beyond 3 days. The half-life (mean ± SEM) of both types of TN-containing beads was significantly extended for 3 h immersion in comparison to 0.5 h immersion (26.1 ± 5.9 vs 10.9 ± 1.0 and 17.0 ± 2.1 vs 5.1 ± 1.9; P < 0.001). However, neither increasing the chitosan concentration, nor immersion time did result in any significant increase in the release of VM. The current study demonstrated an improved control of TN release impregnated in beads. It can be concluded that chitosan beads might be considered as a novel carrier for TN delivery to infected bone for local anti-infective therapy.

The Use of Natural Biopolymer of Chitosan as Biodegradable Beads for Local Antibiotic Delivery: Release Studies

Background: Chitosan is a naturally occurring biopolymer which has been widely used in a variety of biomedical applications including local antibiotic delivery due to its excellent mechanical properties, biodegradability and biocompatibility. Beads are spherical, porous carriers which are prepared from various materials including chitosan. Objectives: The current study aimed to fabricate a new controlled delivery system for local anti-infective treatment and to study its release behavior. Materials and Methods: Twenty beads were prepared from 1% or 2% chitosan solutions and immersed in vancomycin (VM) or teicoplanin (TN) solutions. The antibiotic release kinetics was determined by linear regression analysis supposing first order kinetics. Results: Immersion for 3 h resulted in significant increase in the total TN release that differed from 0.5 h of immersion, except for the 1% beads immersed in VM. Increasing the chitosan concentration significantly increased the total release and antibiotic load of beads. The release of TN was more delayed compared to that of VM, which allowed a gradual release beyond 3 days. The half-life (mean ± SEM) of both types of TN-containing beads was significantly extended for 3 h immersion in comparison to 0.5 h immersion (26.1 ± 5.9 vs 10.9 ± 1.0 and 17.0 ± 2.1 vs 5.1 ± 1.9; P < 0.001). However, neither increasing the chitosan concentration, nor immersion time did result in any significant increase in the release of VM. Conclusions: The current study demonstrated an improved control of TN release impregnated in beads. It can be concluded that chitosan beads might be considered as a novel carrier for TN delivery to infected bone for local anti-infective therapy.

C-Reactive Protein Induces Release of Both Endothelial Microparticles and Circulating Endothelial Cells In Vitro and In Vivo: Further Evidence of Endothelial Dysfunction

Inflammation is pivotal in atherosclerosis. A key early event in atherosclerosis is endothelial dysfunction. C-reactive protein (CRP), the prototypic marker of inflammation in humans, is a risk marker for cardiovascular disease, and there is mounting evidence to support its role in atherothrombosis. CRP has been shown to promote endothelial dysfunction both in vitro and in vivo. Emerging biomarkers of endothelial dysfunction include circulating endothelial cells (CECs) and endothelial microparticles (EMPs). However, there is a paucity of data examining the effect of CRP on CEC and EMP production in vitro and in vivo. In this report, we treated human aortic endothelial cells (HAECs) with increasing concentrations of CRP (0-50 μg/mL) or boiled CRP. We counted CECs and EMPs by flow cytometry. Although CRP treatment resulted in a significant increase in release of both CECs and EMPs, boiled CRP failed to have an effect. Pretreatment of HAECs with sepiapterin or diethylenetriamine NONOate, both of which preserve nitric oxide (NO), resulted in attenuation of CRP's effects on CECs and EMPs. CD32 and CD64 blocking antibodies but not CD16 antibody or lectin-like oxidized LDL receptor 1 small interfering RNA (LOX-1 siRNA) prevented CRP-induced production of CECs and EMPs. Furthermore, delivery of human CRP to Wistar rats compared with human serum albumin resulted in significantly increased CECs and EMPs, corroborating the in vitro findings. We provide novel data that CRP, via NO deficiency, promotes endothelial dysfunction by inducing release of CECs and EMPs, which are biomarkers of endothelial dysfunction.

