prion replication—once again blaming the dendritic cell
TRANSCRIPT
NATURE MEDICINE • VOLUME 5 • NUMBER 11 • NOVEMBER 1999 1235
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THE CULPRIT UNDERLYING all transmissiblespongiform encephalopathy (TSE)
diseases is believed to be a proteina-ceous infectious pathogen known asscrapie prion (PrPSc) (ref. 1). All prion-related diseases are believed to sharethe same basic pathogenic mechanism,involving the structural conversion ofnormal cellular prion (PrPC) into an in-fectious form. This infectious PrPreplicates, and eventually causes theneurological disorders associated withTSE (ref. 1).
Accumulating evidence suggests thatthe host lymphoid system is involved inthe infectious PrP replication leading tothe development of TSE (refs. 2–5).However, the identity of the cell type(s)responsible for prion replication re-mains controversial. Previous studieshave reported that mice lacking B cellsare resistant to PrPSc infection by the pe-ripheral route6, whereas more recent
studies have reported that B cells are notthe direct targets of infectious prion.However, B cells are required for the dif-ferentiation of other cell types that maybe directly involved in the propagationof PrPSc (ref. 7). The exact identity ofthese cells is not known, but it is knownthat they are present in the spleen andnot in the peripheral blood8.
In this issue of Nature Medicine, Brownet al. provide evidence that folliculardendritic cells (FDCs), which reside inthe spleen and depend on B-cell signalsfor maturation, are required for thereplication of a mouse ME7 scrapiestrain9. They report that FDCs them-selves produce PrPC, and that PrPSc repli-cation in mouse spleen occurs inPrPC-expressing FDCs, but not other
bone marrow (BM)-derived cell types,such as lymphocytes and myeloid cells.This conclusion is based on experimentswith chimeric mice. Using irradiatedSCID mice receiving BM transplants,Brown et al. tested the potential for PrPSc
replication in mice bearing PrP+/+ orPrP–/– FDCs, which are derived from thehost, along with PrP+/+ or PrP–/– lympho-cytes and myeloid cells, which were de-rived from BM grafts.
They report that when host FDC andBM-derived donor cells both expressPrPC, PrPSc, injected by the peritoneal orthe intracerebral routes, replicates inthe spleen and leads to TSE–related dis-ease (Fig. 1a). However, when the FDCfrom the host mice do not express PrPC,but the BM-grafted cells do, PrPSc repli-cation does not occur in the spleen, andthe mice remain healthy. This suggeststhat FDC expression of PrPC is necessaryfor PrPSc replication (Fig. 1b). Finally,
Prion replication—once again blaming the dendritic cellThe lymphoid system is known to be involved in the propagation and spread of scrapie. However, the identity of
the cell type responsible for scrapie replication remains controversial. A new study provides evidence that thefollicular dendritic cells in the spleen are the targets of this infectious form of prion (pages 1308–1312).
MAN-SUN SY &PIERLUIGI GAMBETTI
suboptimal anti-retroviral therapy.What measures may be taken to pre-
vent CTL escape and its detrimentalconsequences? Efforts to boost T-helper cell responses12–13 through im-munotherapeutic vaccination, or tobroaden the CTL response14, for exam-ple through peptide-pulsed dendriticcell infusions16, are approaches nowbeing developed. Further efforts shouldaim to improve our understanding ofthe mechanism by which particularHLA molecules seem to affect speed toprogression14,16, a finding also sug-gested by the study of Evans et al.Similarly, we need to gain a better un-derstanding of the factors that deter-mine the immunodominance of theCTL response, and to develop vaccinescapable of inducing effective CTL ac-tivity. Further studies of CTL escape inHIV and SIV infection will help to elu-cidate the precise conditions in whichevasion of the immune response occursand, with viral load measurements,should provide important insights intothe relative contribution of these mu-tations to disease progression.
Future therapeutic efforts thereforeneed to focus not only on anti-retrovi-
ral treatment, but also on harnessingthe immune response to obtain optimaland durable control of viremia.
1. Ogg, G.S. et al. Quantitation of HIV-1-specific cy-totoxic T lymphocytes and plasma load of viralRNA. Science 279, 2103–2106 (1998).
2. Schmitz, J.E. et al. Control of viremia in simian im-munodeficiency virus infection by CD8+ lympho-cytes. Science 277, 333–338 (1999).
