Immunological Impairment Research Articles

Immuno-hematological abnormalities of human immunodeficiency virus-1 infected highly active antiretroviral naïve adults in Zaria, Northern Nigeria

Background: In HIV-1-infected adults, hematological derangements include peripheral cytopenias and bone marrow abnormalities. The level of the CD4+ T lymphopenia is presently considered as one of the best markers of HIV induced immune impairment. Multiple interacting factors contribute to the hematological manifestations of HIV infection. We compare the prevalence of peripheral cytopenias and the level of immunological deterioration in treatment naive HIV-1 infected patients. Patients and Methods: Four hundred consecutive HIV-1 infected patients undergoing pre-treatment (Highly active antiretroviral therapy naive) assessment were recruited at the HIV sub-specialty clinic of Ahmadu Bello University Teaching Hospital, Zaria, Nigeria between May 2003 and October 2004. A cross-sectional study of baseline hematological and immunological parameters was undertaken after confirmation of HIV-1 infection by double ELISA methods; complete blood count and CD4+ T cell count (Dynal ® ) were analyzed by manual techniques. Results: Anemia is the most common peripheral cytopenia, 73% in male and 54% in the females. Neutropenia occurred in 10.8% of the patients while 6.25% had thrombocytopenia. Packed cell volume and Absolute lymphocyte count is associated with a reduction of CD4+ T cell counts but this is not consistent following gender and CDC clinical/immunological staging. CD4+ T cell counts decreases significant from asymptomatic stage A through symptomatic stages B to stage C p Conclusion: This study demonstrates that anemia and low CD4+ T cells are the commonest hematological complication in HIV-1 infected patients and these cytopenias deteriorates with increasing advanced stages of the infection.

A potential role of karyopherin a2 in the impaired maturation of dendritic cells observed in glioblastoma patients

Aim: Patients with glioblastomas demonstrate well-documented immunological impairments including decreased numbers of mature dendritic cells (DCs). Recent data identified karyopherin a2 (KPNA2), a nucleocytoplasmic shuttling receptor, as diagnostic and prognostic biomarker for gliomas. The aim of this ongoing study is to correlate parameters of immunity and nucleocytoplasmic transport in glioblastoma patients. Methods: We preoperatively collected serum from 17 patients with glioblastomas and determined DC subsets (HLA DR+ Lin-, CD34-, CD45+, CD123+, CD11+ were analyzed) using a 6-color flow cytometry panel. Expression levels of KPNA2 and nuclear accumulation of p53 were evaluated semi-quantitatively by immunohistochemistry. O 6 -methylguanine DNA methyltransferase (MGMT) and isocitrate dehydrogenase-1 (IDH-1) status were assessed by pyrosequencing and immunohistochemistry, respectively. Results: Median expression levels for both KPNA2 and p53 were 5-10%. IDH-1-R132H mutation and MGMT promoter hypermethylation was detected in 3/16 and 1/9 patients, respectively. Mean counts of total mature DCs, myeloid DCs and plasmacytoid DCs were 9.6, 2.1, 3.4 cells/μL. A preliminary analysis suggests an association between low KPNA2 nuclear expression and increased numbers of mature DCs. However, this correlation did not reach statistical significance so far (P = 0.077). Conclusion: Our preliminary data may indicate a role of KPNA2 in the impaired maturation of DCs observed in glioblastoma patients.

Relevance of αβ-DNTs as Prognostic Factor on Clinical Outcome and Role on Tumor Surveillance in Haematological Diseases and Solid Tumor: Preliminary Evaluations

