Author Topic: Analysis of single nucleotide polymorphisms in XMRV patient-derived integration  (Read 2416 times)

Val

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Analysis of single nucleotide polymorphisms in XMRV patient-derived integration sites reveals contamination from cell lines acutely infected by XMRV
Alice Rusmevichientong1, Jaydip Das Gupta2, Petra S. Elias1, Robert H. Silverman2, and Samson A. Chow1,*

Abstract

We analyzed xenotropic murine leukemia virus-related virus (XMRV) integration site sequences previously identified from human prostate tissues for single nucleotide polymorphisms (SNPs) to discriminate between patient and potential cell line sourcesof the proviruses. The SNPs of two integration sites were identical to those in cell lines but not the patients, whereas the data on the remaining 12 integration sites were inconclusive. Our results provide direct evidence for contamination during analysis of XMRV integration sites.

Robyn

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Dr. Yes

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Analysis of single nucleotide polymorphisms in XMRV patient-derived integration sites reveals contamination from cell lines acutely infected by XMRV
Alice Rusmevichientong1, Jaydip Das Gupta2, Petra S. Elias1, Robert H. Silverman2, and Samson A. Chow1,*

Abstract

We analyzed xenotropic murine leukemia virus-related virus (XMRV) integration site sequences previously identified from human prostate tissues for single nucleotide polymorphisms (SNPs) to discriminate between patient and potential cell line sourcesof the proviruses. The SNPs of two integration sites were identical to those in cell lines but not the patients, whereas the data on the remaining 12 integration sites were inconclusive. Our results provide direct evidence for contamination during analysis of XMRV integration sites.

I assume these are the same two integration sites identified by Towers' group?
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Gerwyn

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I assume these are the same two integration sites identified by Towers' group?


Yes they are throwing Judy and the WPI under a bus the day before the conference


Tango

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http://microb230.med.upenn.edu/PDF%20Library/18369476.pdf

Quote
HTLV-1 Integration into Transcriptionally Active Genomic Regions Is Associated with Proviral Expression and with HAM/TSP


Kiran N. Meekings1, Jeremy Leipzig2, Frederic D. Bushman2, Graham P. Taylor3, Charles R. M. Bangham1*
1 Department of Immunology, Wright-Fleming Institute, Imperial College London, London, United Kingdom, 2 Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America, 3 Department of Genitourinary Medicine and Communicable Diseases, Wright-Fleming Institute, Imperial College London, London, United Kingdom
Abstract
Human T-lymphotropic virus type 1 (HTLV-1) causes leukaemia or chronic inflammatory disease in ,5% of infected hosts. The level of proviral expression of HTLV-1 differs significantly among infected people, even at the same proviral load (proportion of infected mononuclear cells in the circulation). A high level of expression of the HTLV-1 provirus is associated with a high proviral load and a high risk of the inflammatory disease of the central nervous system known as HTLV-1- associated myelopathy/tropical spastic paraparesis (HAM/TSP). But the factors that control the rate of HTLV-1 proviral expression remain unknown. Here we show that proviral integration sites of HTLV-1 in vivo are not randomly distributed within the human genome but are associated with transcriptionally active regions. Comparison of proviral integration sites between individuals with high and low levels of proviral expression, and between provirus-expressing and provirus non- expressing cells from within an individual, demonstrated that frequent integration into transcription units was associated with an increased rate of proviral expression. An increased frequency of integration sites in transcription units in individuals with high proviral expression was also associated with the inflammatory disease HAM/TSP. By comparing the distribution of integration sites in human lymphocytes infected in short-term cell culture with those from persistent infection in vivo, we infer the action of two selective forces that shape the distribution of integration sites in vivo: positive selection for cells containing proviral integration sites in transcriptionally active regions of the genome, and negative selection against cells with proviral integration sites within transcription units.

Quote
Supporting Information
Figure S1 HTLV-1 integration in vivo and in vitro is identical at
the nucleotide level. Integration of HTLV-1 in vivo is indistinguishable
from that in vitro at the nucleotide level (in vitro data combined from the
co-culture sites obtained in this report and sites reported byDerse et al

MLVs and HTLV do the same - integrate into the same site!!!!

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Dr. Yes

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Even IF one were to assume that there was contamination in Silverman's lab that explains those two integration sites identified in the earlier study on prostate cancer, there were 10 other sites as yet unexplained by contamination, and above all there is the IHC and FISH evidence for MRV infection. 

None of this makes any comment on the Lombardi study, of course, particularly not on the WPI and NCI results.
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Adam

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Even IF one were to assume that there was contamination in Silverman's lab that explains those two integration sites identified in the earlier study on prostate cancer, there were 10 other sites as yet unexplained by contamination, and above all there is the IHC and FISH evidence for MRV infection. 

None of this makes any comment on the Lombardi study, of course, particularly not on the WPI and NCI results.

I agree Doc.

I took this to mean that there is evidence for contamination that has to be taken into account when conducting studies - evidence that if not taken into account might cast doubt on the validity of those studies.

I did not take it to mean that Silverman has suddenly decided to scupper the WPI and/or XMRV research.

