Virology Journal - Latest Comments

The modified description for FMD serotype Asia1 epidemic situation

Xin Yang — 2012-08-31T14:03:41Z

There was an inappropriate description for the epidemic situation of Asia1 FMDV in China (the first sentence of the second paragraph in Background of the paper "FMD serotype Asia1 has been epidemic in China for more than 50 years" ) . The truth is that the first serotype Aisa1 FMDV of China was isolated in 1958. After that, there was no reports about the Asia 1 FMDV untill the year of 2005. So we modified the sentence to "In China, the first FMDV serotype Asia1 was isolated in 1958 ".

Reply to Dr. Dusty Miller's comment.

Chandravanu Dash — 2012-06-27T13:07:14Z

We thank Dr. Miller for his insightful comments. Our rebuttals to his comments are described below.

As described in the abstract of our manuscript, our report builds on and challenges the published data that prostate epithelial cells lines do not express A3G protein (Refs 13-16 in the manuscript). Since we detected A3G in prostate epithelial cell lines (LNCaP and DU145) by western blot and mass spectrometry, the major goal of our findings was to point out discrepancy in A3G detection in the prior studies (Refs 13-16 in the manuscript).

Dr. Miller contests that our conclusions ¿XMRV replicates efficiently in prostate epithelial cells by downregulating A3G expression¿ is not supported by data. We acknowledge that in our report we did not measure XMRV replication. This is because it had already been suggested that "XMRV replicates efficiently in prostate epithelial cells (LNCaP and DU145 cells)¿ by Paprotka et al (2011), Dong et al (2006) and Steiler et al (2010). Given that we were claiming A3G is present in prostate cancer cells, we relied on these reports to conclude that XMRV replicates efficiently in prostate epithelial cells.

How about our claim that ¿XMRV uses a novel mechanism to counteract the antiviral effects of A3G¿? Our data provided strong evidence that XMRV infection downregulated A3G in LNCaP and DU145 cells (As pointed out by Dr. Miller himself). Therefore, we suggested that XMRV may have evolved a novel mechanism to counteract A3G in prostate epithelial cells. This is mainly based on the fact that XMRV does not encode accessory genes that are known to degrade A3G in complex retroviruses. Dr. Miller suggests that the 50% downregulation of A3G is due to packaging to A3G in XMRV virions. Although plausible, we believe that packaging of A3G into the virions may not reduce total A3G levels to 50% since A3G is a constitutively expressed protein. It is also important to point out that as per Paprotka et al. (2011), out of 22 clones from infected LNCaP cells a single clone exhibited G-to-A mutation and these mutations were highly specific for A3F rather than A3G. So the question is if A3G is being packaged into the virions why we do not see A3G specific mutations in XMRV genome. These are some of the questions that are being investigated in our lab currently.

Conclusions not supported by the data

A Dusty Miller — 2012-06-13T10:12:28Z

This report claims two major conclusions (first and last paragraphs of the Abstract):

1. "Although XMRV is thought to use XPR1 for cell entry, it infects A549 cells that do not express XPR1, suggesting usage of other receptors or co-receptors."

2. "XMRV replication was observed in GHOST cells that express CD4 and each of the chemokine receptors ranging from CCR1- CCR8 and BOB suggesting that infectivity in hematopoietic cells could be mediated by use of these receptors."

Neither conclusion is supported by the data.

First, some background. The Xenotropic and Polytropic retrovirus Receptor (XPR1) was discovered independently by three research groups in 1999 [1-3]. These groups screened cDNA expression libraries from different sources for production of proteins that could mediate xenotropic [1,2] or polytropic [3] retrovirus entry into cells that were normally nonpermissive for virus infection. The fact that they all came up with the same receptor provides evidence that there is only a single receptor for these viruses. Additional support for a single virus receptor comes from a subsequent study using human/hamster radiation hybrid cell lines to map genes in the human genome that could act as receptors for xenotropic retroviruses, which again found only one gene, XPR1 [4]. Furthermore, XPR1 mRNA was found to be expressed in all human tissues tested [1,2], thus, there is no need to postulate additional receptors to explain the ability of xenotropic and polytropic retroviruses, including XMRV, to infect many cell types.

