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Chinese herbal medicines (CHMs) are classified as dietary supplements. Interactions with western medications, the presence of contaminants or adulterants, or a mis-labeled or mis-used CHM may lead to toxicological emergencies that can be undetected in death investigations. Laboratories must be able to efficiently analyze cases in which CHMs are suspected. Five extractions were evaluated for their ability to extract pharmacologically active compounds from herbal matrices: water, ethanol, microwave-assisted (MAE), ethanol : chloroform, and acid-wash. Anticonvulsive and other pharmacologically active compounds in Gou Teng, Tian Ma, and Jiang Can purchased from Beijing, China and New York were compared in the powder and the extracts using Direct Analysis in Real Time-Mass Spectrometry (DART-MS). Approximately 0.25 g of macerated herb was used per extraction. The water and ethanol extractions were simple liquid extractions. For the MAE, powdered herb was soaked in 65% ethanol, microwaved, and concentrated. The ethanol : chloroform extraction involved soaking in 1 : 1 ethanol : chloroform, sonication, and concentration. In the acid-wash extraction, powdered herb was soaked in acetic acid, followed by addition of sodium hydroxide, hexane extraction, and reconstitution in ethyl acetate. The powdered herbs and extracts were analyzed using a Jeol JMS T100LC AccuTOF DART-MS in positive and negative mode. Of the evaluated methods, no single extraction worked for all active compounds from the three CHMs. The MAE extract contained the most pharmacologically active compounds, while the acid-wash contained the least for the three products. Gou Teng purchased from different sources did exhibit a difference in pharmacologically active compounds, potentially from different species., With the popularity of natural products, rapid analysis of pharmacologically active analytes from natural matrices is critical for public health. Microwave-assisted extraction with DART-MS provides efficient analysis of analytes from herbal matrices.

Cannabinoid-based products submitted by consumers experiencing adverse effects were analyzed to identify and quantitate ingredients. Product testing identified several synthetic cannabinoids and products with inaccurate or incomplete labeling.

Additional file 1: Supplemental tables and figures.

INTRODUCTION: Electronic cigarettes (e-cigarettes) have rapidly evolved since their introduction to the U.S. market. The rebuildable atomizer (RBA) offers user-driven modification to the heating element (coil) and wicking systems. Different coil materials can be chosen, based on user needs and preferences. However, the heating element of an e-cigarette is believed to be one-source for toxic metal exposure. METHODS: E-cigarette coils from Kanthal and nichrome wires were constructed in a contact and non-contact configuration and heated at four voltages. The maximum temperatures of the coils were measured by infrared temperature sensing when dry and saturated with 100% vegetable glycerin or 100% propylene glycol. The metal composition of each coil was analyzed with Scanning Electron Microscopy-Energy Dispersive X-Ray (SEM-EDX) when new, and subsequently after 1, 50, and 150 heat cycles when dry. RESULTS: The coils reached temperatures above 1000 °C when dry, but were below 300 °C in both liquid-saturated mediums. Metal analysis showed a decrease of 9–19% chromium and 39–58% iron in Kanthal wire and a decrease of 12–14% iron and 39–43% nickel in nichrome wire after 150 heat cycles. Significant metal loss was observed after one heat cycle for both coil alloys and configurations. CONCLUSIONS: The loss of metals from these heat cycles further suggests that the metals from the coils are potentially entering the aerosol of the e-cigarette, which can be inhaled by the user.

Rational Pressure overload induces adaptive and maladaptive cardiac remodeling processes in the heart. Part of the maladaptive process is the cross-talk between cardiomyocytes and macrophages which is dependent on the function of the Activating Transcription Factor 3, ATF3. Yet, the molecular mechanism involved in cardiomyocytes-macrophages communication leading to macrophages recruitment to the heart and cardiac maladaptive remodeling is currently unknown. Methods and results Isolated peritoneal macrophages from either wild type or ATF3-KO mice were cultured in serum free medium to collect conditioned medium (CM). CM was used to probe an antibody cytokine/chemokine array. The interferon γ induced protein 10kDa, CXCL10, was found to be enriched in wild type macrophages CM. Wild type cardiomyocytes treated with CXCL10 in vitro, resulted in significant increase in cell volume as compared to ATF3-KO cardiomyocytes. In vivo, pressure overload was induced by phenylephrine (PE) infusion using micro-osmotic pumps. Consistently, CXCL11 (CXCL10 competitive agonist) and CXCL10 receptor antagonist (AMG487) attenuated PE-dependent maladaptive cardiac remodeling. Significantly, we show that the expression of the CXCL10 receptor, CXCR3, is suppressed in cardiomyocytes and macrophages derived from ATF3-KO mice. CXCR3 is positively regulated by ATF3 through an ATF3 transcription response element found in its proximal promoter. Finally, mice lacking CXCR3 display a significant reduction of cardiac remodeling processes following PE infusion. Conclusions Chronic PE infusion results in a unique cardiomyocytes-macrophages cross-talk that is mediated by IFNγ. Subsequently, macrophages that are recruited to the heart secrete CXCL10 resulting in maladaptive cardiac remodeling mediated by the CXCR3 receptor. ATF3-KO mice escape from PE-dependent maladaptive cardiac remodeling by suppressing the IFNγ-CXCL10-CXCR3 axis at multiple levels.