Why would he?

jemal

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I don't think this was posted yet, this is the link to the article:
http://jvi.asm.org/cgi/content/abstract/JVI.05624-11v1

Full article is behind a pay wall.

I hope that Adam is right by the way.

Gerwyn

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I agree Doc.

I took this to mean that there is evidence for contamination that has to be taken into account when conducting studies - evidence that if not taken into account might cast doubt on the validity of those studies.

I did not take it to mean that Silverman has suddenly decided to scupper the WPI and/or XMRV research.

Why would he?
[/quote

id Silverman had not wanted to scupper the research he would have said that there was no evidence for contamination in the other ten integration sites and contamination is not a possible explanation which explains the findings because integration hot spots occur in human DNA which do not occur in the DNA of cell lines. The fact that XMRV integrates into different chromosomes in human beings compared to that in cell lines

That would have changed the meaning of the entire paper!

Adam

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There are loads of reasons he could do that, but this is not evidence as it clearly happens a lot.

HTLV integrates into precisely the same nucleotides in infected humans and in vitro cell lines.


This is the situation Towers and Silverman say that they are totally unaware of  - yea sure!

Would everyone like to email this info to them and to everyone else.  Next they will claim humans cannot be shown to exist.

"to the best of our knowledge" hides a multitude of sins it seems.

I cannot agree that Silverman is saying I was not aware. He is just doing the science for himself to confirm or cast doubt on Towers findings. He would look a right twat if he ignored/discounted Tower's findings and then later discovered there were correct.

With regards HTLV - as we have said many times before, it would be wrong to accept other virus model as being gospel for HGRV's - the likelihood that any two viruses, however closely related, act in precisely the same way, even to my non-scientific unscientifically educated mind, seems remote. For example, identical twins are not the same people despite their shared DNA.

Gerwyn

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I cannot agree that Silverman is saying I was not aware. He is just doing the science for himself to confirm or cast doubt on Towers findings. He would look a right twat if he ignored/discounted Tower's findings and then later discovered there were correct.

With regards HTLV - as we have said many times before, it would be wrong to accept other virus model as being gospel for HGRV's - the likelihood that any two viruses, however closely related, act in precisely the same way, even to my non-scientific unscientifically educated mind, seems remote. For example, identical twins are not the same people despite their shared DNA.

tHE dna OF SHARED TWINS IS IDENTICAL ADAM


if Silverman had not wanted to scupper the research he would have said that there was no evidence for contamination in the other ten integration sites and contamination is not a possible explanation which explains the findings because integration hot spots occur in human DNA which do not occur in the DNA of cell lines. The fact that XMRV integrates into different chromosomes in human beings compared to that in cell lines
 
 That would have changed the meaning of the entire paper!

HTLV integrates into exactly the same nucleotide in infected persons as it does in the DNA of cell lines might have been worth a mention too!


 

Adam

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The analogy of twins was to show that even though two things are identical (DNA) - enviroment - for instance - shapes them differently and they behave differently.

I bow to everyone's superior knowledge here - but out of interest again ask - why would Silverman scupper the research?

Gerwyn

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The analogy of twins was to show that even though two things are identical (DNA) - enviroment - for instance - shapes them differently and they behave differently.

I bow to everyone's superior knowledge here - but out of interest again ask - why would Silverman scupper the research?

Yes Adam why indeed?

Patricia

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http://niceguidelines.blogspot.com/2011/09/dr-robert-h-silverman-xmrv-is.html

 Dr Robert H. Silverman: XMRV is contamination   
  Analysis of single nucleotide polymorphisms in XMRV patient-derived integration sites reveals contamination from cell lines acutely infected by XMRV
 
   Alice Rusmevichientong1, Jaydip Das Gupta2, Petra S. Elias1, Robert H. Silverman2, and Samson A. Chow1,*:
 
  1 Department of Molecular and Medical Pharmacology, and UCLA AIDS Institute, UCLA School of Medicine, Los Angeles, California 90095 2 Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195 *
 
 Corresponding author. Mailing address: Department of Molecular and Medical Pharmacology, UCLA School of Medicine, CHS 23-133, 605 Charles E. Young Dr. South, Los Angeles, CA 90095, USA. Tel: (310) 825-9600. Fax: (310) 825-6267.
 E-mail: schow@mednet.ucla.edu 
 
 ABSTRACT We analyzed xenotropic murine leukemia virus-related virus (XMRV) integration site sequences previously identified from human prostate tissues for single nucleotide polymorphisms (SNPs) to discriminate between patient and potential cell line sources of the proviruses. The SNPs of two integration sites were identical to those in cell lines but not the patients, whereas the data on the remaining 12 integration sites were inconclusive. Our results provide direct evidence for contamination during analysis of XMRV integration sites.

See also: XMRV / HTLV integrate into precisely the same nucleotides in infected humans and in vitro cell lines



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bakercape

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Yes Adam why indeed?

I wonder if Silverman has been having his grant apps denied recently. Just as the WPI has after the Science paper. 

I wonder if his grant app requests are a matter of public record the last two years? 
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