3. Evans, D.T. et al. Virus-specific CTL responses se-lect for amino-acid variation in Env and Nef.Nature Med. 5, 1270–1276 (1999).
4. Phillips, R.E. et al. Human immunodeficiency virusgenetic variation that can escape cytotoxic T cellrecognition. Nature 354, 453–459 (1991).
5. Brander, C. & Walker, B.D. The HLA class I re-stricted CTL response in HIV infection: systematicidentification of optimal epitopes. In: HIVMolecular Immunology Database. (eds. Korber,B.T.M. et al.) 14–26 (Los Alamos NationalLaboratory Press, Los Alamos, New Mexico,1998)
6. Ossendorp, F. et al. A single residue exchangewithin a viral CTL epitope alters proteasome-me-diated degradation resulting in lack of antigenpresentation. Immunity 5, 115–124 (1996).
7. Brander, C. et al. Efficient processing of the im-munodominant, HLA-A*0201 restricted HIV-1CTL epitope despite multiple variations in the epi-tope flanking sequences. J. Virol. (in the press).
8. Goulder, P.J.R. et al. Late escape from an immun-odominant cytotoxic T-lymphocyte response as-sociated with progression to AIDS. Nature Med. 3,212–217 (1997).
9. Koenig, S. et al. Transfer of HIV-1-specific cyto-toxic T lymphocytes to an AIDS patient leads toselection for mutant HIV variants and subsequentdisease progression. Nature Med. 1, 330–336(1995).
10. Borrow, P. et al. Antiviral pressure exerted by HIV-1-specific cytotoxic T lymphocytes (CTLs) duringprimary infection demonstrated by rapid selec-tion of CTL escape virus. Nature Med. 3, 205–211(1997).
11. Price, D.A. et al. Positive selection of HIV-1 cyto-toxic T lymphocyte escape variants during pri-mary infection. Proc. Natl. Acad. Sci. USA 94,1890–1895 (1997).
12. Rosenberg, E.S. et al. Vigorous HIV-1-specificCD4+ T-cell responses associated with control ofviraemia. Science 278, 1447–1450 (1997).
13. Kalams, S.A. et al. Association between virus-spe-cific CTL and helper responses in HIV-1 infection.J. Virol. 73, 6715–6720 (1999).
14. Carrington, M. et al. HLA and HIV-1:Heterozygote advantage and B*35-Cw*04 disad-vantage. Science 283, 1748–1752 (1999).
15. Dhodapkar, M.V. et al. Rapid generation of broadT-cell immunity in humans after a single injectionof mature dendritic cells. J. Clin. Invest. 104,173–180 (1999).
16. Kaslow, R.A. et al. Influence of human MHC geneson the course of HIV infection. Nature Med. 2,405–411 (1996).
1Partners AIDS Research Center
Massachusetts General Hospital
13th Street, Bldg 149, Rm 5218
Charlestown, Massachusetts 02120, USA
Email: [email protected] AIDS Research Center
Massachusetts General Hospital
13th Street, Bldg 149, Rm 5212D
Charlestown, Massachusetts 02129, USA
Email: [email protected]
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1236 NATURE MEDICINE • VOLUME 5 • NUMBER 11 • NOVEMBER 1999
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when host FDCs express PrPC, butdonor lymphoid and myeloid cells donot, PrPSc replication does occur in thespleen and the mice develop disease(Fig. 1c). This suggests that FDC expres-sion of PrPC is sufficient for PrPSc repli-cation.
These results raise questions regard-ing the mechanisms of PrPSc replicationin FDCs. Why are FDCs uniquely sus-ceptible to infectious forms of PrP,whereas other hematopoietic cells areresistant? Is this due to some uniquefeatures of the FDCs cell in the spleen(such as their anatomical location, nor-mal physiologic functions or morphol-ogy)? Are the normal PrPC glycoformsexpressed in FDC different quantita-tively or qualitatively from the PrPC gly-coforms expressed in other cell types?By what mechanisms do the infectedFDCs transmit PrPSc into the central ner-vous system? When the infectious PrP isinjected intracerebrally, what is themechanism by which infectious PrPtravels from the central nervous systemto the FDCs in the spleen? Are similardendritic cells in other anatomical sitesalso susceptible to infection (e.g.Langerhans’ cells in the skin)?