So, what about the current authors' first claim that XMRV infects A549 cells, but they don't express XPR1, suggesting XMRV can enter cells by using other receptors? They do present data that XMRV can infect A549 cells, but they do not provide any data showing that the cells do not express XPR1. Nor do they provide a literature reference for their statement, repeated multiple times, that A549 cells lack XPR1. A quick Google search reveals a gene expression atlas report (http://www.ebi.ac.uk/gxa/gene/ENSG00000143324?ef=cell_line#efv=A549) that shows XPR1 is indeed expressed in A549 cells. Both gene expression studies listed found XPR1 RNA expression in A549 cells, with one showing higher than average levels and the other showing lower than average levels. Thus, in the absence of data to the contrary, A549 cells do indeed express XPR1, and no additional receptors need be postulated.

What about the authors' second claim that XMRV might use some of the same receptors as does HIV? While they show data that XMRV can replicate in GHOST cells that express CD4 and various chemokine receptors, they've left out the crucial control - cells not expressing these receptors! GHOST cells are derived from HOS human osteosarcoma cells, and a report from 1990 clearly shows that HOS cells are highly infectable by the prototypical NZB strain of xenotropic virus [5]. So, it is likely that the extra receptors in the GHOST cells have nothing to do with xenotropic retrovirus infection. In summary, none of the data presented support the authors' overall conclusion that receptors in addition to XPR1 can mediate XMRV entry.

So what's going on here? I've reread this paper several times to see if I've missed something. Instead, I found additional mistakes such as mislabeling of the panels in the two data figures. Perhaps a few critical paragraphs were inadvertently left out? What about Dr. Xue Wang, Dr. Mingjie Zhang, Dr. Robin Biswas, and Dr. Chintamani D. Atreya, who are acknowledged for critical review of this article? And how did peer review miss such seemingly obvious problems with this report? Perhaps the authors will explain to me what I've missed.

References
1. Tailor CS, Nouri A, Lee CG, Kozak C, Kabat D (1999) Cloning and characterization of a cell surface receptor for xenotropic and polytropic murine leukemia viruses. Proc Natl Acad Sci USA 96: 927-932.
2. Battini JL, Rasko JE, Miller AD (1999) A human cell-surface receptor for xenotropic and polytropic murine leukemia viruses: possible role in G protein-coupled signal transduction. Proc Natl Acad Sci USA 96: 1385-1390.
3. Yang YL, Guo L, Xu S, Holland CA, Kitamura T, et al. (1999) Receptors for polytropic and xenotropic mouse leukaemia viruses encoded by a single gene at Rmc1. Nat Genet 21: 216-219.
4. Rasko JE, Battini JL, Kruglyak L, Cox DR, Miller AD (2000) Precise gene localization by phenotypic assay of radiation hybrid cells. Proc Natl Acad Sci USA 97: 7388-7392.
5. Sommerfelt MA, Weiss RA (1990) Receptor interference groups of 20 retroviruses plating on human cells. Virology 176: 58-69.

Insufficient evidence

Laura Kasman — 2012-03-30T09:59:40Z

In my opinion, this report lacks some critical pieces of information:
1) Histological or molecular evidence that the lesions pictured are in fact warts and contain human papillomavirus
2) Independent chemical analysis verifying that the natural oil product used to treat the lesions in fact contained vitamin A, how much vitamin A it contained, and what other substances it also contained
3) Controls for other compounds the lesions came in contact with during the course of the 7 month treatment. New hand soaps or lotions for example, that were not combined with previous treatments. This would best be controlled for with additional cases.
4) Additional cases. Spontaneous remission of HPV lesions, if that is what these were, is frequent and well documented.

That prior OTC wart treatments were unsuccessful, and that untreated lesions disappeared along with the treated ones would seem to favor the conclusion that these lesions were not warts and they spontaneously regressed.

Is ECV304 an endothelial cell line?

Masanori Terajima — 2012-03-30T09:52:06Z

I am surprised to see that ECV304 cell line is still being used as an endothelial cell line after two papers in 1999 and 2000 clearly showed that it is not.

The two papers are

1: Dirks WG, MacLeod RA, Drexler HG. ECV304 (endothelial) is really T24 (bladder carcinoma): cell line cross- contamination at source. In Vitro Cell Dev Biol Anim. 1999 Nov-Dec;35(10):558-9. PubMed PMID: 10614862.

2: Brown J, Reading SJ, Jones S, Fitchett CJ, Howl J, Martin A, Longland CL, Michelangeli F, Dubrova YE, Brown CA. Critical evaluation of ECV304 as a human endothelial cell model defined by genetic analysis and functional responses: a comparison with the human bladder cancer derived epithelial cell line T24/83. Lab Invest. 2000 Jan;80(1):37-45. PubMed PMID: 10653001.