Karin Wåhlén, Patrik Olausson, Anders Carlsson, Nazdar Ghafouri, Björn Gerdle, Bijar Ghafouri Pain and Rehabilitation Centre, Department of Medical and Health Sciences, Linköping University, Linköping, Sweden Abstract: Chronic widespread pain (CWP) is a complex pain condition that is difficult to treat. The prevalence of CWP approximates ~10% of the general population, with higher prevalence in women. Lack of understanding of molecular mechanisms has been a challenge for diagnosis and treatment of chronic pain. The aim of this study was to explore the systemic protein changes in CWP compared to those in healthy controls (CON). By applying 2-dimensional gel electrophoresis, we analyzed the protein pattern of plasma samples from women with CWP (n=16) and healthy women (n=23). The proteomic data were analyzed using multivariate statistical models, and altered proteins were identified using mass spectrometry. The proteome analysis was further validated by gel-free Western blot. Multivariate statistical data analysis of quantified proteins revealed 22 altered proteins in women with CWP, compared to CON group. Many of the identified proteins are previously known to be involved in different parts of the complement system and metabolic and inflammatory processes, e.g., complement factor B, vitamin D-binding protein, ceruloplasmin, transthyretin and alpha-2-HS-glycoprotein. These results indicate that important systemic protein differences exist between women with CWP and healthy women. Further, this study illustrates the potential use of proteomics to detect biomarkers that may provide new insights into the molecular mechanism(s) of chronic pain. However, further larger investigations are required in order to confirm these findings before it will be possible to identify proteins as potential pain biomarkers for clinical use. Keywords: inflammation, biomarker, painomics, complement system, GC protein

DNA vaccination represents a novel means of expressing Ag in vivo for the generation of both humoral and cellular immune responses. The current study uses this technology to elicit protective immunity against experimental autoimmune encephalomyelitis (EAE), a T cell-mediated autoimmune disease of the central nervous system that serves as an experimental model for multiple sclerosis. RT-PCR verified by Southern blotting and sequencing of PCR products of four different C-C chemokines, macrophage-inflammatory protein-1α (MIP-1α), monocyte-chemotactic protein-1 (MCP-1), MIP-1β, and RANTES, were performed on brain samples from EAE rats to evaluate mRNA transcription at different stages of disease. Each PCR product was then used as a construct for naked DNA vaccination. The subsequent in vivo immune response to MIP-1α or MCP-1 DNA vaccines prevented EAE, even if disease was induced 2 mo after administration of naked DNA vaccines. In contrast, administration of the MIP-1β naked DNA significantly aggravated the disease. Generation of in vivo immune response to RANTES naked DNA had no notable effect on EAE. MIP-1α, MCP-1, and MIP-1β mRNA transcription in EAE brains peaked at the onset of disease and declined during its remission, whereas RANTES transcription increased in EAE brains only following recovery. Immunization of CFA without the encephalitogenic epitope did not elicit the anti-C-C chemokine regulatory response in DNA-vaccinated rats. Thus, modulation of EAE with C-C chemokine DNA vaccines is dependent on targeting chemokines that are highly transcribed at the site of inflammation at the onset of disease.

The diversity of Ag-specific receptors on T cells homing to an inflammatory infiltrate in the central nervous system has been analyzed. Experimental autoimmune encephalomyelitis, a T cell-mediated inflammatory disease of the central nervous system, was induced in Lewis rats with a CD4+, CD8- T cell line specific for peptide 68-86 of myelin basic protein. Within the line a wide array of TCR V beta genes was transcribed including the V beta 8, V beta 10, V beta 15, V beta 16, and V beta 19 families. Accumulation of T cells at the site of inflammation was determined by using RNA-polymerase chain reaction amplification of rearranged TCR V beta transcripts derived from brain. By 8 to 10 h after i.p. infusion of the pathogenic T cell line, TCR V beta transcripts, including mainly V beta families that were predominantly rearranged by the line, could be identified in brains. Restricted TCR V gene transcripts with predominance of the V beta 8 family were identified in brain 48 h after injection, before onset of disease. Paralysis was apparent by 4 to 5 days after injection. At this time diverse V beta gene transcripts were detected in brain, reaching a maximum by day 9, when paralyzed rats have recovered. By day 14 a second stage of limited heterogeneity in the T cell infiltrate could be identified with predominant expression of V beta 8, V beta 9, V beta 10, and V beta 19. Interestingly, three out of these four V beta families were predominantly expressed within the encephalitogenic line. Thus, T cell migration to brain in experimental autoimmune encephalomyelitis is characterized by a rapid penetration of T cells followed by a selective trapping of T cells before the clinical manifestations of disease. When clinical disease was present the T cell infiltrate was diverse, whereas in the post-acute phase of disease the T cells in the central nervous system had limited heterogeneity with selective accumulation of T cells transcribing the same V regions that were detected in the line that incited disease.