An Atypical Role for Collapsin Response Mediator Protein 2 (CRMP-2) in Neurotransmitter Release via Interaction with Presynaptic Voltage-gated Calcium Channels

Collapsin response mediator proteins (CRMPs) specify axon/dendrite fate and axonal growth of neurons through protein-protein interactions. Their functions in presynaptic biology remain unknown. Here, we identify the presynaptic N-type Ca(2+) channel (CaV2.2) as a CRMP-2-interacting protein. CRMP-2 binds directly to CaV2.2 in two regions: the channel domain I-II intracellular loop and the distal C terminus. Both proteins co-localize within presynaptic sites in hippocampal neurons. Overexpression in hippocampal neurons of a CRMP-2 protein fused to enhanced green fluorescent protein caused a significant increase in Ca(2+) channel current density, whereas lentivirus-mediated CRMP-2 knockdown abolished this effect. Interestingly, the increase in Ca(2+) current density was not due to a change in channel gating. Rather, cell surface biotinylation studies showed an increased number of CaV2.2 at the cell surface in CRMP-2-overexpressing neurons. These neurons also exhibited a significant increase in vesicular release in response to a depolarizing stimulus. Depolarization of CRMP-2-enhanced green fluorescent protein-overexpressing neurons elicited a significant increase in release of glutamate compared with control neurons. Toxin block of Ca(2+) entry via CaV2.2 abolished this stimulated release. Thus, the CRMP-2-Ca(2+) channel interaction represents a novel mechanism for modulation of Ca(2+) influx into nerve terminals and, hence, of synaptic strength.

Flavonoid Treatment of Murine Macrophages Does Not Alter Ingestion or Intracellular Survival of Listeria monocytogenes

It was recently reported that the aryl hydrocarbon receptor (AhR) plays a role in innate immunity to Listeria monocytogenes infection in mice. In this study the authors analyzed whether incubation of murine macrophages with various flavonoids that have agonist or antagonist activity for the AhR (β-naphthoflavone, α-naphthoflavone, quercetin, luteolin) affect the uptake and survival of L. monocytogenes. Using the RAW264.7 and J774 murine macrophage cell lines and bone marrow–derived macrophages, the authors found no significant difference between flavonoid-treated and control macrophages. They also found that macrophages incubated with flavonoids alone did not exhibit a significant increase in release of tumor necrosis factor (TNF)-α, a crucial cytokine in anti-Listeria resistance.

Biological roles of monoterpene volatiles derived from rough lemon (Citrus jambhiri Lush) in citrus defense

Volatile compounds of plants, including monoterpenes, are a possible source of signal molecules that induce defense systems to protect plants from tissue damage. Volatile compounds from rough lemon leaves were trapped by solid-phase microextraction fibers in sealed vials, and subsequent gas chromatography–mass spectrometry and gas chromatography analyses identified the profile of the major components, mainly various monoterpenes. Among several monoterpenes examined, citral, citronellal, and linalool significantly inhibited the spore germination and hyphal growth of Alternaria alternata. The effect of linalool was fungistatic, while the effects of citral and citronellal were partially fungicidal. Wounding of rough lemon leaves induced a significant increase in release of monoterpenes. The release of linalool was the most abundant and was 14.5 times that of unwounded rough lemon leaves. Unlike the wounding treatment, microbe attack did not significantly change monoterpene releases, and there was statistically no difference found in the peak areas from microbe-treated and untreated leaves. Linalool, limonene, and β-pinene also had insect-repellant effects on wild-type Drosophila melanogaster. Expression patterns of defense-related genes in rough lemon and rice significantly changed after treatment with vapors of monoterpene volatiles. Taking these results together, monoterpene volatiles are likely to play roles in the defense of rough lemon against microbe and insect pathogens.

Lack of Effect of Oral 4‐Phenylbutyrate on Serum Alpha‐1‐Antitrypsin in Patients with α‐1‐Antitrypsin Deficiency: A Preliminary Study

ABSTRACTObjective:In homozygotes with ZZ genotype alpha‐1‐antitrypsin (α1AT) deficiency, mutant α1ATZ protein (α1ATZ) accumulates in hepatocytes, rather than being secreted into the blood. Homozygous individuals experience emphysema as a result of reduced levels of circulating α1AT in the lung with which to inhibit connective tissue breakdown. Homozygotes may also experience liver disease from the accumulation of α1ATZ within hepatocytes, which causes liver damage. A previous study indicated that the compound 4‐phenylbutyrate (4‐PBA) mediated a significant increase in release of α1ATZ from cells in tissue culture and in a mouse model of α1AT deficiency. The authors hypothesized that 4‐PBA could be used to treat both the liver and lung disease of humans with α1AT deficiency.Methods:In this preliminary, open label study the authors evaluated the effect of 14 days of oral 4‐PBA therapy on α1AT blood levels in 10 patients with α1AT deficiency.Results:There was no significant increase in α1AT blood level associated with 4‐PBA administration. Symptomatic and metabolic side effects were significant.Conclusion:4‐PBA did not increase α1AT blood levels in humans with α1AT deficiency in this preliminary trial.

