Induction of Aminolevulinic Acid Synthase Gene Expression

Induction of Aminolevulinic Acid Synthase Gene Expression Induction of aminolevulinic acid synthase gene expression, down-regulation ferrochelatase and enhancement of metabolite, protoporphyrin IX, excretion by co-therapy with isoniazid and rifampicin (1. Isoniazid and rifampicin induced liver injury by regulating 5-aminolevulinate synthase and ferrochelatase and enhancing protoporphyrin IX 2. Mechanism of rifampicin and isoniazid induced cell death in L-02 cell line and mice) Abstract Isoniazid(INH) and rifampicin(RFP) are first-line antituberculosis drugs, co-therapy with INH and RFP is highly effective. However, the combination of these two drugs frequently cause liver injury or liver failure in humans. The risk of hepatotoxicity is considerably higher in patients receiving both RFP and INH than in those receiving either RFP or INH alone. Numerous studies have been conducted to investigate the mechanism of injury after isoniazid or rifampicin used in various animal models, however, the important mechanism for the combination of isoniazid and rifampicin in humans remains unclear. Here we investigated this combination induced hepatotoxicity using L-02 cells and mice. Introduction Tuberculosis remains a global public health problem whose effects have major impact in developing countries. World Health Organization estimates that there were 8.6 million new TB cases in 2012 and 1.3 million TB deaths. The currently recommended treatment for new cases of drug-susceptible TB is a six-month regimen of four first-line drugs: isoniazid, rifampicin, ethambutol and pyrazinamide. (Global tuberculosis report 2013). However, the combination of isoniazid(INH) and rifampicin(RFP) frequently cause liver injury or liver failure. The risk of hepatotoxicity is considerably higher in patients receiving the combination than in those receiving either RFP or INH alone. The mechanisms leading to liver failure in humans were poorly understood. Recently, a new mechanism ,independent of INH metabolism, is found in the RFP and INH co-therapy induced liver injury. Li et al. (Li, et al. 2013) found that co-therapy with RFP and INH targets porphyrin biosynthesis and results in hepatic protoporphyrin IX (PPIX) accumulation and liver injury . PPIX is an intermediate in porphyrin biasynthesis. Normally the concentrations of PPIX is very low in the liver. However, in some cases the concentration abnormally elevated in blood and liver, such as erythropoietic protoporphyria. High concentrations of PPIX in the liver are known to cause liver injury (Anstey and Hift 2007; Casanova-Gonzalez, et al. 2010). Using hPXR mice, Li et al. demonstrated that the accumulation of endogenous PPIX is through PXR-mediated transcriptional activations of aminolevulinic synthase-1(ALAS1) genes. ALAS1 is the rate-limiting enzyme of heme synthesis in the liver and is drug-responsive, providing heme for CYPs and other hemoproteinsis. Activation of PXR can up regulate ALAS1 expression in liver (Fraser, et al. 2003). RFP upregulate ALAs1 increasing heme-biosynthesis in the liver and overproducing PPIX through activating PXR signalling pathway. However, PPIX accumulation strongly suggests that ferrochelatase became a ratelimiting enzyme during INH-RFP treatment (Lyoumi, et al. 2013). Ferrochelatase (FECH) ,the final enzyme in the heme biosynthetic pathway, catalyses ferrous iron inserted into precursor porphyrin protoporphyrin IX to form heme, and when defective or deficient, causing accumulation of protoporphyrin IX. Ferrochelatase is active in cells that produce 80% heme in the bone marrow (Bloomer, et al. 1991) and the rest in hepatocytes (Bonkowsky, et al. 1975). The excess protoporphyrinIX becomes insoluble in bile and exerts cholestatic effects leading to architectural changes in the