Conclusions not supported by the data

A Dusty Miller — 2012-01-13T10:25:48Z

This report claims two major conclusions (last paragraph of Abstract):

1. XMRV replicates efficiently in prostate epithelial cells by downregulating A3G expression.

2. Our data suggest a novel mechanism by which retroviruses can counteract the antiviral effects of A3G proteins.

Neither conclusion is supported by the data.

First, some background. Many publications to date have shown that XMRV can be mutated by APOBEC3 proteins (A3A - A3G) present at various levels in human cells. Sequencing of the RNA genomes of XMRV viruses produced from particular human cells shows that some genomes are intensely mutated (hypermutated) while others show only background mutation rates likely due to errors in reverse transcription. Rates of hypermutation vary for viruses produced from different human cells, from almost none for XMRV produced by 22Rv1 prostate cancer cells (Paprotka et al., 2010), to ~25% for DU145 prostate cancer cells (Paprotka et al., 2010), to almost 100% for virus produced by human peripheral blood mononuclear cells (PBMC) (Chaipan et al., 2011). The results obtained for human PBMC clearly show that XMRV is not able to circumvent the effects of A3 restriction in human cells.

So, what about the current authors' conclusion that XMRV replicates efficiently in prostate epithelial cells (LNCaP and DU145 cells) by downregulating A3G? The first problem with this claim is the lack of a definition or assay for 'efficient replication'. No measurements of XMRV replication (or hypermutation) were performed by the authors. Previous reports cited by the authors in support of 'efficient' replication (manuscript refs. 3 and 18) documented XMRV replication by showing that XMRV proteins and reverse transcriptase increased with time after exposure of prostate cancer cell lines to XMRV, but provided no evidence that XMRV replication was more or less efficient than that of any other viruses. Next, we already know that XMRV virus produced by DU145 cells is ~25% hypermutated (Paprotka et al., 2010), so XMRV is not completely resistant to at least one of the A3 proteins present in DU145 cells. Even so, XMRV has been shown to 'efficiently' spread in DU145 cells (ref. 3). This is not unexpected, because ~75% of the virus produced from DU145 cells is infectious. Thus, in cells like DU145 that make low levels of A3 proteins, it is unnecessary for XMRV to completely downregulate A3 proteins to replicate 'efficiently'.

The authors do provide some evidence that A3G protein levels are reduced by ~50% in XMRV-infected LNCaP and DU145 cells compared to the parental cell lines. But, could a two-fold reduction in A3G levels mediated by XMRV be responsible for 'efficient' XMRV replication in these cells? In the case of XMRV-infected DU145 cells, where we know that ~25% of the virus produced is hypermutated and ~75% is active, would it matter if there was 2-fold more A3G, resulting in ~50% hypermutation and ~50% active virus production? We don't know because the authors have not measured XMRV replication rates in cells with different A3G levels, but one would expect the virus to replicate well under either condition.

Lastly, there is a simple explanation for the decrease in A3G levels the authors report; that A3G packaging into virions, which is required for virus hypermutation, is responsible for the decrease in cellular levels of A3G. This possibility would be easy to address by assay for A3G associated with XMRV virions in the cell culture medium. If so, there is no reason to propose an A3G regulatory mechanism involving XMRV.

What about the authors' second major conclusion, that their findings suggest a novel mechanism by which retroviruses can counteract the antiviral effects of A3G proteins? The mechanism, if any, is certainly not robust, and its existence is not supported by the data provided. At the very least, the authors should measure A3G mRNA levels in infected and uninfected cells to see if XMRV might be regulating A3G transcription, which would suggest the production of some transcription factor by XMRV, and might provide some support for the suggested regulatory mechanism. This experiment is critical for the authors' claim that XMRV downregulates A3G expression.

On a final note, the representative A3G protein data presented for LNCaP cells +/- XMRV in Fig. 3B don't support the average results from three experiments shown in Fig. 3C. In Fig. 3B, the ratio of A3G in LNCaP+XMRV cells to that in LNCaP cells appears to be about 1:5, the ratio of b-actin about 2:1, so the overall ratio of A3G, normalized to b-actin, is about 1:10 or 10%. The average ratio shown in Fig. 3C is 40% with a very tight error bar, inconsistent with the results shown in Fig. 3B being included in this average. In contrast, the results in Fig. 3B and 3D for the DU145 cells appear consistent.