Lack of effect of oral 4-phenylbutyrate on serum alpha-1-antitrypsin in patients with alpha-1-antitrypsin deficiency: a preliminary study.

In homozygotes with ZZ genotype alpha-1-antitrypsin (alpha1AT) deficiency, mutant alpha1ATZ protein (alpha1ATZ) accumulates in hepatocytes, rather than being secreted into the blood. Homozygous individuals experience emphysema as a result of reduced levels of circulating alpha1AT in the lung with which to inhibit connective tissue breakdown. Homozygotes may also experience liver disease from the accumulation of alpha1ATZ within hepatocytes, which causes liver damage. A previous study indicated that the compound 4-phenylbutyrate (4-PBA) mediated a significant increase in release of alpha1ATZ from cells in tissue culture and in a mouse model of alpha1AT deficiency. The authors hypothesized that 4-PBA could be used to treat both the liver and lung disease of humans with alpha1AT deficiency. In this preliminary, open label study the authors evaluated the effect of 14 days of oral 4-PBA therapy on alpha1AT blood levels in 10 patients with alpha1AT deficiency. There was no significant increase in alpha1AT blood level associated with 4-PBA administration. Symptomatic and metabolic side effects were significant. 4-PBA did not increase alpha1AT blood levels in humans with alpha1AT deficiency in this preliminary trial.

Activities of extract and constituents of Banisteriopsis caapi relevant to parkinsonism

Dopamine deficiency is characteristic of Parkinson's disease (PD) and treatments aim at elevating levels by administration of its precursor l-dihydroxyphenylalanine ( l-DOPA), or inhibiting monoamine oxidases (MAOs), thus preventing its breakdown. Reports of improvements in PD patients treated with Banisteriopsis caapi extracts stimulated investigation of B. caapi stem extract and its two ingredients, harmine and harmaline for these activities. Tests for MAO inhibition using liver homogenate showed that extract and harmaline showed a concentration-dependent inhibition of MAO A (IC 50 1.24 μg/ml and IC 50 4.54 nM, respectively) but had little effect on MAO B activity. The extract at 2.5 mg/ml caused a highly significant increase in release of [ 3H]dopamine from rat striatal slices, as did 200 μM harmine and 6 μM harmaline. In both these experiments, the amount of harmine present could not account for the total activity of the extract. The ability of harmine and harmaline to stimulate dopamine release is a novel finding. These results give some basis to the reputed usefulness of B. caapi stem extract in the treatment of PD.

Complex array of cytokines released by vasoactive intestinal peptide

A complex mixture of five cytokines has been shown to be released by vasoactive intestinal peptide (VIP). Cytokines were measured in paired samples of culture medium and astroglial cytosol by capillary electrophoresis. This is the first description of VIP-mediated release for TNF-α, IL-3, G-CSF and M-CSF from astrocyte cultures. Kinetic studies after VIP treatment demonstrated a gradual but incomplete depletion of cytosolic cytokine levels, with differences observed among the cytokines. Significant increases in release were apparent within 15–30 min for all cytokines. As the recognized VIP receptors (VPAC1 and VPAC2) are linked to adenylate cyclase and also interact with pituitary adenylate cyclase activating polypeptide-38 (PACAP-38), both this homologous peptide and 8-bromo cAMP were investigated and compared to VIP-mediated release. Treatment with 1 mM 8-bromo cAMP produced cytokine release similar in amount to 0.1 nM PACAP-38, but significantly less (<50%) in comparison to 0.1 nM VIP. PACAP-38 and VIP exhibited similar EC 50’s for the release of G-CSF and TNF-α; however, the maximal release was 4–6 times greater for VIP than for PACAP-38. This similarity in potency suggested a VPAC-like receptor; however, the greater efficacy for VIP in comparison to PACAP-38, combined with a lack of cAMP production at subnanomolar concentrations of VIP, suggested a mechanism not currently associated with VPAC receptors. For M-CSF, IL-3 and IL-6, the EC 50’s of VIP were 3–30 times more potent than those of PACAP-38 in producing release. These studies suggested that multiple mechanisms mediate cytokine release in astrocytes: (1) a low efficacy release produced by PACAP-38 that is cAMP-mediated and (2) a high efficacy, VIP-preferring mechanism that was not linked to cAMP. In summary, subnanomolar concentrations of VIP released a complex array of cytokines from astrocytes that may contribute to the mitogenic and neurotrophic properties of this neuropeptide in the central nervous system.