So today, guess what we had at church? A polka party! Haha, just kidding, we had a brunch (bet you would have never guessed that one). I had every intention of not going...big surprise, lol. I like my routines. I eat breakfast before church, and I have the same thing every day (almost). So going to church and
1) being expected to eat
2) not knowing what there would be to eat and
3) feeling like certain people who know about my ED would be watching me to see how I am 'really' doing (if you are reading this Elise, I'm not talking about you)
Well, that kinda made me really nervous. But with a little encouragement from Neil, I decided to go. Plus, I thought it might be even more conspicuous if I didn't go.
Anyways, I did well. So that did feel nice.
Soooooo, now to food in general. Heh.
I've already got the "What the hell are you doing?!" from my therapist on Thursday. And I am (not) looking forward to getting chewed out by my nutritionist today for my food records.
Lack of structure = Lots of Megan sleeping = Lots of not eating.
So the new equation should be to add an alarm to the mix and hope for the best.
Perhaps I was sleeping a lot because I moved myself out all by myself. I guess this doesn't sound too unusual, but it really speaks volumes considering I wasn't even able to push my cart two years ago moving out.
Anyways, we've already established that I feel like crap when I don't eat. And it gets me nowhere. So I have done better over the weekend and such. And I'm supposed to start doing back-to-scale. Part of the doing not-so-well definitely had something to do with the 'high' of seeing the number drop.
Sunday, May 25, 2008
Monday, May 12, 2008
So, I don't totally suck at life
Yay. I was really upset for the last couple of weeks over a paper I had written. It was based on an article that might as well been written in Greek. This can be told be the title of the paper, 'Synergistic Roles of Antibody and Interferon in Noncytoclytic Clearance of Sindibis Virus from Different Regions of The Central Nervous System'. And interestingly enough, I have seen articles with much longer titles.
So, I did what I do best...procrastinate. Not totally, I spent hours trying to decipher what the article meant, but I did not start the paper until the night before it was due. Oops?
Basically we had to explain the article in a page and a half or less and then devise a new experiment based on what was missing from the article. And since I don't even have a BS, I was definitely not confident that whatever hypothesis I came up with would make any sense or not be totally stupid.
It was due at 5 pm. I turned it in a 4:58. And then I realized around 5:30 that I had not made the margins 1" all around, and in the rubric, that means it would not even be graded and would have to be resubmitted late. But on the bright side, I got to try and add some stuff to my paper and make it more complete. But I still thought I didn't do a good job.
Apparently not. I got a perfect score and the comment, "Very solid paper. Good job!" So that made me feel good. And when I reread it, it did seem like a professional paper (IMHO).
And since I am a dork, I am going to post it.
Burdeinick-Kerr et al. (2007) examined the roles of antibodies and interferons in clearing infection of Sindibis Virus (SINV) from different parts of the central nervous system (CNS). The purpose of this experiment was to look at the roles of specific parts of the adaptive and innate immune system. SINV is an alphavirus from the family Togaviridae that is transmitted by mosquitoes and is related to eastern (EEE) and western equine encephalitis (WEE) viruses. Alphaviruses are single stranded positive sense RNA viruses that are enveloped and present icosahedral symmetry. SINV presents symptoms such as a flu-like fever, polyarthritis and rashes in humans. However, in mice, SINV targets the neurons of the central nervous system (CNS), causes encephalomyelitis and is potentially fatal.
To facilitate this experiment, researchers examined mice that were wild type (WT), mice with severe combined immunodeficiency (SCID), beta interferon (IFN-β) knockout (BKO), IFN-γ knockout (GKO), IFN-γ receptor knockout (GRKO), antibody knockout (μMT), and antibody and IFN-α knockout (?MT/GKO) mice. IFN-β? are essential to the early innate immune response, and without it, mice deficient in these die before they can mount a specific immune response. IFN-β/γ and IFN-γ are all necessary for antibody mediated clearance of the virus.
SCID mice cleared infection of SINV in all parts of the CNS through passive transport of antibodies, but the production of virus increased as the levels of antibody decreased. Thus, they did not clear any virus from any part of the CNS, but presented no signs of neurological disease. WT mice, however, were able to clear infection within eight days.
The lack of IFN-β in BKO mice did not affect the production of IFN-α in the CNS and therefore did not affect SINV clearance or recovery.
Both GKO and GRKO mice cleared the virus from the brain, brain stem and lumbar spinal cord, but this clearance was not maintained. GKO mice had virus in the brain or brain stem and GRKO mice had virus in the lumbar spinal cord.