The mutual interactions between the host immune system and cancer cells may either promote or control oncogenesis. Cancer immunotherapies remain viable approaches to sustain patient survival. However, positive clinical response of phase I/II clinical trials remains still to low. The Double negative T cells (DNTs) have emerged as new subset of T cells that contributes specifically to anti- tumor immunity since involved in immune regulation and tolerance acting as regulatory T cells (Treg) and/or cytotoxic T cells. DNTs express either αβ or γδ T-cell receptors (TCR) or lack CD4, CD8 and CD56. Functional studies showed that DNTs might have a direct in vitro anti-tumor activity against lymphoma and melanoma cells. However no data are available on their prognostic significance, modulation during the therapy response and their ability to kill tumor cells in cooperation with other immune cells such as DCs to explain their role in human anti-lymphoma immunity. Knowledge of the DNTs role in tumor surveillance and as prognostic factor for clinical outcome has a strong clinical valence and might allow us to design new approaches of therapeutic strategy.The aim of this study was to evaluate the DNTs during therapy response in lymph nodes (LN), bone marrow (BM) and peripheral blood (PB) of both lymphoma (Ly), Renal Cell Carcinoma (RCC) and Metastatic Melanoma (MM) patients (pts) in order to assess their role on clinical outcome, progression and tumor surveillance.PB and BM samples of 90 Ly pts, with NHL and cHL were collected at diagnosis and during the follow-up (1-6-12 months after chemo- or immuno-chemotherapy therapy). PB samples of 16 healthy donors were collected as control. Twenty fresh LN tissues from pts clinically suggestive of Ly were quickly processed. To evaluate the interaction of DNTs with tumor burden either 22 samples from RCC pts and 30 samples from MM pts were collected. The ontogeny, functional attitude and TCR clonality of DNT subsets (TCRαβ+ and TCRγδ+) were evaluated by following antibodies: CD3, CD4, CD8, CD56, CD45, TCRαβ, CD45Ra, CD45Ro, CCR7, CD27, CD28, CD30, CD69, GITR, CD95, CD178, CD152, IFN-γ, TNF-α, perforin and granzyme B. For functional studies, DNTs were purified from PBMCs through a negative selection and then cultured for 2 weeks. Data were acquired by 8-colour flow cytometer and analysed using Kaluza software. Other immune cells such as DCs, MDSCs and Treg was also detected to evaluate the correlation with DNTs. 7 cases of LNs received a finally diagnosis of reactive follicular hyperplasia (RFH) so they were considered as controls. All pts provided their informed consent in accordance with the Declaration of Helsinki.We observed a significant decrease (p =0.006) of circulating αβ-DNTs in untreated Ly pts (20.5 cells/ul ± 4.8) as compared with healthy controls (31.3 cells/ul ± 3.4) (fig.1-2) and their number seemed to be modulated during the follow-up. Their frequency in 1-month post-cht or disease relapsed pts was significantly decreased (p=0.006) as compared with the diagnosis as well as when compared indolent with aggressive histotypes (p=0.0001). In HL pts the frequency of αβ-DNT was significantly increased as compared with other histotypes (p=0.005) (Fig.3) Furthermore, the αβ-DNTs in LNs of Ly pts were significantly reduced as compared to to RFH-LNs (p=0.006) (fig.5) and are related to the aggressiveness of the disease. When evaluated either in MM and RRC pts the αβ-DNTs were significantly decreased (p=0.001) as compared with healthy controls and more interestingly when compared with Ly pts (p=0.001) given the greater immunological impairment of RCC/MM tumor burden (fig.6). Finally, ex vivo expanded DNTs acquired an immunomodulatory cytokine profile, characterized by the secretion of IFN-gamma and granzyme B, which are known as central components of anti-tumor immune responses supporting the hypothesis of αβ-DNT potential use in immunotherapeutic strategies (fig.4)Our preliminary data, showed an inverse correlation between the frequency of circulating αβ-DNTs and the tumor condition as well as that they could play an important role in both the development and the progression of lymphomas. This is the first study that compares the frequency of αβ-DNTs in three different pathologies correlating to tumor burden. In addition, it is likely that ex-vivo expanded DNTs exert an anti-tumor activity thus suggesting their possible use as a new strategy for adoptive immune-therapy. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Impaired haemophilus influenzae type b transplacental antibody transmission and declining antibody avidity through the first year of life represent potential vulnerabilities for HIV-exposed but -uninfected infants.

To determine whether immune function is impaired among HIV-exposed but -uninfected (HEU) infants born to HIV-infected mothers and to identify potential vulnerabilities to vaccine-preventable infection, we characterized the mother-to-infant placental transfer of Haemophilus influenzae type b-specific IgG (Hib-IgG) and its levels and avidity after vaccination in Ugandan HEU infants and in HIV-unexposed U.S. infants. Hib-IgG was measured by enzyme-linked immunosorbent assay in 57 Ugandan HIV-infected mothers prenatally and in their vaccinated HEU infants and 14 HIV-unexposed U.S. infants at birth and 12, 24, and 48 weeks of age. Antibody avidity at birth and 48 weeks of age was determined with 1 M ammonium thiocyanate. A median of 43% of maternal Hib-IgG was transferred to HEU infants. Although its level was lower in HEU infants than in U.S. infants at birth (P < 0.001), Hib-IgG was present at protective levels (>1.0 μg/ml) at birth in 90% of HEU infants and all U.S. infants. HEU infants had robust Hib-IgG responses to a primary vaccination. Although Hib-IgG levels declined from 24 to 48 weeks of age in HEU infants, they were higher than those in U.S. infants (P = 0.002). Antibody avidity, comparable at birth, declined by 48 weeks of age in both populations. Early vaccination of HEU infants may limit an initial vulnerability to Hib disease resulting from impaired transplacental antibody transfer. While initial Hib vaccine responses appeared adequate, the confluence of lower antibody avidity and declining Hib-IgG levels in HEU infants by 12 months support Hib booster vaccination at 1 year. Potential immunologic impairments of HEU infants should be considered in the development of vaccine platforms for populations with high maternal HIV prevalence.

