Antibodies against lytic and latent Kaposi's sarcoma-associated herpes virus antigens and lymphoma in the European EpiLymph case-control study.

BACKGROUND: Kaposi's sarcoma-associated herpes virus is associated with primary effusion lymphoma and multicentric Castleman's disease. METHODS: Seropositivity to lytic and latent Kaposi's sarcoma herpes virus (KSHV) antigens were examined in 2083 lymphomas and 2013 controls from six European countries. RESULTS: Antibodies against KSHV latent and lytic antigens were detectable in 4.5% and 3.4% of controls, respectively, and 3.6% of cases (P>0.05). The KSHV seropositivity was associated with splenic marginal zone lymphoma (SMZL) (odds ratio (OR)=4.11, 95% confidence interval (CI)=1.57-10.83) and multiple myeloma (OR=0.31, 95% CI=0.11-0.85). CONCLUSION: The KSHV is unlikely to contribute importantly to lymphomagenesis among immunocompetent subjects. However, the observed association with SMZL may underline a chronic antigen mechanism in its aetiology.

HIV-specific cytotoxic T-cells in HIV-exposed but uninfected Gambian women.

A crucial requirement in the rational design of a prophylactic vaccine against the human immunodeficiency virus (HIV) is to establish whether or not protective immunity can occur following natural infection. The immune response to HIV infection is characterized by very vigorous HIV-specific cytotoxic T-lymphocyte (CTL) activity. We have identified four HIV-1 and HIV-2 cross-reactive peptide epitopes, presented to CTL from HIV-infected Gambians by HLA-B35 (the most common Gambian class I HLA molecule). These peptides were used to elicit HIV-specific CTLs from three out of six repeatedly exposed but HIV-seronegative female prostitutes with HLA-B35. These women remain seronegative with no evidence of HIV infection by polymerase chain reaction or viral culture. Their CTL activity may represent protective immunity against HIV infection.

PIP: A crucial requirement in the rational design of a prophylactic vaccine against HIV is to establish whether or not protective immunity can occur following natural infection. The immune response to HIV infection is characterized by very vigorous HIV-specific cytotoxic T-lymphocyte (CTL) activity. Four HIV-1 and HIV-2 cross-reactive peptide epitopes were identified, presented to CTL from HIV-infected Gambian women by HLA-B35 (the most common Gambian class 1 HLA molecule). The study population consisted of 20 women: 14 had been prostitutes for more than 5 years and reported little condom usage and 6 were long-term sexual partners of HIV-infected men. Peptide-stimulated cultures were also set up from 8 known seropositive donors with HLA-B35 or B53, and from a control group of volunteers at low-risk of HIV infection with HLA-B35 (12 Gambian and 7 European) and 2 Gambians with HLA-B53. Specific CTL activity against one or more peptides was repeatedly detected after 10-14 days in the peptide-stimulated cultures from 3 of the 6 high-risk seronegative women with HLA-B35, but not in their three counterparts with HLA-B53 nor in any of the low-risk volunteers. The strongest responses were generated toward the HIV-1 pol peptide, which lies close to the active site of reverse transcriptase, and to the nef peptide, which is conserved between HIV-1 and -2. HIV-specific CTL in seronegative subjects could potentially be a response to acute HIV infection, before the development of antibodies, but the women were still seronegative and virus-culture negative 3 months after the CTL were first detected, making recent infection extremely unlikely. These women remain seronegative with no evidence of HIV infection by polymerase chain reaction or viral culture. Their CTL activity may represent protective immunity against HIV infection.

Seroepidemiology of human herpesvirus 8 (HHV-8) infection in injecting drug users.