μMT mice were able to clear infections from the brain stem and spinal cord, but not the brain itself. These mice also experienced reactivation of the infection in all areas of the CNS. However, the levels of virus were lower than SCID and μMT/GKO mice.
μMT/GKO mice were unable to clear SINV from the brain, but the levels of virus were lower than with SCID mice. Initially, μMT/GKO mice were able to clear SINV in the brain stem, but levels of virus rose after 18 to 22 days. Clearance of SINV in the spinal cord was slow, and at day 12, 2/3 of the mice had not cleared the infection.
This experiment provides many practical implications for understanding the respective roles of antibodies and interferons in mounting a successful immune response. IFN-β was important in controlling the replication of SINV in the CNS, but had no role in the clearance of the virus. Antibody production was useful in clearing SINV from the brain, but did not prevent persistent infection. IFN-γ was important in clearing SINV from the brain stem and spinal cord and was implicated in prevention of persistent infection of the host. The role of IFN-γ or γ was most apparent with the μMT mice. Even though it seemed as if WT, GKO and GRKO mice had completely cleared SINV, at around day 12 there was virus replicating in all of the mice. Because there was some reactivation of the virus, it is likely that the viral RNA in the CNS is persistent, even with adequate immune response.
An important question that was not answered is what are the specific functions of IFN-β, as compared to IFN-α? And what is the combined effect of missing both IFN-β and IFN-α? Mice that are missing the receptors for IFN succumb to alphavirus infection before an acquired immune response can be mounted.
To complete this experiment, IFN-α knockout (AKO), IFN-β knockout (BKO), AKO/BKO, WT and SCID mice would be used, with the WT mice being the control. The AKO, BKO, WT, and SCID mice could be bought and the AKO/BKO mice could be bred. Tail snips would be digested from each of the mice and they would be genotyped by using PCRs. SINV would be injected intracerebrally (i.c.) into mice that were 4 weeks old and tissue cultures would be taken from the brain, brain stem and spinal cord. These tissue cultures would be serially diluted and go under plaque formation to find the amount of infectious virus present. An enzyme immunoassay (EIA) would be used to analyze the presence and amounts of immunoglobulin M (IgM), IgG, IgG1, IgG2a, and IgG2b in each region of the CNS (brain, brain stem, and spinal cord). The tissues from each of the EIA would be centrifuged as well and then would be assayed for IFN- α and IFN-β.
Another important question that was not answered was what additional factors relate to clearance; focusing on the differences between clearance of SINV by ?MT/GKO and SCID mice. SCID mice lack any kind of acquired immune response; they have neither B nor T cells. Essentially, they are completely defenseless against viral infection. ?MT/GKO mice also have a number of things that make them vulnerable to virus attack. Not being able to produce antibodies is the result of mature B cells not being produced. The mice are not only lacking mature B cells, but also are deficient in CD4+ and CD8+ T cells. This issue is made more obvious by the fact that these mice cannot secrete cytokines (IFN-γ). This suggests that there is some other role of B cells other than production of antibodies that helps with clearance of virus.
Present research suggests that there are some other antiviral factors that are important in clearance of the virus in addition to antibodies. A previous study by Lysenko (2005) observed that the action of complement and neutrophil-like cells were independent of antibody presence and adaptive immunity in the clearance of S. pneumoniae. Other cytokine factors may be useful in resisting viral infection. In vitro, Tumor necrosis factor alpha TNF-α, interleulkin-1 (IL-1) and interleukin-6 (IL-6) have contributed to helping to fight infection, but their role in vivo is not well known. The present study used TNF-α and IL-6 to test for cytokine production.
An experiment that could be used to test the hypothesis that the previous factors are useful in viral clearance would be to use μMT/GKO mice as a control and then look at the effects of removing each factor and the effect that has on viral clearance and their relation to SCID mice. The experiment would be very similar to the one described above; creating a knockout strain of mouse for TFN-α and IL-6. A complement protein that could be targeted for knockout would be the membrane cofactor protein (CD46).
There would be 5 groups overall, the control group (μMT/GKO), TFN-α knockout (TAKO), IL-6 knockout (ILKO), CD46 knockout (CKO) and SCID mice. Their genomes would be found through PCR. Mice that were 4 months old would be given SINV i.c. and tissue cultures would be taken from the brain stem, the spinal cord and the brain and these cultures would be serially diluted and plaqued to find the amount of virus present. Since these defenses against SINV are antibody-independent, EIA/ELISA would not be a good candidate to assay this experiment. A better technique would be to use ELISPOT to assay for the activity of TNF-α, IL-6 and CD46.