Publications cited:

Chaipan, C., Dilley, K.A., Paprotka, T., Delviks-Frankenberry, K.A., Venkatachari, N.J., Hu, W.-S., and Pathak, V.K. (2011). Severe restriction of xenotropic murine leukemia virus-related virus replication and spread in cultured human peripheral blood mononuclear cells. J Virol 85, 4888-4897.

Paprotka, T., Venkatachari, N.J., Chaipan, C., Burdick, R., Delviks-Frankenberry, K.A., Hu, W.S., and Pathak, V.K. (2010). Inhibition of xenotropic murine leukemia virus-related virus by APOBEC3 proteins and antiviral drugs. J Virol 84, 5719-5729.

Cure?

Karla Bravo — 2012-01-04T10:30:06Z

Would this vaccine work as a cure for people already infected or only as a prevention vaccine?

Standing on the shoulders of giants...

Luis Branco — 2011-10-27T16:35:30Z

After this manuscript was published in preliminary form we recalled a report by Niklasson, Jahrling and Peters published in 1984 by the Journal of Clinical Microbiology entitled "Detection of Lassa virus antigens and Lassa virus-specific immunoglubulins IgG and M by enzyme-linked immunosorbent assay" that reported persistence of LASV-specific IgM in a relevant nonhuman primate model of LF. Niklasson, Jahrling and Peters observed that rhesus macaques surviving LF continued to generate significant serum titers of LASV-specific IgM until at least day 532 post-infection. The authors considered that, "this unexpected persistence of virus-specific IgM in the nonhuman primate model deserves further investigation and should also be evaluated in patients. The finding raises concerns about possible virus persistence and also suggests that the mere finding of virus-specific IgM in a single serum sample tested by ELISA may not imply recent infection." The current study demonstrates that LASV-specific IgM persists in human LF patients and provides evidence that LASV-specific IgM is not a reliable serological marker for acute LASV Infection. Moreover, in Niklasson et al isolation of nonhuman primates in controlled laboratory environments suggests that re-exposure to LASV is not responsible for the prolonged virus-specific IgM titers in the serum of convalescent animals. The findings in NHP suggest that prolonged IgM responses observed in humans is a common immunological response to LASV infection and opens discussions to its implications in the pathogenesis of LF.

Chromosomally integrated HHV-6 in renal transplant patients

Paolo Lusso — 2011-10-27T16:34:03Z

Sir:

The recent report by Csoma et al.¹ identifies HHV-6A as the viral variant most frequently detected in patients who received renal transplantation, but we noticed some important drawbacks in the study that suggest caution in the interpretation of these results. First and foremost, although the authors introduce the issue of chromosomally integrated HHV-6 (ciHHV-6) in the discussion, they did not test their patients for ciHHV-6. Individuals with ciHHV-6 have high levels of both cell-associated and cell-free HHV-6 DNA in the absence of active infection². For these reasons, clinical studies should exclude, or at least consider separately, such individuals. Although in this study viral load data are not specified for each PCR-positive patient (only mean values and ranges are presented in the text), the numbers of HHV-6 DNA copies at the high end in both plasma (6x10⁵mL) and blood cells (2.1x10⁶/1.5x10⁶ cells) are those typically detected in individuals with ciHHV-6^2,3. Active HHV-6 infection rarely causes a viral load of over 1 million copies per ml and in these situations the elevated viral load is transient and correlates with dramatic clinical manifestations. Since all of the patients with HHV-6 viremia in this study had mild disease, ciHHV-6 status can be presumed in at least some of these patients.

The estimated prevalence of ciHHV-6 is approximately 0.85% among blood donors and newborns in the US and UK^{2, 6}. However, the prevalence of ciHHV-6 in two small studies of renal transplant patients was 1.92% and 2.13%, respectively^{8, 9}. Based on these figures, one might expect to find 4 ciHHV-6 individuals out of 200 as tested in this study.

