The effect of dexmedetomidine on postoperative behaviour change in children: a randomised controlled trial.

Children may develop changes in their behaviour following general anaesthesia. Some examples of negative behaviour include temper tantrums and nightmares, as well as sleep and eating disorders. The aim of this study was to determine whether dexmedetomidine reduces the incidence of negative behaviour change after anaesthesia for day case surgery in children aged two to seven years. Children were randomly allocated to one of three groups: a premedication group received 2 mg.kg-1 intranasal dexmedetomidine; an intra-operative group received 1 mg.kg-1 intravenous dexmedetomidine; and a control group. The primary outcome was the incidence of negative behaviour on postoperative day 3 using the Post-Hospitalisation Behaviour Questionnaire for Ambulatory Surgery (PHBQ-AS) and the Strength and Difficulties Questionnaire (SDQ). Secondary outcomes included: the incidence of negative behaviour on postoperative days 14 and 28; anxiety at induction; emergence delirium; pain; length of recovery and hospital stay; and any adverse events. The data for 247 patients were analysed. Negative behaviour change on postoperative day 3 was similar between all three groups when measured with the PHBQ-AS (47%, 44% and 51% respectively; adjusted p=0.99) and the SDQ (median scores 7.5, 6.0 and 8.0 respectively; adjusted p=0.99). The incidence of negative behaviour in the group who received dexmedetomidine intra-operatively was less at postoperative day 28 (15% compared with 36% in the dexmedetomidine premedication group and 41% in the control group, p<0.001). We conclude that dexmedetomidine does not reduce the incidence of negative behaviour on postoperative day 3 in two to seven-year olds having day case procedures.

Intensity-Modulated Radiotherapy With a Simultaneous Integrated Boost in Rectal Cancer.

AIMS: Dose-response curves suggest that higher doses of radiotherapy improve the complete response rate in rectal cancer. The UK adopted the EXPERT trial dose and fractionation, 45 Gy in 25 fractions to the pelvis with a sequential 9 Gy in five fractions to the gross tumour, in patients where the aim was to maximise the complete response. In the Oxford University Hospital NHS Foundation Trust (Oxford, UK) we deliver a biological equivalent dose (BED5) in selected patients using intensity-modulated radiotherapy (IMRT) with a simultaneous integrated boost (SIB) in 25 fractions. We carried out a retrospective analysis of our series to: (i) document the toxicity of this protocol; (ii) ascertain whether dose constraints from RTOG 0822 were appropriate; (iii) assess the response. MATERIALS AND METHODS: The demographics and treatment details for all consecutive patients treated with this protocol were collected using electronic systems. Patients received 45 Gy to the elective nodes and 52 Gy using a SIB to the gross tumour with capecitabine chemotherapy using IMRT or RapidArc plans. Acute toxicity was collected prospectively during weekly reviews. For the purpose of this study, a dedicated gastrointestinal radiologist reviewed all baseline and post-treatment magnetic resonance images and assigned a magnetic resonance tumour regression grade (mrTRG). RESULTS: Seventy-one patients were identified. Seventy completed radiotherapy with a median overall treatment time of 34 days (range 32-36 days); 67.6% received full-dose chemotherapy, with 21.2% receiving a reduced dose. There was a 4.2% incidence of grade 3+ non-haematological toxicity and 1.5% grade 3 + haematological toxicity. 4.2% were admitted during their radiotherapy, with one death due to a pelvic abscess. The RTOG 0822 constraints were achieved in ≥75% of cases, other than the high-dose bladder constraint. mrTRG 1-2 was seen in 47.8%, with mrTRG 1 seen in 23.9%. CONCLUSIONS: We suggest that our protocol shows acceptable acute toxicity, with promising mrTRG results, and could be adopted by centres as an IMRT equivalent dose for EXPERT dose and fractionation.

Neoadjuvant radiotherapy in rectal cancer - less is more?