Management on bisphosphonate-related osteonecrosis of the jaw in cancer patients

In the treatment of multiple myeloma,bone metastatic tumor of malignant tumors and hypercalcemia, bisphosphonate (BP) administrates according to the BP injection formulation. Since the potency ratio of the bone through intravenous administration is higher than that of the oral medication, and due to the healing delay trendency and immunological deterioration from the effects of steroids and anti-cancer drugs used in the treatment of cancer,bisphosphonate-related osteonecrosis of the jaw(BRONJ) occurs in cancer patients at a high probability.The treatment for BRONJ of patients with cancer is often carried out conservatively, for first priority to suppression of the cancer , and life prognosis. However , the decision often deteriorates the quality of life of the patients.

Quercetin mitigates Adriamycin-induced anxiety- and depression-like behaviors, immune dysfunction, and brain oxidative stress in rats.

This study was designed to evaluate the effect of quercetin, a natural flavonoid, on behavioral alterations, brain oxidative stress, and immune dysregulation caused by a chemotherapeutic agent, Adriamycin (ADR; 7mg/kg of body weight). Different subsets of male Wistar rats were used to determine the benefit of quercetin on ADR-related depression-like and anxiety-like behaviors in the forced swim test, open field, and elevated plus maze, respectively. Quercetin (60mg/kg of body weight) was administered 24, 5, and 1h before the test session of forced swim test (FST) or at the same time points before the elevated plus maze/open field (EPM/OF) tests. Other subsets of rats were sacrificed after quercetin injections to assess the plasma corticosterone level, the brain oxidative status, and the immune cell count. Our results indicate that quercetin alleviated the anxio-depressive-like behavior, attenuated the brain oxidative stress, and suppressed the corticosterone excess that appeared following ADR treatment. The ADR-induced immune disturbance was slightly diminished after quercetin administration, especially for the lymphocyte count. This study suggests that quercetin can mitigate the neurobehavioral and immunological impairments that manifest in ADR-treated rats. Therefore, the combination of quercetin treatment with the chemotherapeutic regimen seems to be beneficial against chemotherapy-related complications.

Manifestaciones estomatológicas en pacientes VIH/SIDA

El presente estudio se realizó en 128 pacientes infectados por el virus de inmunodeficiencia humana, en el Hospital Guillermo Almenara Irigoyen, con la finalidad de encontrar la frecuencia de lesiones orales y la ubicación de éstas en los pacientes VIH/SIDA de acuerdo a la edad y sexo; además establecer la relación existente entre la cifra de linfocitos CD4 con las lesiones detectadas. La muestra fue tomada de manera probabilística; el método utilizado para la recolección de datos fue la observación y la información obtenida se registro en una ficha previamente diseñada. De la muestra 74 pertenecían al sexo masculino y 57 al femenino y sólo el 57,8% presentaban lesiones estomatológicas. En relación a la edad, el mayor número de pacientes se encontraba ubicado en el grupo de 26 a 35 años (43,8%). Las lesiones que se presentaron con mayor frecuencia fueron: la candidiasis pseudomembranosa (21,9%) ubicándose principalmente en lengua, paladar duro y carrillo, úlceras aftosas (14,1%) ubicándose principalmente en carrillo, lengua y labio, la candidiasis eritematosa (10,1%) principalmente en paladar blando, queilitis angular (10,1%) y la leucoplasia vellosa (7%). Existe una relación significativa entre la presencia de candidiasis pseudomembranosa y la leucoplasia oral vellosa con niveles de CD4 menores de 200 cel/mm3 (p&amp;lt; 0,05).

