AIDS-associated cryptococcosis: a comparison of epidemiology, clinical features and outcome in the pre- and post-HAART eras. Experience of a single centre in Italy.

Objectives To assess the prevalence, clinical and immunological characteristics, risk factors and survival of patients with AIDS-related cryptococcosis in the era of highly active antiretroviral therapy (HAART). Methods All newly diagnosed cryptococcosis cases identified retrospectively from among a series of AIDS patients hospitalized consecutively at a single institution in Italy in 1985-1996 (pre-HAART period, n=165) and 1997-2006 (post-HAART period, n=40) were analysed comparatively. Results The prevalence of cryptococcosis decreased from 4.7% (165/3543) to 2.2% (40/1805) between the pre- and post-HAART periods (P=0.0001). There were no differences in the clinical features or immunological status of the patients between the two cohorts. The variables associated with the occurrence of cryptococcosis in the post-HAART era were older age (P<0.001), no previous diagnosis of HIV infection (P<0.001) and infection in homosexual males (P=0.004). During the post-HAART period, immune reconstitution inflammatory syndrome associated with cryptococcosis was observed in five patients (19.3%) a median of 15 weeks after the start of HAART. Thirty-day survival (P=0.045) and overall survival (P=0.0001) were significantly better among patients diagnosed with cryptococcosis in the post-HAART compared to those diagnosed in the pre-HAART era. Conclusions The AIDS-associated cryptococcosis observed in Western countries in the HAART era has similar clinical and immunological characteristics to that observed in the pre-HAART era, but a significantly better outcome.

Post-kala-azar dermal leishmaniasis as an immune reconstitution inflammatory syndrome in a patient with acquired immune deficiency syndrome.

Post-kala-azar dermal leishmaniasis (PKDL) is a complication of visceral leishmaniasis (VL) observed mainly in Sudan and India where it follows treated VL in 50% and 10% of cases, respectively. We report a 46-year-old patient with acquired immune deficiency syndrome who, 7 months after diagnosis of VL, developed PKDL and uveal leishmaniasis following HAART-induced immune recovery. In southern Europe PKDL seems to be an emerging clinical presentation among human immunodeficiency virus (HIV)-infected patients experiencing HAART-induced immune recovery after a previous diagnosis of VL. The best treatment among HIV-infected patients remains to be determined.

Persistent parvovirus B19-induced anemia in an HIV-infected patient under HAART. Case report and review of literature.

Recent reports document resolution of human parvovirus B19-related pure red blood cell aplasia (PB19-PRCA) in HIV-infected patients upon commencement of highly active antiretroviral therapy (HAART). This article describes a patient with PB19-PRCA who, despite fully suppressive HAART, required cyclic administration of intravenous human immunoglobulin over a period of 17 months before PB19 seroconversion and subsequent resolution of relapsing severe anemia. All reports in the English literature describing PB19-related hematologic abnormalities in the post-HAART era are also described herein.

Long-term remission of Kaposi sarcoma-associated herpesvirus-related multicentric Castleman disease with anti-CD20 monoclonal antibody therapy.

Kaposi sarcoma-associated herpesvirus (KSHV)-related multicentric Castleman disease (MCD) is potentially lethal. Growing evidence indicates that, as in Epstein-Barr virus-driven lymphoproliferative disorders after transplantation, KSHV DNA burden in peripheral blood mononuclear cells (PBMCs) may represent the most accurate marker of disease activity. This report describes a patient with human immunodeficiency virus who was followed up clinically and by quantitative polymerase chain reaction for KSHV DNA sequences in PBMCs for more than 3 years following the diagnosis of KSHV-related MCD. Therapy with the antiherpesvirus agent cidofovir, antihuman interleukin-6 antibody BE-8, antiblastic chemotherapy, and combination antiretroviral agents did not achieve durable clinical or virologic remission of the disease. By contrast, administration of the anti-CD20 monoclonal antibody rituximab was well tolerated and allowed a 14-month remission of clinical symptoms and KSHV viremia. Rituximab should be added to the therapeutic armamentarium for KSHV-related MCD.

Human herpesvirus 8 DNA in the skin and blood of patients with Mediterranean Kaposi's sarcoma: clinical correlations.