hepatobiliary system ranging from mild inflammation to fibrosis and cirrhosis (Anstey and Hift 2007). MATERIALS AND METHODS PI staining L-02 were allowed to adhere on glass bottom dishs for 4h, followed by INH,RFP or INH/RFP. The medium was removed after h and cells were stained with for 30 min. Nuclei were stained with 4’,6-diamidino-2-phenylindole (DAPI) and images were recorded with a fluorescence microscope. Western blotting L-02 cells cultured in flask were harvested using 0.25% trypsin (Hyclone, Thermo Scientific, Waltham, Mass). After centrifugation at 1000r and lysis using buffer for Western blotting (), total proteins were collected by following the kit instructions. Protein concentrations were determined using the BCA Protein Assay Kit (). After heating at 95 °C for 5 minutes in sample buffer, proteins were separated on SDS-PAGE using 10% polyacrylamide gels before electroblotting onto PVDFmembrane(). Nonspecific binding was blocked by incubation for 2 hours in 5% (w/v) nonfat milk. The following primary antibodies were used overnight at 4 °C: Rabbit anti-human FECH antibody(; 1:1000); Rabbit anti-human ALAs1 antibody( 1:500); Rabbit anti-human BCRP antibody(; 1:500). Bound antibodies were detected using horseradish peroxidase-conjugated secondary antibodies (Beijing Biosynthesis Biotechnology Co. LTD). Finally, the membranes were visualized by chemiluminescence. RNA Isolation and Real time Polymerase Chain Reaction for ALAs1 and FECH Cell Culture L-02 cells, a human fetal hepatocyte line, purchased from Cell Bank of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, purchased from Shanghai, China, were cultured according to the manufacturer’s instructions 15 at 37 °C in 5% CO2. Cell culture materials were procured from Corning() Discussion Nevertheless, the ability of chemicals to activate PXR is species dependent. RFP is a human PXR specific activator that weakly affect on mouse (Lehmann, et al. 1998). INH hepatotoxicity is thought to be dependent on metabolic activation by arylamine N-acetyltransferase and CYP2E1, but Li found AcHZ and hydrazine do not cause INH-related hepatotoxicity. Hepatic heme synthesis leading to protoporphyria and possible impact with other metabolic systems (Davies, et al. 2005). References Primary Sources Secondary Sources Uncategorized References Anstey, A. V., and R. J. Hift,  2007, Liver disease in erythropoietic protoporphyria: insights and implications for management. Postgrad Med J 83(986):739-48. Bloomer, J. R., et al.  1991, Heme synthesis in protoporphyria. Curr Probl Dermatol 20:135-47. Bonkowsky, H. L., et al. 1975, Heme synthetase deficiency in human protoporphyria. Demonstration of the defect in liver and cultured skin fibroblasts. J Clin Invest 56(5):1139-48. Casanova-Gonzalez, M. J., et al.  2010, Liver disease and erythropoietic protoporphyria: a concise review. World J Gastroenterol 16(36):4526-31. Davies, R., et al.  2005, Hepatic gene expression in protoporphyic Fech mice is associated with cholestatic injury but not a marked depletion of the heme regulatory pool. Am J Pathol 166(4):1041-53. Fraser, D. J., A. Zumsteg, and U. A. Meyer,  2003, Nuclear receptors constitutive androstane receptor and pregnane X receptor activate a drug-responsive enhancer of the murine 5-aminolevulinic acid synthase gene. J Biol Chem 278(41):39392-401. Lehmann, J. M., et al.  1998, The human orphan nuclear receptor PXR is activated by compounds that regulate CYP3A4 gene expression and cause drug interactions. J Clin Invest 102(5):1016-23. Li, F., et al.  2013, Human PXR modulates hepatotoxicity associated with rifampicin and isoniazid co-therapy. Nat Med 19(4):418-20. Lyoumi, S., et al.  2013, PXR-ALAS1: a key regulatory pathway in liver toxicity induced by isoniazid-rifampicin antituberculosis treatment. Clin Res Hepatol Gastroenterol 37(5):439-41.