These findings also raise interestingquestions about the role of FDCs in the
pathogenesis of human prion diseases.Both kuru and new variant Creuzfeldt-Jakob disease are contracted by the con-sumption of contaminated tissues.Therefore, the infectious PrP must enterthe host through the gastrointestinaltract, where the first cell type the infec-tious agents encounters is the epithelialcells. We have found that high levels ofnormal PrPC are present in the epithelialcells lining the gastrointestinal tract ofmice and human (Liu, T. et al., unpub-lished results), and infectious PrPs havebeen detected in the gastrointestinaltract of naturally infected sheep10. Butso far, all the experiments that suggestinvolvement of PrPSc replication in theFDC have been carried out either byperitoneal or intracerebral infection.Therefore, it will be important to deter-mine whether the FDCs in the spleenare also required for replication whenthe animals are infected orally. If FDCsare indeed involved in infection by theoral route, then what are the mecha-nisms by which infectious PrP travelfrom epithelial cells to the FDC in thespleen? Nonetheless, the observationthat PrPSc replicate in FDCs even whenthey are presented intracerebrally mayhave implications for other forms ofhuman prion diseases that are not orally
transmitted, such as in cases of inher-ited or sporadic prion diseases. It will beuseful to learn if infectious PrP is pre-sent in the FDC in the spleen of thesepatients.
The results provided by Brown andcolleagues argue that follicular dendriticcells are involved in the pathogenesis ofthe prion-related diseases. However,most of the evidence presented is basedon immunohistochemical stainingusing a dendritic cell-specific mono-clonal antibody and a polyclonal anti-body against normal PrPC. Althoughcolocolization and immunohistochemi-cal staining in tissues can be informa-tive, they are not the most sensitive orquantitative methods of protein detec-tion. Additional in vitro experimentswill be required to determine whetherFDCs alone are sufficient for infectiousPrP replication. Recent advances in theunderstanding of the growth and differ-entiation of DCs should improve ourability to maintain these cells in cultureand answer this question.
The levels of the normal PrPC expres-sion can also influence the develop-ment of prion diseases1, and the levelsof normal PrPC expression inhematopoietic cells are quite differentbetween human and mouse.Experiments using monoclonal anti-bodies against prion have shown thatnormal PrPC is expressed at high levelsin most of the human peripheral bloodleukocytes, including T cells, B cells,monocytes and dendritic cells.However, only a small population ofmurine leukocytes expresses detectablelevels of normal PrPC (Liu, et al., submit-ted). Therefore, additional studies willbe needed to determine whether obser-vations made in murine models are di-rectly relevant to human prion disease.It is possible that leukocytes in humanand mouse may play different parts inthe pathogenesis.
Dendritic cells are believed to be a tar-get of HIV infection, and may be an es-sential reservoir for HIV. Data nowsuggest that these same cells are likelyto be important in prion infection.Follicular dendritic cells are ideally lo-cated in the spleen, where they serve tocapture blood-borne foreign antigens.Unfortunately, in HIV and prion infec-tion, these cells may be doing moreharm than good, by storing the infec-tious agent and allowing it to replicateand spread to other cell types.Determination of methods to keep PrPSc
Fig. 1 Bone marrow cells obtained from either PrP+/+ or PrP–/– mice were transplanted into irradiatedSCID/PrP+/+ or SCID/PrP–/– mice. Grafted mice were challenged with the ME7 scrapie strain either in-tracerebrally or intraperitoneally 28 days after grafting. Disease development was determined histo-logically and confirmed by a standard clinical end-point bioassay.
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NATURE MEDICINE • VOLUME 5 • NUMBER 11 • NOVEMBER 1999 1237
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MANY ANIMAL MODELS and ongoing clin-ical trials have used suicide-gene
therapy for the treatment of gliomas inthe brain. A common theme of thesetherapies is the delivery of the herpessimplex virus-1 thymidine kinase (HSV-1-TK) gene through replication-incom-petent adenoviral (Ad) or retroviralvectors to tumor cells. After viral infec-tion, gancyclovir treatment selectivelykills transduced cells and non-specifi-cally damages other cells in close prox-imity, by what is known as the‘bystander effect’1. In this issue ofNature Medicine, Dewey et al. raise im-portant concerns about the long-termeffects of adenoviral-mediated suicide-gene therapy in the brain2.