AIM: There is significant international variation in the use of neoadjuvant radiation prior to total mesorectal excision. The MERCURY group advocate selective neoadjuvant chemoradiotherapy (CRT). We have performed a retrospective, single-centre study of patients treated with CRT, where only the circumferential resection margin is threatened, with the aim of identifying whether a more selective approach to CRT provides acceptable local relapse rates (LRRs). METHOD: All consecutive patients who underwent radical surgery for rectal adenocarcinoma over a 5-year period (2007-2012) in the Oxford University Trust were considered. Electronic hospital systems were reviewed to obtain patient and tumour demographics, treatment and follow-up information. All patients were classified into risk categories according to National Institute for Health and Care Excellence guidance. Data were analysed using Microsoft Excel and R. RESULTS: Two hundred and seventy-two patients were identified: 123, 89 and 60 in the high-, intermediate- and low-risk categories, respectively. Seventy-nine per cent of those in the high-risk group, 6% in the intermediate and 5% in the low-risk group underwent CRT. The overall 5-year LRR and distant recurrence rate (DRR) were 5.2% and 17.8%, respectively. The 5-year LRR for those who went straight to surgery was 2.0% and for those who had neoadjuvant CRT it was 7.4%. The DRR for these two groups was 8.5% and 18.9%, respectively. CONCLUSION: Our series demonstrates that the use of CRT only in margin-threatening tumours, results in an exceptionally low LRR for those without margin-threatening disease. In routine clinical care, this strategy can minimize the significant morbidity of multimodal treatment and allow earlier introduction of systemic therapy to minimize distant recurrence.

The treatment of all MRI-defined low rectal cancers in a single expert centre over a 5-year period: is there room for improvement?

AIM: Outcomes following treatment for low rectal cancer still remain inferior to those for upper rectal cancer. A clear definition of 'low' rectal cancer is lacking and consensus is more likely using a definition based on MRI criteria. This study aimed to determine disease presentation and treatment outcome of low rectal cancer based on a strict anatomical definition. METHOD: A low rectal cancer was defined as one with a lower border below the pelvic attachment of the levator muscles on sagittal MRI. One hundred and eighty consecutive patients with tumours defined by this criterion between 2006 and 2011 were identified from a prospectively managed departmental database. RESULTS: One hundred and eighteen patients (66%) underwent curative resection and 12 (7%) palliative resection. Eleven patients (6%) were entered into a 'watch and wait' (W&W) protocol; 10 others (5%) were not fit to undergo any operation. Some 26 patients (14%) had nonresectable local or metastatic disease. An R0 resection was the most important factor influencing survival after curative surgery. R+ resections occurred in 12% of non-abdominoperineal excisions, 11% of abdominoperineal excisions and 47% of extended resections. Overall survival was similar in the curative resections compared with the W&W patients. In 23 of the 96 (24%) treated with neoadjuvant chemoradiotherapy there was a persistent clinical or a pathological complete response. CONCLUSION: In curative resections, a clear margin is the most important determinant of survival. In 24% of the patients treated with neoadjuvant chemoradiotherapy, surgery could potentially have been avoided. There is scope for improvement in the treatment of locally advanced rectal cancers.

Transanal endoscopic microsurgery for rectal cancer.

Since its introduction in the 1980s, total mesorectal excision (TME) has been the standard surgical technique for treating rectal cancer. This procedure involves removing the rectum and the surrounding envelope of fat along the plane of the mesorectal fascia. Resecting this embryological unit reduces the local recurrence rate by removing all local lymph nodes, including those with occult metastatic disease; however, this surgery is associated with mortality and morbidity. Complications include incontinence for patients given an anastomosis, long-term stoma formation, and sexual and bladder dysfunction. Local excision of rectal cancer using the transanal endoscopic microsurgery (TEM) technique is associated with fewer complications, and therefore, is used as an alternative in specific circumstances. We outline the technique, its indications, imaging appearances and complications.

CD4 T-cell cytokine correlates of diagnostic tests for latent tuberculous infection in HIV-1-infected subjects.