Importunate Persuasions: Anxiety of Authorship and the Female Fight for Self-Sovereignty

In Margaret Cavendish’s essay The Blazing World she creates a world of her own where she rules as a sovereign and is afforded a power that would not otherwise be possible for her: â€Å"†¦if any should like the world I have made and be willing to be my subjects, they may imagine themselves such, and they are such, I mean in their minds†¦; but if they cannot endure to be subjects, they may create worlds of their own and govern themselves as they please† (1785).In this passage, Margaret Cavendish exposes her own idiosyncratic philosophy concerning her position in society as a woman, female author and a member of a court that was ostracized. By prolifically writing about herself, she attempts to exercise her right to a voice and uses it as an instrument of power and resistance in an oppressed and powerless situation. However, her language bears traces of an internalization of the oppressive social structure and an anxiety of authorship1 that prevents her from suc cessfully establishing herself as autonomous.In this essay, I will attempt to demonstrate how Margaret Cavendish, through her poetry and prose, endeavors to achieve self-sovereignty through singularity but fails due to fear of social alienation from not just the patriarchal hegemony but also from the women of her era that perpetuated it. In The Poetess’s Hasty Resolution, Margaret Cavendish establishes herself as not only a poet but a gifted one at that. â€Å"Reading my verses, I liked them so well/Self-love did make my judgment to rebel/Thinking them so good, I thought more to write† (1-3).Here, Margaret introduces her desire for self-sovereignty and her initial willingness to exercise it through the vocation of writing. She writes of a â€Å"self-love† initiated by the sound of her own voice and empowers her to fight against the status quo, â€Å"to rebel†. She decides to go about her rebellion through writing and putting forth the female voice. Howeve r, she compromises her own self-adulation with the criticism that she receives. She recognizes and notes that â€Å"others† appose her voicing her opinion: â€Å"Considering not how others would them like† (4).By interjecting this criticism in with her self-congratulatory treatise, she refutes them with an impervious tone in her language, as if she intended to rebel and dismiss the reader’s response to her style. Conversely, she also acknowledges them, within the first four lines of her poem, which alludes to a deep concern. This indicates a woman who cares deeply for what others think. This concern could be due to her position in society. Being the wife of a Duke and lady to an ostracized Queen, it was well within her interest to be aware of her social milieu.Moreover, the aristocracy was used to people caring about what they thought and effecting how others acted and spoke, in other words, exercising their hegemony. While she is amongst this power structure, s he pushes the limits of her position and acceptance by speaking out and seeks to establish agency, which was not readily afforded to women in the seventeenth century. Even though Margaret Cavendish’s rank was high enough to enjoy an element of immunity, she expresses concern over the fragility of her position.In A True Relation of My Birth, Breeding and Life Margaret appreciates how the breadth of her status is afforded to her through marriage, â€Å"second wife to the Lord Marquis of Newcastle, for my lord having had two wives, I might easily have been mistaken, especially if I should die and my lord marry again† (1780). Her language seems humble yet uncertain. One could postulate that this uncertainty is due to her position being conditional of a male counterpart. In her texts, she relies heavily upon a male for information and education.In A True Relation†¦ she diminishes her own ability â€Å"†¦I had a natural stupidity† (1779) and relays how she w ould be dependent upon a male member of her household to explain matters to her: â€Å"†¦and when I read what I understood not, I would ask my brother†¦ he being learned, the sense of meaning thereof† (ibid). Quickly following this passage, she resorts to gender performance1: â€Å"†¦my serious study could not be much, by reason I took great delight in attiring, fine dressing and fashions† (ibid). By positing herself within a socially accepted arena for women, she at once becomes less offensive to her female readership.However, she shows her â€Å"anxiety of authorship† that Sarah Gilbert and Susan Gubar define as a female author’s anxiety of being judged by male readers, critics, etc. thus they will compromise their own voice in an attempt to placate the male gaze2 and save themselves from alienation: â€Å"Her battle, however, is not against her (male) precursor’s reading of the world but against his reading of her. In order to de