Formalin inactivation of the lactate dehydrogenase-elevating virus reveals a major neutralizing epitope not recognized during natural infection.

Five hybridomas that secrete monoclonal antibodies which neutralize the infectivity of lactate dehydrogenase-elevating virus (LDV) were isolated from BALB/c mice primed with Formalin-inactivated LDV. Competition analyses indicated that all five neutralizing monoclonal antibodies recognize contiguous, if not identical, epitopes on the envelope glycoprotein of LDV (VP-3) which are not recognized by nonneutralizing VP-3-specific monoclonal antibodies isolated from the same fusion. Despite the presence of neutralizing activity, polyclonal anti-LDV antibodies obtained from persistently infected mice did not compete for binding to LDV with four of the five neutralizing monoclonal antibodies tested. The results indicate that the envelope glycoprotein of LDV possesses a major neutralizing epitope which is poorly recognized, if at all, by mice during a natural infection but is rendered immunogenic by Formalin inactivation of the virus. The epitope was also not immunogenic in a rabbit, since its polyclonal LDV-neutralizing antibodies did not inhibit binding of the mouse monoclonal antibodies to LDV. Passive immunization with the neutralizing monoclonal antibodies did not protect mice from LDV infection and did not alter the course of infection. Neutralizing monoclonal antibodies have been used to select a neutralization escape variant by a novel combination of in vitro and in vivo isolation.     

Monoclonal antibody protection from age-dependent poliomyelitis: implications regarding the pathogenesis of lactate dehydrogenase-elevating virus.

Over 90% of cyclophosphamide-treated, 6- to 7-month-old C58/M mice developed fatal paralytic disease after infection with a virulent strain of lactate dehydrogenase-elevating virus (LDV), with a mean onset of paralysis of about 16 days. Passive immunization with polyclonal antibodies or with a group of anti-LDV monoclonal antibodies (MAbs) with single-epitope specificity 1 day before or at the time of LDV infection prevented the development of paralytic disease without interfering with the replication of LDV in permissive macrophages, the primary host cells of LDV. In situ hybridization of spinal cord sections with an LDV-specific cDNA probe indicated that the MAb specifically prevented the cytocidal infection of motor neurons by LDV without blocking the infection of smaller nonneuronal cells in the spinal cord. The protective antibodies recognize at least two different epitopes on the glycoprotein of LDV, VP-3. Passive immunizations with other anti-LDV MAbs, which recognize at least three other epitopes on VP-3 of LDV, afforded no protection. In contrast to the protective effect of anti-LDV MAb injection before or at the time of LDV infection, their administration postinfection exerted relatively little protection, though it delayed the appearance of paralytic symptoms. However, repeated injections of MAbs until at least 7 days postinfection also afforded a high degree of protection. The results indicate that protective MAbs may interfere with two stages in the development of LDV-induced paralytic disease. When administered at the time of LDV infection, they prevent the initial infection of spinal cord motor neurons. After this initial event, repeated injections of MAb are required to inhibit the spread of LDV between neurons until the endogenous production of protective anti-LDV antibodies in these mice.     

Cytohistologic correlation of urothelial lesions secondary to photodynamic therapy

Qualitative cytologic evaluations of urinary bladder washings were performed on a selected population following photodynamic therapy for recurrent transitional cell carcinoma of the bladder. The seven patients were monitored trimonthly by cystoscopy, multiple biopsies and cytopreparations. Cancers reappeared in two of the five patients who initially responded to therapy. In the remaining two patients, the recurrent neoplasms were therapeutically refractory. Cytology detected recurrent cancer prior to biopsy confirmation and/or cytoscopic identification. Exfoliative cytology was correlated with the histopathology of the concurrent biopsies; a possible source for a false-positive cytodiagnosis was the cellular atypia of reepithelialized bladder mucosa. Dysplasia was not identified cytologically or histologically.     

Role of ultrasonography in diagnosing early rheumatoid arthritis and remission of rheumatoid arthritis - a systematic review of the literature

Introduction

Ultrasonography (US) might have an added value to clinical examination in diagnosing early rheumatoid arthritis (RA) and assessing remission of RA. We aimed to clarify the added value of US in RA in these situations performing a systematic review.

Methods

A systematic literature search was performed for RA, US, diagnosis and remission. Methodological quality was assessed; the wide variability in the design of studies prohibited pooling of results.

Results

Six papers on the added value of US diagnosing early RA were found, in which at least bilateral metacarpophalangeal (MCP), wrists and metatarsophalangeal (MTP) joints were scanned. Compared to clinical examination, US was superior with regard to detecting synovitis and predicting progression to persistent arthritis or RA. Eleven papers on assessing remission were identified, in which at least the wrist and the MCP joints of the dominant hand were scanned. Often US detected inflammation in patients clinically in remission, irrespective of the remission criteria used. Power Doppler signs of synovitis predicted X-ray progression and future flare in patients clinically in remission.

Conclusions

US appears to have added value to clinical examination for diagnosing of RA when scanning at least MCP, wrist and MTP joints, and, when evaluating remission of RA, scanning at least wrist and MCP joints of the dominant hand. For both purposes primarily power Doppler US might be used since its results are less equivocal than those of greyscale US.     

Overlapping motifs (PTAP and PPEY) within the Ebola virus VP40 protein function independently as late budding domains: involvement of host proteins TSG101 and VPS-4

The VP40 protein of Ebola virus can bud from mammalian cells in the form of lipid-bound, virus-like particles (VLPs), and late budding domains (L-domains) are conserved motifs (PTAP, PPxY, or YxxL; where "x" is any amino acid) that facilitate the budding of VP40-containing VLPs. VP40 is unique in that potential overlapping L-domains with the sequences PTAP and PPEY are present at amino acids 7 to 13 of VP40 (PTAPPEY). L-domains are thought to function by interacting with specific cellular proteins, such as the ubiquitin ligase Nedd4, and a component of the vacuolar protein sorting (vps) pathway, tsg101. Mutational analysis of the PTAPPEY sequence of VP40 was performed to understand further the contribution of each individual motif in promoting VP40 budding. In addition, the contribution of tsg101 and a second member of the vps pathway, vps4, in facilitating budding was addressed. Our results indicate that (i) both the PTAP and PPEY motifs contribute to efficient budding of VP40-containing VLPs; (ii) PTAP and PPEY can function as L-domains when separated and moved from the N terminus (amino acid position 7) to the C terminus (amino acid position 316) of full-length VP40; (iii) A VP40-PTAP/tsg101 interaction recruits tsg101 into budding VLPs; (iv) a VP40-PTAP/tsg101 interaction recruits VP40 into lipid raft microdomains; and (v) a dominant-negative mutant of vps4 (E228Q), but not wild-type vps4, significantly inhibited the budding of Ebola virus (Zaire). These results provide important insights into the complex interplay between viral and host proteins during the late stages of Ebola virus budding.     

Cutting edge: OFF cycling of TNF production by antigen-specific CD8+ T cells is antigen independent

Although they are known for their capacity to kill infected cells, Ag-specific CD8(+) T cells elaborate other effector mechanisms, including TNF and IFN-gamma, that contribute to defense against infection. Ag-specific CD8(+) T cells rapidly turn ON and turn OFF IFN-gamma production in direct response to Ag contact, presumably to minimize the potential immunopathology that could result from inappropriate secretion of this inflammatory mediator. In this study, we show, using in vitro propagated and directly ex vivo-analyzed Ag-specific CD8(+) T cells, that in contrast to Ag-dependent ON/OFF cycling of IFN-gamma production, the cessation of TNF production by the same IFN-gamma producing cells is rapid and Ag independent.