SETTING: Public human immunodeficiency virus (HIV) clinic and tuberculosis (TB) clinics in Kampala, Uganda. OBJECTIVE: To examine TB-specific CD4 T-cell single and polyfunctional cytokine correlates of clinical diagnostic tests for latent tuberculous infection (LTBI) in HIV-1-infected subjects. DESIGN: Thirty antiretroviral therapy-naïve HIV-1-infected adults without active TB disease underwent clinical tuberculin skin test (TST), interferon-gamma release assay (IGRA), and in vitro flow cytometry analysis on cells stimulated with purified protein derivative (PPD) and TB antigens early secreted antigenic target 6 + culture filtrate protein 10 (EC) for frequencies of interleukin (IL) 2, IL-17, interferon-gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) expressing cells. RESULTS: PPD-specific CD4 T-cell expression of TNF-α and IFN-γ was higher in the TST-positive than in the TST-negative group. EC-specific CD4 T-cell expression of TNF-α and IL-2 was higher in the TST+ group than in the TST- group. Expression of both PPD and EC-specific expression of IL-2, IFN-γ and TNF-α were greater in IGRA-positive than in IGRA-negative subjects. The TST+ group exhibited greater polyfunctionality than the TST- group. All cytokine combinations that contained TNF-α correlated strongly with TST size. CONCLUSION: While IL-2, IFN-γ and TNF-α correlate with clinical tests of LTBI, TNF-α is the dominant cytokine correlating with both TST size and magnitude of IGRA response.

How useful is endorectal ultrasound in the management of early rectal carcinoma?

INTRODUCTION: Endorectal ultrasonography (EUS) is used to T stage early rectal tumours and select patients to whom transanal endoscopic microsurgery (TEM) could be offered. Published papers have shown that EUS can have good accuracy, but there is little literature on how EUS influences patient management. The study aim is to ascertain the value of EUS in the management of early rectal tumours. METHODS: Patients with adenomas/early rectal carcinoma being considered for TEM were prospectively studied. Each patient underwent EUS. The surgeon recorded the expected T stage, confidence level of the T stage and management plan for each patient on a proforma before and after the ultrasound result was revealed. Comparison was made between the ultrasound stage and final pathological stage where available. RESULTS: Ninety-six patients were referred over 2 years. Nine were out of reach of the rigid probe and were excluded. Proformas were completed on 53/87 patients (age range 28-87 years, mean age 66 years, 30 males/23 females). Forty-eight patients had a pathological report to compare with the EUS T stage. Ultrasound agreed with the pathological T staging in 43 patients (90%). Patient management was changed in five patients. In 30% of (16/53) patients, EUS increased the confidence level for T staging. CONCLUSION: Although EUS has a high accuracy in predicting the T stage of early rectal cancers, it never changes the management plan for lesions thought to be benign. It seldom changes the pre-operative selection process when clinical examination is considered with other imaging modalities (MRI/CT). EUS should be reserved for answering specific questions in difficult cases rather than for all patients.

Laxative-free CT colonography.

OBJECTIVES: The aim of this study was to determine if the introduction of faecal tagging to CT colonography (CTC) made the examination easier to tolerate or reduced the number of false-positives. METHODS: Our department changed bowel preparation for CT colonography from Picolax (Ferring Pharmaceuticals Ltd, London, UK) to Gastrografin (Bracco Diagnostics Inc, Princeton, NJ) only with a modified diet. Questionnaires were given to a subgroup of patients within these cohorts. The numbers of false-positives were compared between two cohorts before and after this change. false-positives were defined as lesions reported on CT that were not confirmed by subsequent endoscopic examination. Polyps were matched if they were in the same or adjacent segments, and were within 5 mm of the reported size. RESULTS: 412 patients were identified from the Picolax cohort, and 116 from the Gastrografin cohort. 62 patients in each group completed questionnaires. Gastrografin produced less diarrhoea; 34% had five or more bowel motions in the previous day and night, compared with 77% for Picolax (p<0.001), although more patients found drinking it unpleasant compared with Picolax (85% reported drinking Picolax as "easy" vs 61% for Gastrografin; p=0.002). Picolax produced more non-diagnostic examinations, although this difference was not statistically significant. There was not a significant reduction in the numbers of false-positives (2 out of 112 for Gastrografin group, 14 out of 389 for the Picolax group; p=0.54). CONCLUSION: Switching from Picolax to Gastrografin as a CTC preparation technique produced less diarrhoea, but did not reduce the number of false-positives.

Polyfunctional Mycobacterium tuberculosis-specific effector memory CD4+ T cells at sites of pleural TB.