A cross-sectional study was carried out in injecting drug users (IDUs) from Greece to assess the seroprevalence of human herpesvirus 8 (HHV-8) and to identify potentially associated risk factors. A total of 288 IDUs were tested for K8.1 antibodies to HHV-8 lytic antigen. Associations between HHV-8 serostatus and potential risk factors were examined using univariate and multivariate logistic regression analysis. Seroprevalence of HHV-8 was 24.3% (95% CI 19.5-29.7), increasing with age from 19.4% in those aged <30 years to 52.9% in those aged 40 years (P for trend=0.003). No statistically significant associations between HHV-8-positive status and gender, educational level, age at first drug injection, needle sharing, number of imprisonments, complications from drug overdose, HIV and HCV were observed. In the multivariate logistic regression analysis, older age (40 vs. <40 years, OR 3.30, 95% CI 1.14-9.56) and report of septicaemia/abscess (yes vs. no, OR 1.80, 95% CI 1.01-3.18) were each independently associated with higher HHV-8 seroprevalence. HHV-8 is highly prevalent in the IDU population in Greece. The independent association between HHV-8 and reported abscess or septicaemia supports the hypothesis that poor hygiene conditions in the setting of drug injection may contribute to HHV-8 transmission.

Treatment of classic Kaposi sarcoma with a nicotine dermal patch: a phase II clinical trial.

BACKGROUND: Kaposi sarcoma (KS), a malignancy of dermal endothelial cells that is caused by human herpesvirus 8 (HHV8) infection, is sensitive to perturbations of immunity. Nicotine might be effective against KS because of its immunologic and vascular effects and because smoking is associated with a low risk of KS. OBJECTIVE AND STUDY DESIGN: We conducted a masked, randomized phase 2 clinical trial of transdermal nicotine and placebo patches to assess the safety and efficacy of nicotine against classic KS (cKS). SUBJECTS AND METHODS: Three cKS lesions, predominantly nodules, in each of 24 non-smoking patients were randomly assigned to 15 weeks continuous treatment with nicotine patch (escalated to 7 mg), identical masked placebo patch or no patch. Changes in lesion area and elevation from baseline through six follow-up visits, by direct measurement and by two independent readers using digital photographs of the lesions, were compared using non-parametric and regression methods. Changes in longitudinal levels of HHV8 antibodies and DNA in blood cells were similarly assessed. RESULTS: There were no systemic or serious adverse events, and compliance was good. One patient resumed smoking and discontinued patches, and two patients withdrew at week 12 for unrelated indications. Six (29%) of the remaining 21 suspended use of patches to relieve local skin irritation; four of these six completed the trial at reduced dose. Treatment assignment was not associated with significant or consistent changes in cKS lesion area or elevation, HHV8 viral load or antibodies. CONCLUSION: Transdermal nicotine and placebo patches caused no serious toxicities but had no demonstrable effect on nodular cKS lesions or HHV8 levels.

Whole-genome association study of antibody response to Epstein-Barr virus in an African population: a pilot.

Epstein Barr virus (EBV) infects 95% of the global population and is associated with up to 2% of cancers globally. Immunoglobulin G (IgG) antibody levels to EBV have been shown to be heritable and associated with developing malignancies. We, therefore, performed a pilot genome-wide association analysis of anti-EBV IgG traits in an African population, using a combined approach including array genotyping, whole-genome sequencing and imputation to a panel with African sequence data. In 1562 Ugandans, we identify a variant in human leukocyte antigen (HLA)-DQA1, rs9272371 (p = 2.6 × 10-17) associated with anti-EBV nuclear antigen-1 responses. Trans-ancestry meta-analysis and fine-mapping with European-ancestry individuals suggest the presence of distinct HLA class II variants driving associations in Uganda. In addition, we identify four putative, novel, very rare African-specific loci with preliminary evidence for association with anti-viral capsid antigen IgG responses which will require replication for validation. These findings reinforce the need for the expansion of such studies in African populations with relevant datasets to capture genetic diversity.

Activity of thalidomide in AIDS-related Kaposi's sarcoma.