"A changing climate will have increasingly grave consequences for the world???s poorest, most vulnerable people. Who says so? The intergovernmental Panel on Climate Change (IPCC) and the 2007/2008 Human Development Report (HDR), Fighting Climate Change: Human Solidarity in a Divided World."

Naked DNA encoding TNFalpha was introduced to BALB/c mice with experimental antiphospholipid syndrome (APS) induced by beta2GPI. Administration of naked DNA encoding TNFalpha resulted in the generation of immunological memory to its gene product, associated with elevated circulating anti-TNFalpha antibodies. Enriched IgG fraction of the mouse anti-TNFalpha was biologically active since it prevented endothelial cell activation by TNFalpha e.g., inhibition of monocyte adhesion to activated endothelial cells (HUVEC). Mice immunized with beta2GPI, vaccinated with TNFalpha DNA at an early stage of disease development, showed decreased titres of circulating anti-beta2GPI antibodies as compared to the group of mice vaccinated with a control naked DNA. The reduction of antiphospholipid antibody production was followed by amelioration of the foetal loss, increased platelet count to normal values as well as normalization of the prolonged aPTT. APS mice which were introduced to the TNFalpha DNA vector at a later stage of the disease development, showed less improvement in their clinical manifestations. The current study suggests a way in which a DNA vaccine can be employed for induction of a protective immunity in experimental APS.

We recently used a modification of gene therapy (naked DNA vaccination) to induce immunological memory against self-pro-inflammatory chemokines such as macrophage inflammatory protein-1 alpha or monocyte chemoattractant protein-1, and against the pro-inflammatory cytokine tumor necrosis factor-alpha. First, DNA constructs encoding each of the different pro-inflammatory mediators together with a repeated immunostimulatory sequence were prepared. Then, experiment animals were subjected to a repeated administration of each construct. Under these conditions, tolerance to the product of each insert was broken, and immunological memory established. Upon induction of experimental autoimmune encephalomyelitis, a T cell-mediated autoimmune disease of the central nervous system that serves as a model for multiple sclerosis or adjuvant induced arthritis, this memory was "turned on" to provide DNA-vaccinated animals a high state of disease resistance. Antibodies to the product of each inserted gene were isolated form these animals. Each antibody was found capable of neutralizing in vitro the chemoattractive properties of each relevant chemokine, thereby transferring disease resistance. Interestingly, the level of their production was dependent on disease severity, that is, each titer was accelerated in accordance with disease progression. Thus, by using a simple gene therapy technique the immune system could be "re-educated" to restrain its own harmful activities.

TNF-alpha is thought to be a key pro-inflammatory cytokine in T cell-mediated autoimmune diseases, particularly in rheumatoid arthritis (RA) and multiple sclerosis (MS). Experimental autoimmune encephalomyelitis (EAE) serves as an animal model for MS. The current study observes a notable TNF-alpha-specific antibody titer generated during the course of EAE, apparently not sufficient to prevent the development of disease. Administration of TNF-alpha-naked DNA vaccine enhanced the production of TNF-alpha-specific antibody titer and conferred EAE resistance. These antibodies were found to be neutralizing in vitro and capable of inhibiting the development of disease when transferred to other EAE rats. Thus, modulation of EAE with TNF-alpha DNA vaccines enhances the regulation of natural immunity to a self pro-inflammatory cytokine and provides a tool by which the immune system is encouraged to elicit anti-self protective immunity to restrain its own harmful reactivity when such a response is needed.

Specific oligonucleotide primers were used to identify and isolate IFN-gamma-inducing factor (IGIF) from the brain of rats with developing experimental autoimmune encephalomyelitis (EAE), a T cell-mediated autoimmune disease of the central nervous system that serves as a model for multiple sclerosis. IGIF was highly transcribed in the brain at the onset and during the course of active EAE. PCR products encoding rat IGIF were used to generate the recombinant protein that was used to induce anti-IGIF neutralizing Abs. These Abs significantly reduced the production of IFN-gamma by primed T cells proliferating in response to their target myelin basic protein epitope and by Con A-activated T cells from naive donors. When administered to rats during the development of either active or transferred EAE, these Abs significantly blocked the development of disease. Splenic T cells from protected rats were cultured with the encephalitogenic myelin basic protein epitope and evaluated for production of IL-4 and IFN-gamma. These cells, which proliferated, exhibited a profound increase in IL-4 production that was accompanied by a significant decrease in IFN-gamma and TNF-alpha production. Thus, we suggest that perturbation of the Th1/Th2 balance toward Th2 cells is the mechanism underlying EAE blockade by anti-IGIF immunotherapy.