Pleural tuberculosis (TB) is a common presentation of Mycobacterium tuberculosis (MTB) infection, and despite spontaneous resolution remains a strong risk factor for reactivation pulmonary TB in a majority of individuals. This study was undertaken to further understand the characteristics of immune cells at sites of pleural TB. A significant shift toward memory CD4+ T cells with an effector phenotype and away from naïve CD4+ T cells in pleural fluid as compared to blood mononuclear cells was found. These data suggest that effector T cells are capable of migrating to sites of active TB infection and/or the differentiation to effector phenotype T cells in situ is highly amplified. Using multi-parameter flow cytometry analysis, a significant portion of MTB-specific CD4+ T cells in the pleural space were polyfunctional demonstrating two, three or four simultaneous functions including IFN-gamma, IL-2, TNF-alpha, and or MIP-1 alpha production. A greater proportion of these polyfunctional cells were of effector memory rather than central memory phenotype. The role of these polyfunctional MTB-specific CD4+ T cells at sites of pleural TB requires further study.

Adenovirus-based vaccines: comparison of vectors from three species of adenoviridae.

In order to better understand the broad applicability of adenovirus (Ad) as a vector for human vaccine studies, we compared four adenovirus (Ad) vectors from families C (Ad human serotype 5 [HAdV-5; here referred to as AdHu5]), D (HAdV-26; here referred to as AdHu26), and E (simian serotypes SAdV-23 and SAdV-24; here referred to as chimpanzee serotypes 6 and 7 [AdC6 and AdC7, respectively]) of the Adenoviridae. Seroprevalence rates and titers of neutralizing antibodies to the two human-origin Ads were found to be higher than those reported previously, especially in countries of sub-Saharan Africa. Conversely, prevalence rates and titers to AdC6 and AdC7 were markedly lower. Healthy human adults from the United States had readily detectable circulating T cells recognizing Ad viruses, the levels of which in some individuals were unexpectedly high in response to AdHu26. The magnitude of T-cell responses to AdHu5 correlated with those to AdHu26, suggesting T-cell recognition of conserved epitopes. In mice, all of the different Ad vectors induced CD8(+) T-cell responses that were comparable in their magnitudes and cytokine production profiles. Prime-boost regimens comparing different combinations of Ad vectors failed to indicate that the sequential use of Ad vectors from distinct families resulted in higher immune responses than the use of serologically distinct Ad vectors from the same family. Moreover, the transgene product-specific antibody responses induced by the AdHu26 and AdC vectors were markedly lower than those induced by the AdHu5 vector. AdHu26 vectors and, to a lesser extent, AdC vectors induced more potent Ad-neutralizing antibody responses. These results suggest that the potential of AdHu26 as a vaccine vector may suffer from limitations similar to those found for vectors based on other prevalent human Ads.

Radiological detection of post-oesophagectomy anastomotic leak - a comparison between multidetector CT and fluoroscopy.

The purpose of this study was to directly compare CT with fluoroscopy for the diagnosis of occult anastomotic leak following oesophagectomy. Patients undergoing oesophagectomy and gastric conduit formation for the treatment of oesophageal cancer were eligible for inclusion. Imaging was performed 6-8 days post-operatively. Patients underwent multislice CT examination of the chest and abdomen with a bolus of oral contrast, followed by fluoroscopic water-soluble contrast swallow (with subsequent use of barium if this was normal). The studies were reviewed by a consultant radiologist, who was blinded to the results of the other modality. Images were reported as showing "no leak", "possible leak" or "definite leak". The presence of mediastinal gas or fluid or extraluminal contrast at CT was recorded. The clinical outcome after reinstituition of oral intake was used as a reference standard. Patient preference for modality was recorded. 52 patients were recruited. Four were found to have leak on CT and fluoroscopy. 11 had possible leak at CT, but normal fluoroscopy: 2 of these had a leak confirmed later, whereas 9 had no leak. 37 had normal CT and fluoroscopy findings, and remained clinically well. The sensitivity, specificity, positive and negative predictive values were 100%, 80%, 40% and 100%, respectively, for CT, and 67%, 100%, 100% and 96%, respectively, for fluoroscopy. The positive predictive value of mediastinal air, air/fluid and extraluminal contrast were 25%, 75% and 50%, respectively. 35 patients found CT more tolerable. In conclusion, CT was better tolerated and more sensitive but less specific than fluoroscopy for detecting occult anastomotic leak.