If the hypothesis is correct that these defenses are antibody- independent, the levels of virus will be increased from the control and will be closer to the levels of the SCID mice. This experiment would help to illustrate how truly complex our immune systems are and would demonstrate that having many different defenses against viral attack is essential because if one thing goes wrong, the host will have other backup defenses and can fight off the attack.
Works Cited
Elena S Lysenko, Adam J Ratner, Aaron L Nelson, and Jeffrey N Weiser. The Role of Innate Immune Responses in the Outcome of Interspecies Competition for Colonization of Mucosal Surfaces. PLoS Pathog. 2005 September; 1(1): e1.
So, I did what I do best...procrastinate. Not totally, I spent hours trying to decipher what the article meant, but I did not start the paper until the night before it was due. Oops?
Basically we had to explain the article in a page and a half or less and then devise a new experiment based on what was missing from the article. And since I don't even have a BS, I was definitely not confident that whatever hypothesis I came up with would make any sense or not be totally stupid.
It was due at 5 pm. I turned it in a 4:58. And then I realized around 5:30 that I had not made the margins 1" all around, and in the rubric, that means it would not even be graded and would have to be resubmitted late. But on the bright side, I got to try and add some stuff to my paper and make it more complete. But I still thought I didn't do a good job.
Apparently not. I got a perfect score and the comment, "Very solid paper. Good job!" So that made me feel good. And when I reread it, it did seem like a professional paper (IMHO).
And since I am a dork, I am going to post it.
Burdeinick-Kerr et al. (2007) examined the roles of antibodies and interferons in clearing infection of Sindibis Virus (SINV) from different parts of the central nervous system (CNS). The purpose of this experiment was to look at the roles of specific parts of the adaptive and innate immune system. SINV is an alphavirus from the family Togaviridae that is transmitted by mosquitoes and is related to eastern (EEE) and western equine encephalitis (WEE) viruses. Alphaviruses are single stranded positive sense RNA viruses that are enveloped and present icosahedral symmetry. SINV presents symptoms such as a flu-like fever, polyarthritis and rashes in humans. However, in mice, SINV targets the neurons of the central nervous system (CNS), causes encephalomyelitis and is potentially fatal.
To facilitate this experiment, researchers examined mice that were wild type (WT), mice with severe combined immunodeficiency (SCID), beta interferon (IFN-β) knockout (BKO), IFN-γ knockout (GKO), IFN-γ receptor knockout (GRKO), antibody knockout (μMT), and antibody and IFN-α knockout (?MT/GKO) mice. IFN-β? are essential to the early innate immune response, and without it, mice deficient in these die before they can mount a specific immune response. IFN-β/γ and IFN-γ are all necessary for antibody mediated clearance of the virus.
SCID mice cleared infection of SINV in all parts of the CNS through passive transport of antibodies, but the production of virus increased as the levels of antibody decreased. Thus, they did not clear any virus from any part of the CNS, but presented no signs of neurological disease. WT mice, however, were able to clear infection within eight days.
The lack of IFN-β in BKO mice did not affect the production of IFN-α in the CNS and therefore did not affect SINV clearance or recovery.
Both GKO and GRKO mice cleared the virus from the brain, brain stem and lumbar spinal cord, but this clearance was not maintained. GKO mice had virus in the brain or brain stem and GRKO mice had virus in the lumbar spinal cord.
μMT mice were able to clear infections from the brain stem and spinal cord, but not the brain itself. These mice also experienced reactivation of the infection in all areas of the CNS. However, the levels of virus were lower than SCID and μMT/GKO mice.
μMT/GKO mice were unable to clear SINV from the brain, but the levels of virus were lower than with SCID mice. Initially, μMT/GKO mice were able to clear SINV in the brain stem, but levels of virus rose after 18 to 22 days. Clearance of SINV in the spinal cord was slow, and at day 12, 2/3 of the mice had not cleared the infection.