Another issue that deserves attention is the low level of HHV-6 latency (6-9%) reported by Csoma et al., which suggests a poor PCR sensitivity, making it difficult to interpret the data on the incidence of active HHV-6 infection. Since HHV-6, especially variant B, is a ubiquitous virus that infects >95% of the population at an early age and persists in a latent form predominantly in blood cells^{10, 11}, the majority of adults harbor HHV-6 DNA sequences in their blood cells.
Finally, the authors report that 8 out of 9 patients with viremia had the A variant of HHV-6, which is in contrast with previous studies of post-renal transplant patients in which the B variant was predominantly detected^{12, 13}. However, detection of the two variants is influenced by the compartment investigated as well as, possibly, by the method used. HHV-6B is found more frequently in PBMCs while HHV-6A is found more frequently in plasma. Nitsche et al. used a variant-specific PCR and found HHV-6A in 92% of plasma samples, but in only 4% of PBMC samples from 336 HSCT patients¹⁴.
Lack of identification of ciHHV-6 has led to a great deal of confusion in the HHV-6 field. The finding of high HHV-6 DNA levels in the blood of mildly ill or asymptomatic individuals with ciHHV-6 may lead to the faulty conclusion that HHV-6 can replicate at significant levels without causing disease, as also suggested in this study. A simple quantitative test on whole blood can establish ciHHV-6 status in almost all cases. Conversely, serum and plasma PCR DNA tests are not sufficient to identify ciHHV-6 status¹⁵. Confirmation can be made by hair follicle or fingernail DNA PCR testing^{15, 16}.

Sincerely,

Paolo Lusso, National Institute of Allergy and Infectious Diseases, NIH, USA
Dharam Ablashi, HHV-6 Foundation, USA
Kristin Loomis, HHV-6 Foundation, USA
Sylvie Rogez, Department of Virology, CHRU Dupuytren, France
Louis Flamand, Rheumatology and Immunology Research Center, Université Laval, Canada

1. Csoma E, Meszaros B, Gall T, Asztalos L, Konya J, Gergely L: Dominance of variant A in Human Herpesvirus 6 viraemia after renal transplantation, Virol J 2011, 8:403
2. Leong HN, Tuke PW, Tedder RS, Khanom AB, Eglin RP, Atkinson CE, Ward KN, Griffiths PD, Clark DA: The prevalence of chromosomally integrated human herpesvirus 6 genomes in the blood of UK blood donors, J Med Virol 2007, 79:45-51
3. Ward KN, Leong HN, Thiruchelvam AD, Atkinson CE, Clark DA: Human herpesvirus 6 DNA levels in cerebrospinal fluid due to primary infection differ from those due to chromosomal viral integration and have implications for diagnosis of encephalitis, J Clin Microbiol 2007, 45:1298-1304
4. Brands-Nijenhuis AV, van Loo IH, Schouten HC, van Gelder M: Temporal relationship between HHV 6 and graft vs host disease in a patient after haplo-identical SCT and severe T-cell depletion, Bone Marrow Transplant 2010,
5. Tohyama M, Hashimoto K: [Drug-induced hypersensitivity syndrome], Nippon Rinsho 2007, 65 Suppl 8:341-343
6. Hall CB, Caserta MT, Schnabel KC, Boettrich C, McDermott MP, Lofthus GK, Carnahan JA, Dewhurst S: Congenital infections with human herpesvirus 6 (HHV6) and human herpesvirus 7 (HHV7), J Pediatr 2004, 145:472-477
7. Pellett PEaGT: Human herpesviruses 6, 7, and 8. Edited by J. Versalovic KCC, G. Funke, J.H. Jorgensen, M.L. Landry, and D.W. Warnock,. Washington, DC, ASM Press, 2011,
8. Lee SO, Brown RA, Eid AJ, Razonable RR: Chromosomally integrated human herpesvirus-6 in kidney transplant recipients, Nephrol Dial Transplant 2011, 26:2391-2393
9. Kidd IM, Clark DA, Sabin CA, Andrew D, Hassan-Walker AF, Sweny P, Griffiths PD, Emery VC: Prospective study of human betaherpesviruses after renal transplantation: association of human herpesvirus 7 and cytomegalovirus co-infection with cytomegalovirus disease and increased rejection, Transplantation 2000, 69:2400-2404
10. Clark DA, Ait-Khaled M, Wheeler AC, Kidd IM, McLaughlin JE, Johnson MA, Griffiths PD, Emery VC: Quantification of human herpesvirus 6 in immunocompetent persons and post-mortem tissues from AIDS patients by PCR, J Gen Virol 1996, 77 ( Pt 9):2271-2275
11. Yoshikawa T, Goshima F, Akimoto S, Ozaki T, Iwasaki T, Kurata T, Asano Y, Nishiyama Y: Human herpesvirus 6 infection of human epidermal cell line: pathogenesis of skin manifestations, J Med Virol 2003, 71:62-68
12. Chapenko S, Folkmane I, Ziedina I, Chistyakovs M, Rozentals R, Krumina A, Murovska M: Association of HHV-6 and HHV-7 reactivation with the development of chronic allograft nephropathy, J Clin Virol 2009, 46:29-32
13. Loginov R, Karlsson T, Hockerstedt K, Ablashi D, Lautenschlager I: Quantitative HHV-6B antigenemia test for the monitoring of transplant patients, Eur J Clin Microbiol Infect Dis 2010, 29:881-886
14. Nitsche A, Muller CW, Radonic A, Landt O, Ellerbrok H, Pauli G, Siegert W: Human herpesvirus 6A DNA Is detected frequently in plasma but rarely in peripheral blood leukocytes of patients after bone marrow transplantation, J Infect Dis 2001, 183:130-133
15. Ward KN, Leong HN, Nacheva EP, Howard J, Atkinson CE, Davies NW, Griffiths PD, Clark DA: Human herpesvirus 6 chromosomal integration in immunocompetent patients results in high levels of viral DNA in blood, sera, and hair follicles, J Clin Microbiol 2006, 44:1571-1574
16. Hubacek P, Virgili A, Ward KN, Pohlreich D, Keslova P, Goldova B, Markova M, Zajac M, Cinek O, Nacheva EP, Sedlacek P, Cetkovsky P: HHV-6 DNA throughout the tissues of two stem cell transplant patients with chromosomally integrated HHV-6 and fatal CMV pneumonitis, Br J Haematol 2009, 145:394-398