An evidence-based review for the management of cystic pancreatic lesions.

The incidental finding of pancreatic cysts is becoming more common because of the increased use of cross-sectional imaging. As a result, the perspective from historical series of symptomatic patients is not always applicable to the current cohort of patients with cystic lesions in their pancreas. In this review, the characteristic radiological features that aid diagnosis are highlighted, and the complementary role of different imaging methods and the appropriate use of tissue sampling are identified. Based on the literature regarding the diagnostic role of imaging in characterizing cystic pancreatic lesions, it is possible to recommend a practical imaging algorithm for the diagnosis of cystic pancreatic lesions.

Analysis of total human immunodeficiency virus (HIV)-specific CD4(+) and CD8(+) T-cell responses: relationship to viral load in untreated HIV infection.

Human immunodeficiency virus (HIV)-specific T-cell responses are thought to play a key role in viral load decline during primary infection and in determining the subsequent viral load set point. The requirements for this effect are unknown, partly because comprehensive analysis of total HIV-specific CD4(+) and CD8(+) T-cell responses to all HIV-encoded epitopes has not been accomplished. To assess these responses, we used cytokine flow cytometry and overlapping peptide pools encompassing all products of the HIV-1 genome to study total HIV-specific T-cell responses in 23 highly active antiretroviral therapy naïve HIV-infected patients. HIV-specific CD8(+) T-cell responses were detectable in all patients, ranging between 1.6 and 18.4% of total CD8(+) T cells. HIV-specific CD4(+) T-cell responses were present in 21 of 23 patients, although the responses were lower (0.2 to 2.94%). Contrary to previous reports, a positive correlation was identified between the plasma viral load and the total HIV-, Env-, and Nef-specific CD8(+) T-cell frequency. No correlation was found either between viral load and total or Gag-specific CD4(+) T-cell response or between the frequency of HIV-specific CD4(+) and CD8(+) T cells. These results suggest that overall frequencies of HIV-specific T cells are not the sole determinant of immune-mediated protection in HIV-infection.

Factors affecting thymic function after allogeneic hematopoietic stem cell transplantation.

Hematopoietic stem cell transplantation (HSCT) is followed by profound immunodeficiency. Thymic function is necessary for de novo generation of T cells after HSCT. Circulating CD45RA(+) naive T-cell levels are predictive of antigen-specific T-cell responses in the absence of graft-versus-host disease (GVHD). These T cells may not represent recent thymic emigrants, since naive T cells may maintain this phenotype if not antigen-activated. To accurately measure thymic output after HSCT and determine the factors that influence thymic function, T-cell receptor excision circles (TRECs) were examined in CD4(+) and CD8(+) cells from a cross-section of patients following HSCT. TREC levels rose weeks after HSCT and could be detected in patients 6 years after HSCT. TREC levels correlated with the frequency of phenotypically naive T cells, indicating that such cells were not expanded progeny of naive T cells present in the donor graft. Chronic GVHD was the most important factor that predicted low TREC levels even years after HSCT. Patients with a history of resolved GVHD had decreased numbers of TREC, compared with those with no GVHD. Because few adults had no history of GVHD, it was not possible to determine whether age alone inversely correlated with TREC levels. Recipients of cord blood grafts had no evidence of decreased TREC induced by immunosuppressive prophylaxis drugs. Compared with unrelated donor grafts, recipients of matched sibling grafts had higher TREC levels. Collectively, these data suggest that thymopoiesis is inhibited by GVHD. Larger studies will be needed to determine the independent contributions of age and preparative regimen to post-transplant thymopoietic capacity.

Functionally inert HIV-specific cytotoxic T lymphocytes do not play a major role in chronically infected adults and children.

