Interferon inactivation

The supernatants containing the interferons were decanted from monolayers, pooled,and tested for freedom from bacteria. Residual arborvirus was inactivated by acid and heat as follows. The liquid was brought to pH 2 by the addition of 0.3N hydrochloric acid in Earle s saline (minus sodium chloride and sodium bicarbonate), kept at 4°C for 24 hours, and then brought back to pH 7 by the addition of 0.3N sodium hydroxide in distilled water. The liquid was then heated at 56°C for 30 minutes. 

Each interferon preparation was ultracentrifuged at 20,000 revolutions per minute for one hour to remove tissue debris and inactivated virus. The supernatant was dialyzed against distilled water (1 400) for 24 hours at4°C. The material was then freeze-dried. The dried product was reconstituted in one-tenth of the original volume in distilled water and dispensed into ampoules. Reconstituted solutions were assayed for interferon activity, examined for toxicity, and tested for sterility. 

McArthur JC (2004) HIV dementia an evolving disease. J Neuroimmunol 157(l-2) 3-10 McArthur JC, Hoover DR, BaceUar H, MUler EN, Cohen BA, Becker JT, Graham NM, McArthur JH, Seines OA, Jacobson LP et al (1993) Dementia in AIDS patients incidence and risk factors. Multicenter AIDS Cohort Study. Neurology 43(ll) 2245-2252 McManus CM, liu JS, Hahn MT, Hua LL, Brosnan CE, Berman JW, Lee SC (2000) Differential induction of chemokines in human microgUa by type I and II interferons. GUa 29(3) 273-280 McQuibban GA, Butler GS, Gong JH, BendaU L, Power C, Clark-Lewis I, OveraU CM (2001) Matrix metaUoproteinase activity inactivates the CXC chemokine stromal ceU-derived factor-1. J Biol Chem 276(47) 43503 3508... 

The shaking of protein solutions may lead to aggregation and precipitation as a result of several mechanisms, such as air oxidation, denaturation at the interface, adsorption to the vessel, or mechanical stress. These possibilities were systematically examined for solutions of human fibroblast interferon [50]. In this example, mechanical stress was identified as the causative factor in the inactivation. The proposed mechanism of inactivation by mechanical stress was through orientation of the asymmetrical protein in the... 

Forty-four years have passed since the initial discovery of interferon (IFN) [1]. Chick chorioallantoic membranes incubated with heat-inactivated influenza virus produced a substance that conferred resistance to infection by live virus [2,3].Termed interferon, this substance was later shown to have properties of a protein and to consist of a family of distinct proteins. 

Disposition According to Microme-dex, data on the metabohsm of interferon beta are lacking. However, this compound is similar to interferon alfa, which is rapidly inactivated in body fluids and tissue. 

These studies demonstrating a protective effect of sialic acid residues on serum glycoproteins provide an explanation for earlier, conflicting observations about the biological effect of, for example, desialylated erythropoietin, which stimulates erythropoiesis only after direct application to bone-marrow cell-cultures, and not after injection into the blood stream.469 In the latter experiment, only the native, sialylated hormone was active. Rapid clearance and inactivation of follicle-stimulating hormone,470 or interferon,471 after treatment with sialidase may be explained by uptake into liver cells. 

In discussing the mechanism of antiviral protection and stimulation of interferon production in the mouse, DeClercq and Merigan46 concluded that there was a direct relationship between the extent of protection against vesicular stomatitis virus, the titers of interferon produced and the doses of tilorone. Giron et al.47, however, found no correlation between interferon induction and protection against MM virus in mice. Protection was achieved at doses far below the doses at which detectable interferon was found in the serum. Both findings may be consistent with differing mechanisms of viral inactivation for the two viruses under study. 

High temperatures can break native S—S bonds and form new S—S bonds that can lock the protein into a denatured configuration. Low pH, sodium dodecyl sulfate. Tween 80 , chaotropic salts, and exogenous proteins have been used to protect proteins from thermal inactivation.f Ethylene glycol at 30-50% was used as a protectant of antiviral activity of p-interferon preparations. Human serum albumin was used inrecombinant human interferon-pser-i7, which resulted in increased thermal stability. Water-soluble polysaccharides such as dextrans and amylose and point-specific (site-directed) mutagenesis have also been used to increase thermal stability of therapeutic proteins and peptides. 

If an intermediate forms that has a solubility less than that of N or D this can lead to aggregation and eventually to precipitation. For example, the addition of moderate amounts of denaturant to bovine growth hormone (bGH) can generate a partially unfolded intermediate of low solubility which aggregates. Similarly, y-interferon (IFN-y) is inactivated by acid treatment or by the addition of salt because the dimeric native state is converted into monomers, which are partially denamred. For both proteins the formation of intermediates leads to inactivation. 

Several models, yet relatively little direct evidence has been presented to explain the genetic regulation of interferon production. No matter what model may be built to accomodate all known facts, it should be composed, in its simplest form, of an operon (interferon gene), a repressor protein, and a mRNA. The repressor protein may combine with the operator of the operon (referred to as transcriptional control in Fig. 7) in this state, the interferon gene would be repressed. It would be derepressed if the repressor is inactivated, e.g. following contact of the cell with the interferon inducer. The derepressed interferon gene would then be transcribed to mRNA, which in turn is translated to interferon. Hypothetically, the repressor protein may also combine with the mRNA for interferon (referred to as translational control), and even with the interferon molecule itself (referred to as posttranslational control in Fig. 7). 

As previously mentioned, interferons are cytokines produced by cells to protect them from viral infection, and anti-interferon strategies are a part of the immune evasion repertoire of most viruses. These mechanisms include the production of soluble versions of interferon receptors, which act as decoys. These decoys bind and inactivate interferons before they reach their destination - normal, membrane-bound receptors.22... 

Because tumor growth is associated with the progressive impairment of immunologic competence, a therapeutic goal is the enhancement of cell-mediated immunity. Cell-mediated immunity may be augmented by nonspecific stimulants such as levamisole, lymphokines, and interferon, which stimulate the antitumor activity of NK cells. Cell-mediated immunity may also be stimulated by specific stimulants such as vaccines, which are composed of killed or inactivated tumor cells or tumor cell fragments. The full range of immunotherapy s usefulness remains to be realized. 

Interferon-a. Alfa-interferon alpha-interferon IFN- LelF leukocyte interferon lymphoblastoid interferon. Family of highly homologous species -specific proteins that inhibits viral replication and cellular proliferation and modulates the immune response. One of the type I interferons. Produced by peripheral blood leukocytes or lymphoblastoid cells upon exposure to live or inactivated virus, double-stranded RNA. Or bacterial products. Multiple sub-types have been identified which contain 163-166 amino acids mol wt approximately 18.000-20,000 daltons. Production by virus stimulated human leukocytes I. Gresser, Proc. Soc. Exp. Biol Med. 108, 799 (1961). Purification C. 

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