fine herself as an author she must redefine the terms of her socialization†, (Gilbert and Gubar, 2027).Gilbert and Gubar’s solution for this is a female author must replace the male precursor or influence with a female, at once helping to her to identify with her own sex and giving her an alliance in which to fight patriarchal control. However, if women themselves perpetuate male hegemony, this can prove difficult. In The Blazing World, Margaret creates a new world in order to experience and exercise the ambition and power that she desired. This new world becomes a metaphor for the real world with which she parallels it.In this new world, it is finally safe for her to tackle male hegemony head on and she attempts to do this with analogies of what men become. In her customized world, men take on animal characteristics. Some examples are: â€Å"worm-men†¦fox-men†¦ant-men†¦ape-men† (1781) most of which could be considered derogatory and most certainly diminutive. By ‘othering’ the male species and making them sub-human, Margaret can successfully exercise control in her realm. She also employs this tactic in The Hunting of the Hare in which Wat, a male, is a hare being hunted and assuming a secondary and fragile position. However her anxiety of authorship recurs.After assigning animal counterparts for the male species in The Blazing World, she quickly reiterates that she indeed receives power from the emperor. The empress and the Duchess, both as Margaret in a fragmented state, are informed and educated by priests and statesmen, both male, of the affairs of the government and the church. These figures, which could arguably also represent elements of Margaret’s own psyche particularly the internalized male gaze, attempt to justify their exclusion of women from places of worship and matters of the state as they are â€Å"importunate persuasions† or threatening figures of change (1782).Furthermore, when sh e herself describes the power one could exercise in their own world, she does so by using masculine pronouns; â€Å"he may create a world of what fashion and government he will†¦as he pleases†¦as he thinks best†¦also he may alter that world†¦Ã¢â‚¬  (1784). Thus, Margaret once again crumbles under the pressure of anxiety of authorship due to fear of social backlash. In a previously quoted passage, Margaret Cavendish uses ‘reason’ as the justification for her self-diminution: â€Å"†¦my serious study could not be much, by reason I took great delight in attiring, fine dressing and fashions† (1779).OED defines reason as â€Å"A statement of some fact (real or alleged) employed as an argument to justify or condemn some act, prove or disprove some assertion, idea, or belief †. However, her usage of the word ‘reason’ evolves. ‘Reason’ later becomes the conduit for her creation of the world in which she can rule as a sovereign, as â€Å"Margaret the First†: â€Å"This is the reason, why I added this†¦to my philosophical observations† (1781). Her language when referring to herself in The Blazing World is authoritarian: â€Å"I shall account myself as a happy creatoress† (1780); â€Å"authoress of a whole world† (1784), etc.Although initially she claims to merely be a scribe to the empress of this imaginary world, there is evidence that Cavendish actually sees herself as the empress. In the first paragraph she sees two worlds, the world in which she exists as Margaret Cavendish and the Blazing World, as antipodes of each other, thus making them parallel: â€Å"†¦and joined them as two worlds at the ends of their poles† (1780). She goes on to describe the world of her creation: â€Å"it is a description of a new world†¦a world of my own creating, which I call the Blazing World† (ibid).As she posits herself in an ultimate position of pow er as â€Å"creatoress† and â€Å"authoress† (idem) she herself is sovereign, thus the empress could easily be interpreted as her. This is further evidenced within the romantic beginning of the story. The empress is heralded as a goddess and the object of the emperor’s affection, paralleling the empress’s story with that of her own: receipt of power and title through marriage. By aligning herself with a female figure of power, she at once establishes a female precursor but also empowers herself in fighting the alienation of hegemonic criticism.The metaphor of this alignment is interesting. One would figure that she had a very powerful female ‘precursor’ in Queen Henrietta Maria; however the Queen’s power was jeopardized by Charles’ execution and her banishment. Also, this particular alignment had proved precarious as it caused the loss of her and her husband’s estate (albeit temporarily). Thus, it was necessary for Margar et to create a new female figure of authority with which she could associate herself. Her internal conflict of desperately wanting to speak out but being checked by fear of societal repercussions is exercised