The highly sensitive quantitation of virus-specific CD8(+) T cells using major histocompatibility complex-peptide tetramer assays has revealed higher levels of cytotoxic T lymphocytes (CTLs) in acute and chronic virus infections than were recognized previously. However, studies in lymphocytic choriomeningitis virus infection have shown that tetramer assays may include measurement of a substantial number of tetramer-binding cells that are functionally inert. Such phenotypically silent CTLs, which lack cytolytic function and do not produce interferon (IFN)-gamma, have been hypothesized to explain the persistence of virus in the face of a quantitatively large immune response, particularly when CD4 help is impaired. In this study, we examined the role of functionally inert CTLs in chronic HIV infection. Subjects studied included children and adults (n = 42) whose viral loads ranged from <50 to >100,000 RNA copies/ml plasma. Tetramer assays were compared with three functional assays: enzyme-linked immunospot (Elispot), intracellular cytokine staining, and precursor frequency (limiting dilution assay [LDA]) cytotoxicity assays. Strong positive associations were observed between cell numbers derived by the Elispot and the tetramer assay (r = 0.90). An even stronger association between tetramer-derived numbers and intracellular cytokine staining for IFN-gamma was present (r = 0.97). The majority (median 76%) of tetramer-binding cells were consistently detectable via intracellular IFN-gamma cytokine staining. Furthermore, modifications to the LDA, using a low input cell number into each well, enabled LDAs to reach equivalence with the other methods of CTL enumeration. These data together show that functionally inert CTLs do not play a significant role in chronic pediatric or adult HIV infection.

Putative immunodominant human immunodeficiency virus-specific CD8(+) T-cell responses cannot be predicted by major histocompatibility complex class I haplotype.

Recent studies of human immunodeficiency virus (HIV)-specific CD8(+) T cells have focused on responses to single, usually HLA-A2-restricted epitopes as surrogate measures of the overall response to HIV. However, the assumption that a response to one epitope is representative of the total response is unconfirmed. Here we assess epitope immunodominance and HIV-specific CD8(+) T-cell response complexity using cytokine flow cytometry to examine CD8(+) T-cell responses in 11 HLA-A2(+) HIV(+) individuals. Initial studies demonstrated that only 4 of 11 patients recognized the putative immunodominant HLA-A2-restricted p17 epitope SLYNTVATL, suggesting that the remaining subjects might lack significant HIV-specific CD8(+) T-cell responses. However, five of six SLYNTVATL nonresponders recognized other HIV epitopes, and two of four SLYNTVATL responders had greater responses to HIV peptides restricted by other class I alleles. In several individuals, no HLA-A2-restricted epitopes were recognized, but CD8(+) T-cell responses were detected to epitopes restricted by other HLA class I alleles. These data indicate that an individual's overall CD8(+) T-cell response to HIV is not adequately represented by the response to a single epitope and that individual major histocompatibility complex class I alleles do not predict an immunodominant response restricted by that allele. Accurate quantification of total HIV-specific CD8(+) T-cell responses will require assessment of the response to all possible epitopes.

Assessment of thymic output in adults after haematopoietic stem-cell transplantation and prediction of T-cell reconstitution.

BACKGROUND: The potential benefits of haematopoietic stem-cell transplantation are tempered by the depletion of T-cells accompanying this procedure. We used a new technique which quantifies the excisional DNA products of T-cell-receptor (TCR) gene rearrangement to measure thymic output directly in patients with multiple myeloma, and thus assessed the contribution of the thymus to immune recovery after transplantation. METHODS: We studied 40 patients, 34-66 years of age, who had been randomly assigned myeloablative chemotherapy and autologous peripheral-blood haematopoietic stem-cell transplantation with unmanipulated grafts or grafts enriched for CD34 stem cells. CD4 and CD8 T-cell counts were measured, thymic output was estimated serially until 2 years after transplantation, and percentages of naive T-cells were measured. FINDINGS: The production of substantial numbers of new naive T cells by the thymus could be detected by 100 days post-transplant; there was a significant inverse relation between age and recovery of new T cells. In the CD34-unselected group, numbers of TCR-rearrangement excision circles returned to baseline after 2 years, whereas in the CD34-selected group, numbers at 2 years were significantly higher than both baseline numbers (p=0.004), and 2-year numbers in the unselected group (p=0.046). Increased thymic output correlated with, and was predictive of, increased naive T-cell numbers and broader T-cell-receptor repertoires. INTERPRETATION: Our results provide evidence that the adult thymus contributes more substantially to immune reconstitution after haematopoietic stem-cell transplantation than was previously thought, and therefore could be a target for therapeutic intervention.