PURPOSE: To assess the toxicity and activity of oral thalidomide in Kaposi's sarcoma (KS) in a phase II dose-escalation study. PATIENTS AND METHODS: Human immunodeficiency virus (HIV)-seropositive patients with biopsy-confirmed KS that progressed over the 2 months before enrollment received an initial dose of 200 mg/d of oral thalidomide in a phase II study. The dose was increased to a maximum of 1,000 mg/d for up to 1 year. Anti-HIV therapy was maintained during the study period. Toxicity, tumor response, immunologic and angiogenic factors, and virologic parameters were assessed. RESULTS: Twenty patients aged 29 to 49 years with a median CD4 count of 246 cells/mm(3) (range, 14 to 646 cells/mm(3)) were enrolled. All patients were assessable for toxicity, and 17 for response. Drowsiness in nine and depression in seven patients were the most frequent toxicities observed. Eight (47%; 95% confidence interval [CI], 23% to 72%) of the 17 assessable patients achieved a partial response, and an additional two patients had stable disease. Based on all 20 patients treated, the response rate was 40% (95% CI, 19% to 64%). The median thalidomide dose at the time of response was 500 mg/d (range, 400 to 1,000 mg/d). The median duration of drug treatment was 6.3 months, and the median time to progression was 7.3 months. CONCLUSION: Oral thalidomide was tolerated in this population at doses up to 1,000 mg/d for as long as 12 months and was found to induce clinically meaningful anti-KS responses in a sizable subset of the patients. Additional studies of this agent in KS are warranted.

Kaposi's sarcoma-associated herpesvirus and Kaposi's sarcoma in Africa. Uganda Kaposi's Sarcoma Study Group.

BACKGROUND: Endemic Kaposi's sarcoma (KS) is a clinically and epidemiologically distinct human immunodeficiency virus negative form of KS occurring in Africa. Kaposi's sarcoma is now the most frequently reported cancer in some areas of Africa. OBJECTIVE: To determine if a KS-associated herpesvirus (KSHV) is present in both endemic HIV-seronegative and HIV-seropositive KS lesions from African patients. METHODS: Paraffin-embedded tissue specimens from Ugandan patients with KS and non-KS tumor control patients attending a university-based oncology clinic were examined in a blinded case-control study. Tissue DNA specimens were examined for detectable KSHV genome by nested polymerase chain reaction performed at two independent laboratories. RESULTS: We identified KSHV in 17 (85%) of 20 KS tissue specimens from HIV-seronegative patients and 22 (92%) of 24 KS tissue specimens from HIV-infected persons. Kaposi's sarcoma lesions from four HIV-infected persons and four HIV-seronegative persons were positive for KSHV. Unlike previous studies in North America and Europe, three (14%) of 22 non-KS cancer control patients' tissue specimens were also positive for KSHV that resulted in an overall odds ratio of 49.2 (95% confidence interval, 9.1 to 335) for detecting KSHV in KS lesions from patients in Uganda. CONCLUSION: As in North America and Europe, KSHV infection is strongly associated with both HIV-seropositive and HIV-seronegative KS in Africa. However, it is likely that infection with this virus is more highly prevalent in Uganda.

HIV-2 antisera cross-neutralize HIV-1.

The neutralization properties of three independent HIV-2 isolates were examined in comparison with four diverse HIV-1 strains. Human sera containing antibodies specific to HIV-2 can cross-neutralize HIV-1. By contrast, HIV-1 sera are group-specific and have no neutralizing effect on HIV-2. Therefore, HIV-2 antigens may be important components for the development of broadly cross-protective AIDS vaccines.

Variability and evolution of Kaposi's sarcoma-associated herpesvirus in Europe and Africa. International Collaborative Group.