This experiment provides many practical implications for understanding the respective roles of antibodies and interferons in mounting a successful immune response. IFN-β was important in controlling the replication of SINV in the CNS, but had no role in the clearance of the virus. Antibody production was useful in clearing SINV from the brain, but did not prevent persistent infection. IFN-γ was important in clearing SINV from the brain stem and spinal cord and was implicated in prevention of persistent infection of the host. The role of IFN-γ or γ was most apparent with the μMT mice. Even though it seemed as if WT, GKO and GRKO mice had completely cleared SINV, at around day 12 there was virus replicating in all of the mice. Because there was some reactivation of the virus, it is likely that the viral RNA in the CNS is persistent, even with adequate immune response.
An important question that was not answered is what are the specific functions of IFN-β, as compared to IFN-α? And what is the combined effect of missing both IFN-β and IFN-α? Mice that are missing the receptors for IFN succumb to alphavirus infection before an acquired immune response can be mounted.
To complete this experiment, IFN-α knockout (AKO), IFN-β knockout (BKO), AKO/BKO, WT and SCID mice would be used, with the WT mice being the control. The AKO, BKO, WT, and SCID mice could be bought and the AKO/BKO mice could be bred. Tail snips would be digested from each of the mice and they would be genotyped by using PCRs. SINV would be injected intracerebrally (i.c.) into mice that were 4 weeks old and tissue cultures would be taken from the brain, brain stem and spinal cord. These tissue cultures would be serially diluted and go under plaque formation to find the amount of infectious virus present. An enzyme immunoassay (EIA) would be used to analyze the presence and amounts of immunoglobulin M (IgM), IgG, IgG1, IgG2a, and IgG2b in each region of the CNS (brain, brain stem, and spinal cord). The tissues from each of the EIA would be centrifuged as well and then would be assayed for IFN- α and IFN-β.
Another important question that was not answered was what additional factors relate to clearance; focusing on the differences between clearance of SINV by ?MT/GKO and SCID mice. SCID mice lack any kind of acquired immune response; they have neither B nor T cells. Essentially, they are completely defenseless against viral infection. ?MT/GKO mice also have a number of things that make them vulnerable to virus attack. Not being able to produce antibodies is the result of mature B cells not being produced. The mice are not only lacking mature B cells, but also are deficient in CD4+ and CD8+ T cells. This issue is made more obvious by the fact that these mice cannot secrete cytokines (IFN-γ). This suggests that there is some other role of B cells other than production of antibodies that helps with clearance of virus.
Present research suggests that there are some other antiviral factors that are important in clearance of the virus in addition to antibodies. A previous study by Lysenko (2005) observed that the action of complement and neutrophil-like cells were independent of antibody presence and adaptive immunity in the clearance of S. pneumoniae. Other cytokine factors may be useful in resisting viral infection. In vitro, Tumor necrosis factor alpha TNF-α, interleulkin-1 (IL-1) and interleukin-6 (IL-6) have contributed to helping to fight infection, but their role in vivo is not well known. The present study used TNF-α and IL-6 to test for cytokine production.
An experiment that could be used to test the hypothesis that the previous factors are useful in viral clearance would be to use μMT/GKO mice as a control and then look at the effects of removing each factor and the effect that has on viral clearance and their relation to SCID mice. The experiment would be very similar to the one described above; creating a knockout strain of mouse for TFN-α and IL-6. A complement protein that could be targeted for knockout would be the membrane cofactor protein (CD46).
There would be 5 groups overall, the control group (μMT/GKO), TFN-α knockout (TAKO), IL-6 knockout (ILKO), CD46 knockout (CKO) and SCID mice. Their genomes would be found through PCR. Mice that were 4 months old would be given SINV i.c. and tissue cultures would be taken from the brain stem, the spinal cord and the brain and these cultures would be serially diluted and plaqued to find the amount of virus present. Since these defenses against SINV are antibody-independent, EIA/ELISA would not be a good candidate to assay this experiment. A better technique would be to use ELISPOT to assay for the activity of TNF-α, IL-6 and CD46.
If the hypothesis is correct that these defenses are antibody- independent, the levels of virus will be increased from the control and will be closer to the levels of the SCID mice. This experiment would help to illustrate how truly complex our immune systems are and would demonstrate that having many different defenses against viral attack is essential because if one thing goes wrong, the host will have other backup defenses and can fight off the attack.
Works Cited
Elena S Lysenko, Adam J Ratner, Aaron L Nelson, and Jeffrey N Weiser. The Role of Innate Immune Responses in the Outcome of Interspecies Competition for Colonization of Mucosal Surfaces. PLoS Pathog. 2005 September; 1(1): e1.
Monday, May 05, 2008
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