Vaccination of macaques against pathogenic simian immunodeficiency virus with Venezuelan equine encephalitis virus replicon particles.

Vaccine vectors derived from Venezuelan equine encephalitis virus (VEE) that expressed simian immunodeficiency virus (SIV) immunogens were tested in rhesus macaques as part of the effort to design a safe and effective vaccine for human immunodeficiency virus. Immunization with VEE replicon particles induced both humoral and cellular immune responses. Four of four vaccinated animals were protected against disease for at least 16 months following intravenous challenge with a pathogenic SIV swarm, while two of four controls required euthanasia at 10 and 11 weeks. Vaccination reduced the mean peak viral load 100-fold. The plasma viral load was reduced to below the limit of detection (1,500 genome copies/ml) in one vaccinated animal between 6 and 16 weeks postchallenge and in another from week 6 through the last sampling time (40 weeks postchallenge). The extent of reduction in challenge virus replication was directly correlated with the strength of the immune response induced by the vectors, which suggests that vaccination was effective.

Human immunodeficiency virus type 1-specific cytotoxic T lymphocyte activity is inversely correlated with HIV type 1 viral load in HIV type 1-infected long-term survivors.

HIV-1-specific cytotoxic T cell (CTL) activity has been suggested to correlate with protection from progression to AIDS. We have examined the relationship between HIV-specific CTL activity and maintenance of peripheral blood CD4+ T lymphocyte counts and control of viral load in 17 long-term survivors (LTSs) of HIV-1 infection. Longitudinal analysis indicated that the LTS cohort demonstrated a decreased rate of CD4+ T cell loss (18 cells/mm3/year) compared with typical normal progressors (approximately 60 cells/mm3/year). The majority of the LTSs had detectable, variable, and in some individuals, quite high (>10(4) RNA copies/ml) plasma viral load during the study period. In a cross-sectional analysis, HIV-specific CTL activity to HIV Gag, Pol, and Env proteins was detectable in all 17 LTSs. Simultaneous analysis of HIV-1 Gag-Pol, and Env-specific CTLs and virus load in protease inhibitor-naive individuals showed a significant inverse correlation between Pol-specific CTL activity and plasma HIV-1 RNA levels (p = 0.001). Furthermore, using a mixed linear effects model the combined effects of HIV-1 Pol- and Env-specific CTL activity on the viral load were significantly stronger than the effects of HIV-1 Pol-specific CTL activity alone on predicted virus load. These data suggest that the presence of HIV-1-specific CTL activity in HIV-1-infected long-term survivors is an important component in the effective control of HIV-1 replication.

Humoral, mucosal, and cellular immunity in response to a human immunodeficiency virus type 1 immunogen expressed by a Venezuelan equine encephalitis virus vaccine vector.

A molecularly cloned attenuated strain of Venezuelan equine encephalitis virus (VEE) has been genetically configured as a replication-competent vaccine vector for the expression of heterologous viral proteins (N. L. Davis, K. W. Brown, and R. E. Johnston, J. Virol. 70:3781-3787, 1996). The matrix/capsid (MA/CA) coding domain of human immunodeficiency virus type 1 (HIV-1) was cloned into the VEE vector to determine the ability of a VEE vector to stimulate an anti-HIV immune response in mice. The VEE-MA/CA vector replicated rapidly in the cytoplasm of baby hamster kidney (BHK) cells and expressed large quantities of antigenically identifiable MA/CA protein. When injected subcutaneously into BALB/c mice, the vector invaded and replicated in the draining lymphoid tissues, expressing HIV-1 MA/CA at a site of potent immune activity. Anti-MA/CA immunoglobulin G (IgG) and IgA antibodies were present in serum of all immunized mice, and titers increased after a second booster inoculation. IgA antibodies specific for MA/CA were detected in vaginal washes of mice that received two subcutaneous immunizations. Cytotoxic T-lymphocyte responses specific for MA/CA were detected following immunization with the MA/CA-expressing VEE vector. These findings demonstrate the ability of a VEE-based vaccine vector system to stimulate a comprehensive humoral and cellular immune response. The multifaceted nature of this response makes VEE an attractive vaccine for immunization against virus infections such as HIV-1, for which the correlates of protective immunity remain unclear, but may include multiple components of the immune system.