OBJECTIVE: To study the evolution of Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus type 8 in Europe and Africa. DESIGN AND METHODS: PCR and sequence analysis of the variable viral membrane glycoprotein gene K1 in 58 tumour and peripheral blood samples from patients with AIDS-related Kaposi's sarcoma (KS), 'classic' (HIV-negative) KS, transplant KS, Multicentric Castleman's Disease, other lymphoproliferative disorders, and healthy KSHV-infected individuals from the UK, Denmark, Sweden, Italy, Spain, Iceland, The Faroe Islands, Greece, The Gambia and Uganda. RESULTS: Three major groups of K1 sequences were found: A, B and C, as defined previously. The K1 gene has evolved, both within and between these three groups, under positive selection. KSHV group B strains predominate in Africa and are more distant from groups A and C, found in Europe, than A and C are from each other. Within group C two subgroups, C' and C", can be identified. Subgroup C" is more closely related to group A in a region of the K1 protein and appears to be phylogenetically close to the branchpoint between A and C. Group A and C strains are currently found in both HIV-1-infected and -uninfected Europeans, and were already present in Europe before the start of the AIDS epidemic. We found some examples of closely related K1 sequences in Italy and Denmark, but in general KSHV strains in Europe did not cluster geographically. CONCLUSION: KSHV strains in East and West Africa are closely related but phylogenetically distant from those in Europe. The two major KSHV groups in Europe are more closely related, with some strains adopting an intermediate phylogenetic position. In Europe, KSHV strains may have been disseminated at least several decades ago. Variability in the K1 region is driven by selection and does not correlate with different KSHV-related pathologies or geographic regions where clinically more aggressive HIV-negative KS ('endemic' KS) is more common.

Prevalence and distribution of HHV-8 in different subpopulations, with and without HIV infection, in Spain.

OBJECTIVE: To estimate the seroprevalence of HHV-8 in several Spanish subpopulations with different risk levels of acquiring HIV-1 infection and from different geographical regions. DESIGN: Cross-sectional seroprevalence study. METHODS: A total of 1699 serum samples from blood donors (613), children under the age of 12 years (100), injecting drug users (IDU) (382), heterosexuals attending a sexually transmitted disease (STD) clinic (273) and homosexual men attending a STD clinic or a HIV-based hospital unit (331) were analysed for anti-HHV-8 antibodies. The presence of antibodies against HHV-8 was tested with an indirect immunofluorescence assay (IFA). A subsample of HHV-8-positive samples was also tested for antibody titre against HHV-8. RESULTS: The overall seroprevalence of antibodies against HHV-8 for the blood donor population was 6.5% (7.0% in Andalusia, 8.0% in Catalonia and 4.5% in the Basque Country). None of the children tested positive for HHV-8. The HHV-8 prevalence was 86.7% in HIV-positive homosexual men and 28.0% in HIV-negative homosexual men (P < 0.001). Of heterosexual men attending STD clinics, 17.2% tested positive for HHV-8; 11.5% of IDU tested positive for HHV-8. HHV-8 antibody titres by groups parallel the distribution of HHV-8 prevalence. No association between HHV-8 antibody titres and CD4 cell count or HIV viral load was identified. CONCLUSIONS: The HHV-8 prevalence among blood donors in Spain is higher than in Northern Europe and the USA, but is similar to that in Northern Italy. The distribution of HHV-8 is compatible with a sexually transmitted agent. The distribution of HHV-8 correlates with that of Kaposi's sarcoma but factors other than HHV-8 seem to explain the Kaposi sarcoma distribution.

Detection of human herpesvirus 8 DNA in semen from HIV-infected individuals but not healthy semen donors.