Dewey et al. studied the effects of ade-noviral-mediated suicide in the rat CNS-1 tumor model, which has been shownto accurately reflect the infiltrativetumor growth observed in humangliomas3. Treatment of CNS-1 tumorswith a single dose of Adv/HSV-1-TK, fol-lowed by gancyclovir, induced chronicinflammation that could still be de-tected at 3 months after therapy.Despite tumor eradication, chronic ac-tive inflammation was demonstrated bythe presence of macrophages and CD8+
T cells at the original tumor site, in ad-dition to microglia and astrocyte activa-tion. There was localized demyelinationin the area of the original tumor. Unlikeprevious studies using adenoviral vec-tors in the brain, Dewey et al. reportthat HSV-1-TK transgene expression isstable, and can be detected for up to 3months after therapy. The distribution
of HSV-1-TK was widespread, and ex-pression was found in both the ispilat-eral and contralateral hemispheres. Toour knowledge this is the first study todocument chronic inflammation afterAdv/HSV-1-TK suicide-gene therapy inthe brain, and raises important issues toconsider in developing strategies fortreatment of brain tumors.
It is important to determine what fac-tors induce the chronic inflammationseen after Adv/HSV-1-TK tumor therapy.There are three potential explanations,each of which have been addressed inthe paper by Dewey et al. The first is thatthe response is directed toward tumorantigen(s) released when CNS-1 cells arekilled and the debris are eliminated bymacrophages. This response would betumor-specific and favorable, as itwould theoretically induce an anamnes-tic response that might prevent tumorrecurrence. The authors demonstratethat this is a reasonable conclusion, asperipheral priming of rats with mito-mycin C-treated CNS-1 cells protectedagainst a lethal intracranial challenge ofCNS-1 cells.
A second possibility is that the chronicinflammation is vector-induced. This re-sponse would not be desirable, as itwould indicate that the immunity wouldnot be tumor-specific. Dewey et al.demonstrate that intracerebral injectionof Adv/HSV1-TK alone leads to the localaccumulation and persistence of CD8+
lymphocytes at the injection site up to 3months after viral inoculation. Thischronic inflammation may be the resultof persistent adenoviral antigen expres-sion, due to the ‘leaky’ nature of first-generation replication-incompetentadenoviral vectors4. Therefore, presenta-tion of these de novo-synthesized viralproteins in the context of major histo-compatibility molecules in the brainmay lead to the generation of anti-viralimmunity manifested as chronic inflam-mation. However Dewey et al. rule outthis possibility by demonstrating thatHSV1-TK expression in the hemispherecontralateral to the tumor does not in-duce chronic inflammation.
Another possibility is thatmacrophages may be infected with re-combinant adenovirus and chronicallystimulate antigen-specific T cells.Indeed, it has been shown thatmacrophages can be infected with re-combinant adenoviral vectors5. Deweyet al. report strong HSV1-TK immunore-activity that overlaps with macrophagedistribution in the ipsilateral hemi-sphere of the brain. However, it is notclear if macrophages participate in theinitiation of the chronic inflammationseen after Adv/HSV1-TK therapy in thismodel system.
A final possibility is shown by experi-ments that demonstrate the synergy be-tween vector and gancyclovirtreatments. A single intracerebral injec-tion of combined Adv/HSV1-TK andgancyclovir led to increased numbers ofCD8+ T cells in the brain 3 months aftertherapy, as compared with what was ob-
Inflammatory thoughts about glioma gene therapyGene therapy for treatment of glioma often involves delivery of herpes simplex virus-1 thymidine kinase gene. A
new study shows that this approach can induce chronic inflammation, and raises important questions about currentadenoviral-based clinical trials (pages 1256–1263).
TAMMY KIELIAN &WILLIAM F. HICKEY
infected cells from spreading their cap-tured prey may be a useful approach toTSE therapy that may also be applicableto other infectious diseases.
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10. Van Keulen, L.J., Schreuder, B.E., Vromans, M.E.,Langeveld, J.P. & Smits M.A. Scrapie-associatedprion protein in the gastrointestinal tract of sheepwith natural scrapie. J. Comp. Pathol. 121, 55–63(1999).
Division of Neuropathology
Institute of Pathology
Case Western Reserve University
School of Medicine
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Email: [email protected] or
Email: [email protected]
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