in interesting ways.Margaret is aware of the perpetuation of the male hegemony through women and illustrates her frustrations through her texts. Women would most likely make up Margaret’s ideal readership; however one can surmise that they have also been the source of a great deal of her criticism. In The Poetess’s Hasty Resolution, Margaret signifies the ‘she’ as the source of her criticism: Will you, said she, thus waste your time in vain On that which in the world small praise shall gain? For shame, leave off, said she, the printer spare He’ll lose by your ill poetry, I fear.Besides the world hath already such a weight Of useless books, as it is overfraught. Then pity take, do the world a good turn, And all you write cast in the fire and bur n. (9-16) In this incendiary passage, Margaret exposes many elements of oppression perpetuated by women upon women. The idealized female precursor and intended source of support, demeans Cavendish’s art and directly exercises male hegemonic social pressures upon her. First, they attempt to induce fear in her of women’s destructive capability upon their husbands if they do not adhere to gendered social norms.Second, these critics try to instill her with an anxiety of influence4 by purporting that there are enough things to read in the world and she is not worthy of authorship. Finally, the female critic entreats her to immediately quit her education and stifle her ambitions, as knowledge and ambition are not becoming of a lady. However, Margaret reifies her own anxiety and realizes the danger of this â€Å"importunate persuasion† (ibid) and persists: â€Å"Then all in haste I to the press it sent/Fearing persuasion might my book prevent† (19-20). The betra yal by her own sex does little to incite camaraderie in Margaret.To empower herself against an oppressive but generative hegemonic control that is elicited upon her by other women, she attempts to distinguish herself through singularity. â€Å"I would dislike if any should follow my fashions, for I always took delight in a singularity† (1779). Thus, one may understand why Margaret would desire to set herself apart from her sex, as they did not prove to be an agreeable group with which to identify. However, to be alienated and excluded was a dire circumstance for a woman in seventeenth century Britain.She had already experienced banishment and understood how stringent the repercussions proved for stepping outside the societal ideal. Thus, if Margaret Cavendish could achieve renown and be considered excellent, she could achieve social currency. This accreditation would be what she would need to stave off the scorn of the more strict members of society. She achieved the respect of some accomplished members of society, mainly men, and cunningly realized that the best place with which to exercise her ‘reason’ was through her status in society.The complex and uncomfortable dynamic of desiring to remain in a position of influence and imperatively needing to utilize her own voice regardless of the heavy hegemonic resistance against it is a brave endeavor but a futile one. In order to remain amongst a group, one must adhere to its rules and be amenable to its policies. Although, Margaret Cavendish couldn’t fully achieve self sovereignty in her own time, she provided future female writers with a precursor to the feminist writers what would succeed her. Through her struggle she gives other ‘authoresses’ the support she herself would have appreciated.Although, she herself never achieved self sovereignty through singularity, she proved to be more than just a â€Å"great emulator† (1779). Works Cited Butler, Judith. â€Å"Gend er Trouble† Leitch 2488-2501. Gilber, Sandra M. and Susan Gubar â€Å"The Madwoman in the Attic: The Woman Writer and the Nineteenth-Century Literary Imagination. † Leitch 2023-2035. Greenblatt, Stephen, ed. The Norton Anthology of English Literature. Vol. 1. 8th ed. New York, NY: W. W. Norton and Company, 2006. 1773-1784. Leitch, Vincent, ed. The Norton Anthology of Theory and Criticism. New York, NY: W. W. Norton and Company, 2001. Footnotes 1.Gender performance is a theory derived by Judith Butler which states that the everyday actions of women and men are a performance of societal expectations and gendered norms and further more that this performance perpetuates the social constructs places upon us that define what it means to be male or female, man or woman, i. e. : men refusing to cry, women wearing high heels etc. 2. The male gaze is a poststructuralist feminist theory stemming from Michel Foucault’s utilization of the panopticon in surveillance society. T his theory places male hegemony in the position of the panopticon and women ‘self correct’ under its gaze.

The Definitive Solution for Ftce General Knowledge Essay Topics 2016 That You Can Learn About Today

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