Current understanding of ZIP and ZnT zinc transporters in human health and diseases

Zinc transporters, the Zrt-, Irt-like protein (ZIP) family and the Zn transporter (ZnT) family transporters, are found in all aspects of life. Increasing evidence has clarified the molecular mechanism, in which both transporters play critical roles in cellular and physiological functions via mobilizing zinc across the cellular membrane. In the last decade, mutations in ZIP and ZnT transporter genes have been shown to be implicated in a number of inherited human diseases. Moreover, dysregulation of expression and activity of both transporters has been suggested to be involved in the pathogenesis and progression of chronic diseases including cancer, immunological impairment, and neurodegenerative diseases, although comprehensive understanding is far from complete. The diverse phenotypes of diseases related to ZIP and ZnT transporters reflect the multifarious biological functions of both transporters. The present review summarizes the current understanding of ZIP and ZnT transporter functions from the standpoint of human health and diseases. The study of zinc transporters is currently of great clinical interest.

Fatal Pneumococcal Meningitis in a 7-Year-Old Girl with Interleukin-1 Receptor Activated Kinase Deficiency (IRAK-4) Despite Prophylactic Antibiotic and IgG Responses to Streptococcus Pneumoniae Vaccines

IRAK-4 deficiency causes IL-1R and TLR signaling failure, resulting in minimal clinical features despite invasive bacterial infection. We report the course of a 7-year-old IRAK-4-deficient girl presenting in the first year with multiple occult Staphylococcus aureus lymphadenitis. She was managed with antibiotic prophylaxis (sulfa/trimethoprim/PenV, then - due to neutropenia - Cefprozil), pneumococcal vaccination (PCV-7, Pneumovax23, PCV-13) and vigilance. Pneumococcal-specific IgG levels were monitored. No bacterial infections occurred on prophylaxis for 6 years after initial presentation. IgG response to pneumococcal polysaccharide was satisfactory but short-lived, requiring frequent boosting. At age 7, patient developed a morning headache and vomited once. Cefprozil was administered and re-dosed. Over 12 h, she was fatigued without other symptoms. Low fever accompanied another emesis. A few hours later she was confused, and purpuric rash appeared. Emergency physicians diagnosed sepsis/meningitis and started vancomycin-ceftriaxone. Respiratory failure and cerebellar herniation occurred <24 h after first symptoms. Blood and CSF grew Streptococcus pneumoniae type 6C resistant to second-generation cephalosporins. The patient's latest PCV-13 vaccination was 6 weeks before death, which included serotype 6A. Immunoglobulins were normal except IgG4 was increased (3.4 g/L). IgG response to vaccine antigens was satisfactory. IgG to 6A is reported to cross-react with 6C, but this was not the case here. Despite antibiotic prophylaxis and repeated vaccination, even older IRAK-4-deficient patients are at high risk of rapidly fatal infection due to emergence of antibiotic resistance. These patients need early assessment at any age, bacterial culturing, alternative empiric antibiotic therapy and close observation when even vaguely unwell. Based on increasingly recognized immunological and/or clinical impairments in B cell function, and possibly other defects, long-term IgG prophylaxis in addition to antibiotics is recommended.

The effects of preparation "Histochrome" in biochemical parameters of blood for patients with cardiopathologies

The effects of the small doses of the preparation Gistochrome, containing natural polyhydroxynaphtoquinoine echinochrom A from flat sea urchin Scaphechinus mirabilis, on blood biochemical parameters have been studied in patients with cardiovascular diseases. Gistochrome administration influenced the LPO-antioxidant protection system, indicating reinforcement of antioxidant protection mechanisms. Gistochrome modulated the immune status and the plasma cytokine profile. Thus, Gistochrome may be recommended as means of additional therapy for patients with cardiovascular diseases for correcting the metabolic, immunological and redox impairments.

Role of IL-10 and TGF-β1 in local immunosuppression in HPV-associated cervical neoplasia.

Cervical cancer is a worldwide disease that constitutes a significant public health problem, especially in developing countries, not only due to its high incidence but also because the most affected population comprises women who belong to marginalized socio-economic classes. Clinical and molecular research has identified immunological impairment in squamous intraepithelial cervical lesions and cervical cancer patients. Human Papillomavirus (HPV) has several mechanisms for avoiding the immune system: it down-regulates the expression of interferon and upregulates interleukin (IL)-10 and transforming growth factor (TGF)-β1 to produce a local immunosuppressive environment, which, along with altered tumor surface antigens, forms an immunosuppressive network that inhibits the antitumor immune response. In this review we analyzed the available data on several deregulated cellular immune functions in patients with NIC I, NIC II and NIC III and cervical cancer. The effects of immunosuppressive cytokines on innate immune response, T-cell activation and cellular factors that promote tumor cell proliferation in cervical cancer patients are summarized. We discuss the functional consequences of HPV E2, E6, and E7 protein interactions with IL-10 and TGF-β1 promoters in the induction of these cytokines and postulate its effect on the cellular immune response in squamous intraepithelial cervical lesions and cervical cancer patients. This review provides a comprehensive picture of the immunological functions of IL-10 and TGF-β1 in response to HPV in humans.