OBJECTIVE: To ascertain the prevalence of Kaposi's sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus (HHV) type 8, and cytomegalovirus (CMV) DNA in semen was investigated. METHODS: Amplification by nested polymerase chain reaction was used to detect viral DNA sequences in samples from 24 HIV-infected gay men, 15 of them with Kaposi's sarcoma (KS), and 115 healthy donors. RESULTS: Six of the 24 HIV-infected patients had detectable HHV-8 DNA in their semen: three of the 15 patients with KS and three of the nine patients without KS. CMV DNA was detected in 20 semen samples from HIV-infected patients. None of the semen samples from healthy donors had detectable HHV-8 DNA and rates of CMV DNA detection were low (3%). CONCLUSIONS: The study demonstrates the presence of HHV-8 in semen from HIV-infected individuals with, or at risk, of developing KS and the potential for sexual transmission of the virus. We found no evidence of HHV-8 in the semen of HIV-uninfected donors.

Human herpesvirus type 8 and Kaposi's sarcoma.

Kaposi's sarcoma-associated herpesvirus or human herpesvirus type 8 (HHV-8) is present in all forms of Kaposi's sarcoma (KS) as well as in primary effusion lymphomas and some cases of Castleman's disease. In KS tissues, HHV-8 is present in endothelial and spindle cells. Current serologic tests suggest that HHV-8 is predominantly found in those at risk of KS and is not as widespread as most other human herpesviruses. HHV-8 encodes various proteins that may play a role in promotion of cellular growth, including cyclin- and G-coupled protein receptor homologues, and anti-apoptotic proteins, including Bcl-2, IL-6 (i.e., interleukin 6), and FLIP (i.e., FLICE inhibitory protein) homologues. In addition, HHV-8 encodes two macrophage inflammatory-like proteins with anti-human immunodeficiency virus and angiogenic potential.

Posterior circulation aneurysms. Results of direct operations.

Twenty-eight patients with aneurysms of the posterior circulation were managed by direct operations, 53.6% of these being done within 14 days of a subarachnoid hemorrhage. Transvenous cardiac pacing was used in 4 of the 13 patients with aneurysms of the basilar bifurcation in order to produce a short period of profound hypotension while final dissection and clip application was performed. There were nine patients with posterior-inferior cerebellar artery/vertebral-junction aneurysms, the only one dying having a large aneurysm. The operative technique used in the majority of the 13 patients with basilar bifurcation aneurysms involved subtotal temporal lobectomy, and the operative exposure so obtained was excellent. Five patients died, one of whom rebled from an aneurysm of the basilar artery origin that could only be wrapped, the total mortality being 17.9%. Seventeen (74%) of the 23 survivors were assessed as showing good results; 5 (22%) as showing fair results, and only 1 (4%) as showing a poor result.

The fetal fibroblast: the effector cell of scarless fetal skin repair.

Human fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (fetal) fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations: cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal fibroblasts can heal fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the fetal fibroblast is the major effector cell for scarless fetal skin repair.

Endoscopic creation and repair of fetal cleft lip.

In utero repair of several life-threatening malformations in the human fetus is now a clinical reality, yet fetal surgery still poses significant risks to both the mother and the unborn child. Preterm labor is a major problem and is directly related to the large hysterotomy required for fetal exposure. Endoscopic surgical manipulation of the fetus, through small uterine "ports," solves this problem and may eventually permit fetal intervention for non-life-threatening malformations. In this pilot study we demonstrated the feasibility of performing endoscopic surgery on the fetus in situ. A lip incision was created and repaired using endoscopic microsurgical techniques in midgestation fetal lambs. This represents the first application of this technique for in utero fetal intervention.

A model for fetal cleft lip repair in lambs.