BACKGROUND: Kaposi's sarcoma is a multifocal lympho-angioproliferative disease that appears in elderly subjects of Mediterranean origin (classical form), young Africans and immunodepressed patients (as a result of organ transplantation or AIDS). In 1994, DNA sequences of a new human herpesvirus, called HHV-8, were detected in skin lesions and peripheral blood of patients with AIDS-related Kaposi's sarcoma by confirmational display analysis and polymerase chain reaction. OBJECTIVE: As HHV-8 in peripheral blood mononuclear cells is detected in about 50% of Mediterranean Kaposi's sarcoma patients and its presence fluctuates in time in the same patient, maybe its detection correlates with the clinical behaviour of the disease. METHODS: By using routine and nested polymerase chain reaction we evaluated the presence of HHV-8-specific DNA sequences in the skin lesions, perilesional healthy skin and peripheral blood mononuclear cells of a group of 40 HIV-negative patients with Mediterranean Kaposi's sarcoma. RESULTS: HHV-8 DNA sequences have been found in 40/40 (100%) lesional skin of Mediterranean Kaposi's sarcoma, in 35/40 (85%) perilesional apparently normal skin and in 24/40 (60%) peripheral blood monuclear cell samples. The results of polymerase chain reaction on peripheral blood monuclear cells were positive in 41% of the patients with slowly evolving disease as opposed to 74% of those with rapidly evolving disease, and in 47.6% of the patients with stage I-II disease as opposed to 73.6% of those with stage III-IV. CONCLUSION: The detection of HHV-8 in peripheral blood monuclear cells seems to correlate with the more aggressive stages and the rapid evolution behaviour of Mediterranean Kaposi's sarcoma.

Activation of latent Kaposi's sarcoma-associated herpesvirus by demethylation of the promoter of the lytic transactivator.

Kaposi's sarcoma-associated herpesvirus (KSHV) is strongly linked to Kaposi's sarcoma, primary effusion lymphomas, and a subset of multicentric Castleman's disease. The mechanism by which this virus establishes latency and reactivation is unknown. KSHV Lyta (lytic transactivator, also named KSHV/Rta), mainly encoded by the ORF 50 gene, is a lytic switch gene for viral reactivation from latency, inasmuch as it is both essential and sufficient to drive the entire viral lytic cycle. Here we show that the Lyta promoter region was heavily methylated in latently infected cells. Treatment of primary effusion lymphoma-delivered cell lines with tetradecanoylphorbol acetate caused demethylation of the Lyta promoter and induced KSHV lytic phase in vitro. Methylation cassette assay shows demethylation of the Lyta promoter region was essential for the expression of Lyta. In vivo, biopsy samples obtained from patients with KSHV-related diseases show the most demethylation in the Lyta promoter region, whereas samples from a latently infected KSHV carrier remained in a methylated status. These results suggest a relationship among a demethylation status in the Lyta promoter, the reactivation of KSHV, and the development of KSHV-associated diseases.

Kaposi's sarcoma-associated herpesvirus LANA2 is a B-cell-specific latent viral protein that inhibits p53.

Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is associated with three proliferative diseases ranging from viral cytokine-induced hyperplasia to monoclonal neoplasia: multicentric Castleman's disease (CD), Kaposi's sarcoma (KS), and primary effusion lymphoma (PEL). Here we report a new latency-associated 1,704-bp KSHV spliced gene belonging to a cluster of KSHV sequences having homology to the interferon regulatory factor (IRF) family of transcription factors. ORFK10.5 encodes a protein, latency-associated nuclear antigen 2 (LANA2), which is expressed in KSHV-infected hematopoietic tissues, including PEL and CD but not KS lesions. LANA2 is abundantly expressed in the nuclei of cultured KSHV-infected B cells. Transcription of K10.5 in PEL cell cultures is not inhibited by DNA polymerase inhibitors nor significantly induced by phorbol ester treatment. Unlike LANA1, LANA2 does not elicit a serologic response from patients with KS, PEL, or CD as measured by Western blot hybridization. Both KSHV vIRF1 (ORFK9) and LANA2 (ORFK10.5) appear to have arisen through gene duplication of a captured cellular IRF gene. LANA2 is a potent inhibitor of p53-induced transcription in reporter assays. LANA2 antagonizes apoptosis due to p53 overexpression in p53-null SAOS-2 cells and apoptosis due to doxorubicin treatment of wild-type p53 U2OS cells. While LANA2 specifically interacts with amino acids 290 to 393 of p53 in glutathione S-transferase pull-down assays, we were unable to demonstrate LANA2-p53 interaction in vivo by immunoprecipitation. These findings show that KSHV has tissue-specific latent gene expression programs and identify a new latent protein which may contribute to KSHV tumorigenesis in hematopoietic tissues via p53 inhibition.