Peripheral immunophenotype and viral promoter variants during the asymptomatic phase of feline immunodeficiency virus infection

Feline immunodeficiency virus (FIV)-infected cats enter a clinically asymptomatic phase during chronic infection. Despite the lack of overt clinical disease, the asymptomatic phase is characterized by persistent immunologic impairment. In the peripheral blood obtained from cats experimentally infected with FIV-C for approximately 5 years, we identified a persistent inversion of the CD4/CD8 ratio. We cloned and sequenced the FIV-C long terminal repeat containing the viral promoter from cells infected with the inoculating virus and from in vivo-derived peripheral blood mononuclear cells and CD4 T cells isolated at multiple time points throughout the asymptomatic phase. Relative to the inoculating virus, viral sequences amplified from cells isolated from all of the infected animals demonstrated multiple single nucleotide mutations and a short deletion within the viral U3, R and U5 regions. A transcriptionally inactivating proviral mutation in the U3 promoter AP-1 site was identified at multiple time points from all of the infected animals but not within cell-associated viral RNA. In contrast, no mutations were identified within the sequence of the viral dUTPase gene amplified from PBMC isolated at approximately 5 years post-infection relative to the inoculating sequence. The possible implications of these mutations to viral pathogenesis are discussed.

Micro-RNA Expression Profile Reveals MiR-222 As a Potential Biomarker For EBV-Positive Diffuse Large B-Cell Lymphoma

BackgroundEBV+ diffuse large B-cell lymphoma of the elderly (EBV+DLBCLe) is considered a provisional entity in the latest World Health Organization classification. It affects individuals older than 50 years without prior documented immunodeficiency.This disorder has unfavorable clinical course even after the advent of immunotherapy associated to anthracycline-based chemotherapy. It is linked to Epstein-Barr virus and the physiopathology is related to the presence of the virus itself, senescence and immunological deterioration. Currently there is not a characteristic pattern of expression of microRNAs in EBV+DLBCLe. AimsTo characterize a signature profile for this new entity and to explore microRNAs as biomarkers and potential alternative therapeutic targets for EBV+DLBCLe. Methods124 cases of patients of DLBCL were treated at Hospital Sao Paulo UNIFESP/EPM between 2000 to 2010 and had paraffin blocks available for immunohistochemical and molecular analyses. Seventy-one of 124 patients were more than 50 years and were potential candidates to be considered EBV+DLBCLe (pilot study). In situ hybridization was used for EBV detection (EBER1, Invitrogen) in a tissue microarray slide. Total RNA was obtained from tumor slides using Recover All Total Nucleic Acid Isolation kit (Applied Biosystems). We obtained cDNAs using Megaplex Pools for microRNA Expression (Applied Biosystems). The cDNA was inserted into two platforms containing 384 human microRNA each (Taqman Low Density Arrays) on 7900 Real Time PCR Systems (Applied Biosystems). Data analyses were made in mathematical-statistical environment “R”. The normalization method 2-deltaCt was performed using the endogenous RNU48, and it was identified as the most stable among samples by software Normfinder. It was also used RNU6 recommended by the manufacturer, in a comparative way. MicroRNAs differentially expressed in EBV+ group compared to EBV negative were identified by means of nonparametric tests rank products (RankProd) and Wilcoxon rank-sum (R-Stats). We considered differentially expressed microRNAs which average fold change above or below 1.5.After, real-time quantitative PCR was performed through 7500 Real time PCR Systems (Applied Biosystems) using TaqMan Small RNA kit assays and normalized with RNU48. Results8.5% of cases of DLBCL were considered EBV+DLBCLe after ISH for EBV. 53.1% of the pilot study were considered GCB and 43.9% non-GCB according to Hans et al. algorithm (2004); 73.7% were classified as worse prognosis (groups 3 and 4) according to Salles et. al(2011) model combining IPI and immunohistochemical markers (bcl-2 and Ki67).We selected four of EBV+ and four of EBV negative samples matched by age, gender, stage and IPI to be analyzed in the PCR platforms. We found 10 deregulated microRNAs among the two groups. However, only seven microRNAs achieved statistically significant differences and would be the start point of a microRNA signature profile proposal to be validated in a larger multicentric cohort (total of 29 EBV+DLBCLe versus 65 DLBCL). Among them let-7g, miR-126, miR-146a, miR-146b, miR-150 and miR-155 were overexpressed in EBV+DLBCLe comparing to EBV-negative DLBCL whereas miR-151 was underexpressed. After validation in 29 EBV+DLBCLe (including 23 new cases) and 65 EBV negative cases we confirmed overexpression of miR-126 in 75.8% (median 2.14 vs 0.14,p< 0.0001), miR-146a in 62% (median 1.94 vs 0.49, p = 0.0035) ,miR-146b in 51.7% (median 1.51 vs 0.11 , p< 0.0001),miR-150 in 96.5 % (median 20.54 vs 2.56,p< 0.0001) and miR-222 in 23,8% of cases (median 0.67 vs 0.08,p< 0.0001,Mann-Whitney) and also confirmed underexpression of miR-151 in 96% of EBV+DLBCL cases. Although miR-222 was overexpressed in ¼ of the cases, it showed high speficity (98%) and positive predictive value (83%), Area Under the Curve= 0.87774, when EBV+ when compared to EBV negative cases. Summary /ConclusionThe merit of the present study is to propose a microRNA signature for a recently described disease and to highlight miR-222 as a possible biomarker and therapeutic target for EBV+DLBCLe. The main routes deregulated are NF-KappaB and PI3K-AKT pathway, being PTEN a target of the overexpressed miR-222. Thus, the findings suggest that antagomiRs for miR-222,that are being tested in some types of cancer, could be also used as adjuvant therapy to R-CHOP in EBV+DLBCL. (Supported by FAPESP 2010/17668-6). Disclosures:No relevant conflicts of interest to declare.