The diversity of Ag-specific receptors on T cells homing to an inflammatory infiltrate in the central nervous system has been analyzed. Experimental autoimmune encephalomyelitis, a T cell-mediated inflammatory disease of the central nervous system, was induced in Lewis rats with a CD4+, CD8- T cell line specific for peptide 68-86 of myelin basic protein. Within the line a wide array of TCR V beta genes was transcribed including the V beta 8, V beta 10, V beta 15, V beta 16, and V beta 19 families. Accumulation of T cells at the site of inflammation was determined by using RNA-polymerase chain reaction amplification of rearranged TCR V beta transcripts derived from brain. By 8 to 10 h after i.p. infusion of the pathogenic T cell line, TCR V beta transcripts, including mainly V beta families that were predominantly rearranged by the line, could be identified in brains. Restricted TCR V gene transcripts with predominance of the V beta 8 family were identified in brain 48 h after injection, before onset of disease. Paralysis was apparent by 4 to 5 days after injection. At this time diverse V beta gene transcripts were detected in brain, reaching a maximum by day 9, when paralyzed rats have recovered. By day 14 a second stage of limited heterogeneity in the T cell infiltrate could be identified with predominant expression of V beta 8, V beta 9, V beta 10, and V beta 19. Interestingly, three out of these four V beta families were predominantly expressed within the encephalitogenic line. Thus, T cell migration to brain in experimental autoimmune encephalomyelitis is characterized by a rapid penetration of T cells followed by a selective trapping of T cells before the clinical manifestations of disease. When clinical disease was present the T cell infiltrate was diverse, whereas in the post-acute phase of disease the T cells in the central nervous system had limited heterogeneity with selective accumulation of T cells transcribing the same V regions that were detected in the line that incited disease.

Electrolytic lesions of the baso-medial-hypothalamus (BMH) cause numerous dysfunctions of the endocrine system, many of which have been implicated in regulating immune responses. The objective of this report was to evaluate the direct effects of electrolytic lesions in the BMH on the immune response of the chicken. Our results indicate that temporary immune response impairment, as evaluated by both T and B lymphocyte related responses, occurred only in birds that expressed both brain damage and endocrine dysfunction, whereas birds expressing brain damage alone had normal immune responses. The possible regulation of immune responses by the BMH via the endocrine system is discussed.

In this paper the application of Quasi Classical Approach to the analysis of distillation columns with dual composition control is presented. The method relies heavily on a computer graphic package because it retains the basic fundaments of classical control theory such as Myquist and Bode plots. Firstly the main measurements and properties in which the methodology is based are briefly described. Then the advantages of using reserved frame controllers is discussed and through the example it is shown that they can handle the principal aspects of a control system. The experimental model of the| column, TOFA, has been chosen to show the applicability of the methodology.

In this paper the application of Quasi Classical Approach to the analysis of distillation columns with dual composition control is presented. The method relies heavily on a computer graphic package because it retains the basic fundaments of classical control theory such as Nyquist and Bode plots. Firstly the main measurements and properties in which the methodology is based are briefly described. Then the advantages of using reserved frame controllers is discussed and through the example it is shown that they can handle the principal aspects of a control system. The experimental model of the column, TOFA, has been chosen to show the applicability of the methodology.

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory condition of the central nervous system with similarities to multiple sclerosis. In both diseases, circulating leukocytes penetrate the blood-brain barrier and damage myelin, resulting in impaired nerve conduction and paralysis. We sought to identify the adhesion receptors that mediate the attachment of circulating leukocytes to inflamed brain endothelium in EAE, because this interaction is the first step in leukocyte entry into the central nervous system. Using an in vitro adhesion assay on tissue sections, we found that lymphocytes and monocytes bound selectively to inflamed EAE brain vessels. Binding was inhibited by antibodies against the integrin molecule alpha 4 beta 1, but not by antibodies against numerous other adhesion receptors. When tested in vivo, anti-alpha 4 integrin effectively prevented the accumulation of leukocytes in the central nervous system and the development of EAE. Thus, therapies designed to interfere with alpha 4 beta 1 integrin may be useful in treating inflammatory diseases of the central nervous system, such as multiple sclerosis.