Differential viral protein expression in Kaposi's sarcoma-associated herpesvirus-infected diseases: Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease.

Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is linked to KS, primary effusion lymphomas (PEL), and a subset of multicentric Castleman's disease (MCD). Transcript mapping studies using PEL cell lines have allowed preliminary classification of viral gene expression into constitutive (class I) and inducible (class II/III) categories. To determine whether viral gene expression differs in vivo, we examined tissue sections of KSHV-infected disorders, using specific antibodies against proteins that are representative of the different expression classes of KSHV genes. ORF73/LANA appears to be a surrogate marker for KSHV infection because it is constitutively expressed in vitro and in vivo in all KSHV-infected cells. Expression of vIRF1, vIL6, and PF-8 proteins in the infected B cells of MCD lymph nodes reproduces the expression pattern observed in TPA-stimulated KSHV-infected B-cell lines. In contrast, the protein expression of the inducible viral genes that we tested in KS and PEL biopsies is restricted to PF-8 and vIL6, respectively. The tightly restricted expression of KSHV proteins in vivo differs from the dysregulated expression of inducible KSHV genes in vitro and suggests that viral gene expression in KSHV-infected cell lines does not accurately reflect what occurs in diseased tissues. These differences may be related to either cell-specific or immune restriction of viral replication.

Intractable pruritus in HIV infection: immunologic characterization.

BACKGROUND: Severe, intractable pruritus, often associated with erythematopapular skin lesions and hypereosinophilia, is a condition observed in some nonatopic, HIV-infected patients. We performed immunovirologic analyses of this condition. METHODS: Immunologic (mitogen-stimulated production of cytokines, tumor necrosis factor-alpha [TNF-alpha], and soluble CD23; serum levels of soluble CD23, ICAM-1, TNF-alpha, IgG, IgE, and IgA) and virologic (HIV viral load) parameters were analyzed in six patients with therapy-resistant pruritus. Hypereosinophilia was present in all these patients. Results were compared to those of seven HIV-seropositive individuals similar to the first one in terms of CD4 counts and clinical staging, but without pruritus. RESULTS: Hypereosinophilia; hyper-IgE and hyper-IgA; augmented interleukin (IL)-4, IL-5, and sCD23; and reduced interferon-gamma production by mitogen-stimulated peripheral blood mononuclear cells (PBMC) were detected when patients with pruritus were compared to HIV controls. HIV viral load was also augmented in patients in whom pruritus was present. CONCLUSIONS: The results suggest that therapy-resistant, intractable pruritus accompanied by hypereosinophilia may be used to define a subset of HIV-seropositive individuals showing prototypic hyperactivation of humoral immunity, and in whom augmented HIV viral load is present.

The A-Myb transcription factor is a marker of centroblasts in vivo.

The A-Myb transcription factor is structurally related to the c-myb proto-oncogene and is involved in the control of proliferation and/or differentiation of mature B lymphocytes. We have shown previously by PCR analysis that A-myb is preferentially expressed in CD38+ CD39- sIgM- mature B cells. We demonstrate here, using in situ hybridization, that A-myb expression is restricted to the dark zone of human tonsils and lymph nodes. Furthermore, we show that A-Myb expression is cell cycle regulated both in tonsillar B cells and in Burkitt's lymphoma cell lines, being detectable only in the S and G2/M phases of the cell cycle and not in G0/G1 phase. Strong proliferation of resting human B cells induced in vitro by a variety of physiologic signals, including anti-mu, CD40 ligand, IL-2, IL-4, IL-6, IL-13, IFN-gamma, TNF-alpha, anti-CD19, and anti-CD20, failed to induce A-myb expression, suggesting that proliferation alone is not sufficient for A-myb expression in the absence of induction of a true centroblast phenotype. Finally, we show that differentiation of germinal center B cells in vitro toward either memory or plasma cells is accompanied by rapid down-regulation of A-myb expression. We conclude that A-myb is a marker of centroblasts generated in vivo.

Expression of a virus-derived cytokine, KSHV vIL-6, in HIV-seronegative Castleman's disease.