Value of cell studies that reflect the natural situation and whether, histiocyte-derived U-937 cells are the cell-line of choice, for studies of HIV infection interactions of monocytes and macrophages

Garry Lynch — 2011-10-27T16:29:52Z

In the article by Ferrucci et al `Cellular phenotype impacts human immunodeficiency virus type 1 viral protein R subcellular localization¿ Virology Journal 8: 397 (10th of August) 2011 (1), applauded is their approach taken in this study of HIV Vpr protein localization, using cells that more closely represent the types of cells infected in nature as compared to numerous previous studies that deployed laboratory cell-lines (e.g., HeLa cells) derived from cell types that do not naturally harbour HIV infection. The latter approach has undoubtedly been useful and beneficial in many different situations to reveal various important aspects of HIV infection and its interactions. However, given the inherent and specialised compositions and proteome(s) of each different cell type that underpins their size, shapes and activities in the complex body system it is reasonable and pertinent to consider whether for other situations red herring or un-natural blind-ends similarly may have also been collaterally promulgated. And some for which we may still remain in the dark of their true biologic activities in their natural cell environment? Perhaps some of the differences in the HIV Vpr localisation (viz cytoplasmic versus nuclear) between cell types that are representative of naturally infected cells (this present study) versus laboratory manipulated cell targets may have uncovered one such instance.

Accepting the limitations of cell-lines as stand-ins for primary cells such as fresh PBMC¿s, raised is the question of whether U-937 cells is the ideal cell-line choice to in-general and collectively represent native monocytes and differentiated macrophages for HIV studies. As it is well known that there are significant differences in the HIV infection of monocytes and their more differentiated macrophage forms (2), it is unreasonable to lump them together as synonymous entities, and although related should be separately demarcated as monocytes and macrophages. U-937 cells were originally derived and developed from a case of histiocytic lymphoma (3). As such histiocytes (ergo U-937 cells) from bone marrow derived myeloid progenitors (4), differ from monocytes, as they are already well differentiated, and committed resident cells of connective tissue and are relatively inactive and immobile compared with other monocytes/macrophages. Furthermore as U-937 cells express cell surface markers that differ from primary monocytes (5, 6), they poorly represent monocytes. Perhaps a follow up study by these investigators is necessary to establish whether the localisation of Vpr in monocytes is the same as for macrophages. Proposed therefore is whether the monocytoid cell line THP-1 (7), derived from a monocytic leukemia, may have been a more preferred cell-line to represent monocytes. And for which THP-1 stimulation to a differentiated macrophage phenotype can be achieved by treatment with phorbol ester (8), that would enable comparison of the spectrum of monocyte to macrophage HIV Vpr protein locales¿ to be realised. With the proviso of course that such laboratory cell-line findings are validated against primary monocytes and macrophages.

With the above technicality raised, the present study by Ferrucci et al is an appropriate addition to the field using cell lines that more closely represent those myeloid-lineage cells infected in nature, than earlier cell studies of unrelated lineage, and necessary to demarcate the Vpr cell localisation properties (and possibly other HIV related proteins) of cells infected in vivo.