Fetal wounds heal without inflammation and scar formation. This phenomenon may, in the future, be applicable to human cleft lip and palate repair. However, extensive experimental work must first be done to document the benefits of in utero repair. We developed a large animal model for creation and repair of a complete cleft lip and alveolus using fetal lambs. The cleft lip and alveolus deformity was created in eight 75-day-gestation fetuses (term = 145 days) and either repaired in three layers or left unrepaired. There were four sham-operated fetuses, and all animals were alive at harvest. Repaired, unrepaired, and control fetuses were harvested at 7, 14, 21, and 70 days following surgery. The unrepaired fetuses demonstrated a complete cleft lip and alveolus with an oronasal fistula. The maxilla was asymmetrical, with the greater segment deviated toward the cleft and with decreased anterior maxillary width. In contrast, repaired cleft lip and alveolus animals showed no scar, normal thickness of the lip, and a symmetrical maxilla. Histologic analysis of the repaired wounds showed evidence of tissue regeneration without scar formation. The results of this preliminary study indicate that the fetal lamb cleft lip and alveolus model is technically feasible with an excellent survival rate. Healing occurs without scar formation. In the repaired animals, the maxilla was symmetrical. This model will be used to document facial growth following in utero repair of a cleft lip and alveolus.

Studies in fetal wound healing, VI. Second and early third trimester fetal wounds demonstrate rapid collagen deposition without scar formation.

The mechanisms that underlie the lack of scarring in fetal wounds are unknown, but probably relate to the control of collagen fibrillogenesis. The role of collagen in the fetal wound matrix is controversial, and several wound implant models have been used to evaluate collagen deposition in fetal wounds. Unfortunately, these models create an artificial wound environment and may thereby affect the results. In order to study fetal wound collagen deposition in linear wounds without artificially altering the wound environment, we applied a highly sensitive immunohistochemical technique that uses antibodies to collagen types I, III, IV, and VI. We found that collagen was deposited in fetal wounds much more rapidly than in adult wounds. Wound collagen deposition occurred in a normal dermal and mesenchymal pattern in second and early third trimester fetal lambs. These findings are consistent with the observation that the fetus heals rapidly and without scar formation. In contrast, wounds in late gestation fetal lambs showed some evidence of scar formation. Further studies may suggest ways to alter the adult wound so that it heals in a fetal manner.

Studies in fetal wound healing: III. Early deposition of fibronectin distinguishes fetal from adult wound healing.

Wound healing in the fetus proceeds through a series of steps that differ in the fetus and the adult. At each phase of this complex process, there is signaling between the tissue cells and the wound microenvironment, signals that are mediated by and through the extracellular matrix. We postulate that these signals occur earlier in fetal wounds, resulting in more rapid repair. To investigate this, we compared the first 24 hours of wound healing in the rabbit fetus and adult, using antibodies against key extracellular matrix macromolecular components: laminin, fibronectin, and type-specific collagens I, III, IV, and V. Fibronectin was the first matrix component to be deposited, and was visualized as early as four hours after fetal wounding and 12 hours after adult wounding. There was no evidence of new laminin or collagen deposition in either the fetal or adult wounds at any time point examined. The early deposition of fibronectin, a matrix adhesion molecule that provides a scaffolding for epithelial migration, may underlie the rapid reepithelialization observed in fetal wounds.

The evaluation of velopharyngeal function using flexible nasendoscopy.

Nasendoscopy is an essential tool in assessing the dynamic function and structure of the velopharyngeal sphincter during speech and swallowing. Flexible fibre-optic nasendoscopy has been used by the cleft palate team at Withington Hospital, Manchester since 1989. Seventy-six patients were referred between 1989 and 1994 for evaluation of velopharyngeal function during speech. Flexible nasendoscopic evaluation was attempted in 50 patients, and successfully carried out in 43 patients. The age range was four years to 77 years (mean 21 years). The patients were divided into two groups: Group 1 consisting of patients with cleft palate and Group 2 comprised of patients with non-overt cleft palate-related velopharyngeal dysfunction of various aetiologies; such as, submucous cleft, post-tonsillectomy, post-adenoidectomy, neurological and post-traumatic. Based on the findings on nasendoscopy, videofluoroscopy and clinical speech/voice analysis the following treatment options were recommended: 17 (40 per cent) for pharyngoplasty, five (11 per cent) for revision pharyngoplasty, 15 (35 per cent) for speech therapy, four for an obturator and one for tonsillectomy. Two previously undetected submucous clefts were diagnosed.