Castleman's disease is a rare B cell lymphoproliferative disorder related to excess interleukin-6 (IL-6)-like activity. Kaposi's sarcoma-associated herpesvirus (KSHV or HHV8), which encodes a functional cytokine (vIL-6), has been found in some patients with Castleman's disease. Lymph nodes from 14 HIV-seronegative Castleman's disease patients were compared to hyperplastic lymph nodes from 25 HIV-seronegative patients as well as Kaposi's sarcoma lesions from 48 patients for KSHV infection and vIL-6, human IL-6, and Epstein-Barr virus EBER expression. While all Kaposi's sarcoma tissues examined were polymerase chain reaction-positive and all control lymph nodes were polymerase chain reaction-negative for KSHV, none had detectable vIL-6 expression. Six of 14 (43%) Castleman's tissues were positive for KSHV by polymerase chain reaction and all 6 had evidence of vIL-6 expression by immunohistochemistry. vIL-6-positive Castleman's disease patients generally had the multicentric plasma cell variant form of the disease and had a rapidly fatal clinical course frequently associated with autoimmune hemolytic anemia and gammopathy. In contrast, 7 (88%) of the 8 vIL-6-negative Castleman's disease patients had localized disease and have remained disease-free after therapy. KSHV vIL-6 expression appears to be limited to hematopoietic cells and is not present in Kaposi's sarcoma spindle cells. These data suggest that Castleman's disease is a syndrome of multiple etiologies involving aberrant IL-6 activity from either endogenous or viral sources.

Risk of Kaposi's sarcoma-associated herpes virus transmission from donor allografts among Italian posttransplant Kaposi's sarcoma patients.

Kaposi's sarcoma-associated herpesvirus (KSHV) is a newly discovered herpes virus found in all forms of Kaposi's sarcoma (KS) including KS among immunosuppressed transplant patients. It is unknown whether this virus is transmitted by organ transplantation or is reactivated during immunosuppression among those patients infected before transplantation. To investigate the risk of KSHV transmission during organ transplantation, we conducted a case-control study of transplant recipients with and without KS matched to their respective donors. Sera were collected at time of transplantation and tested in a randomized and blinded fashion using four KSHV serologic assays testing for antibodies to both latent and lytic phase antigens. Ten (91%) of 11 organ recipients who developed KS were seropositive prior to transplantation by one or more of the assays compared with two (12%) of 17 control organ recipients (OR = 75, 95% CI = 4.7, 3500). KS cases were more likely to have been born in southern Italy where KS is endemic than the recipient controls or either donor group. Only four (36%) of 11 donors to case patients and three (18%) of 17 donors to control patients were seropositive (P = .38, two-tailed Fisher's exact test). KSHV transmission could not be ruled out for the single KS patient seronegative at transplantation and clear evidence for organ-related transmission was found for another KS patient outside of the case-control study. Antibodies to KSHV are detectable in the sera from most transplant recipients before initiation of immunosuppressive treatment suggesting that KS among immunosuppressed transplant patients is primarily due to virus reactivation. KSHV transmission, however, from an infected allograft can occur, and our study reports the first documented case of person-to-person transmission of KSHV.

Simultaneous onset of primary cutaneous B-cell lymphoma and human herpesvirus 8-associated Kaposi's sarcoma.

We report the simultaneous occurrence of Kaposi's sarcoma (KS) and primary cutaneous B-cell lymphoma (CBCL) of the leg in a 79-year-old woman, seronegative for HIV-1, HTLV-1 and HTLV-2. The CBCL underwent complete clinical remission after local radiotherapy, whilst the KS became disseminated within a year following diagnosis. However, 2 years after the diagnosis of KS, the patient died with neurological symptoms. These were presumed to be due to involvement of the central nervous system by lymphoma, although in the absence of an autopsy, this could not be proven. Skin biopsies from the original KS and CBCL lesions, as well as short-term culture of spindle cells from the KS lesion and peripheral blood mononuclear cells (PBMC), were studied by semiquantitative polymerase chain reaction (PCR) using primers specific for DNA sequences of a novel gamma-herpesvirus-8 (HHV-8). PCR studies were strongly positive for the virus on KS cells and PBMC; conversely, a low viral load was found on CBCL cells. A high titre of serum IgG antibodies reacting with the nuclei of the HHV-8 positive cell line BCP-1 was found. These data suggest that reactivation of latent infection with HHV-8 had occurred in this patient, and that HHV-8 is directly involved in KS, but not in CBCL of the leg, an aggressive variant of CBCL.