Intraclonal Complexity Of CLL Fractions In Cell Proliferation Rates, Gene Expression Signatures, and Responses To Autologous T-Cell Help In Peripheral blood and Secondary Lymphoid Tissues

Chronic lymphocytic leukemia (CLL) is a disease of resting and dividing B cells. In order to understand the trafficking and transition of proliferative and resting fractions between periphery and lymphoid tissues, we have tried to [1] define the relative levels of CLL B-cell proliferation in distinct anatomic sites, [2] identify and characterize CLL intraclonal fractions from these sites that differ in time since last replication, and [3] model the abilities of these subsets to repopulate immune deficient mice. In the blood, resting and recently-divided CLL cells can be distinguished by surface expression of CXCR4 and CD5. CXCR4dimCD5br (DIM) cells are an activated subset, CXCR4intCD5int (INT) cells are the clonal bulk, and CXCR4brCD5dim (BR) cells are older and more quiescent. Here we compared in vivo proliferation rates, gene expression profiles, and differences in xenografting of these 3 fractions. In addition, we studied the same parameters in 3 other fractions, CXCR4dimCD5dim (dDIM), CXCR4intCD5br (INT/BR) and CXCR4brCD5br (dBR).CLL B-cell kinetics was analyzed in 7 treatment-naïve patients who drank deuterated “heavy” water (2H2O). At day 13, peripheral blood (PB), lymph node (LN), and bone marrow (BM) samples were collected from each patient, and cells from these 3 sites were analyzed for 2H-labeled CD5+CD19+ cells after FACS sorting into 6 fractions: DIM, dDIM, INT, INT/BR, BR and dBR. Overall, more CLL B-cell proliferation was found in LN than PB and BM; only small numbers of divided cells were found in BM. Interestingly, the DIM, dDIM, and INT/BR had the highest 2H-incorporation in LN and PB; and DIM cells from LN and PB showed similar levels of 2H-incorporation.Gene expression profiling using Illumina Human HT12 BeadChips was then performed on the same 6 fractions from PB and LN. Expression value ratios for fractions from each patient were analyzed using R, and sets of significant genes (fold change >1.5 and P<0.05) were determined. Unsupervised hierarchical clustering grouped together the DIM and dDIM fractions from all other fractions. Although the INT/BR had high 2H-incorporation levels, it clustered with the BR, INT and dBR fractions. Genes upregulated in DIM and dDIM included genes involved in cell proliferation and survival, such as DUSP1, PRKCD and BMF, and chemokine genes CCL3, CCL3L1, CCL3L3, CCL4L2. Genes changed in BR, dBR, and INT/BR included negative regulators of cell survival and proliferation, e.g. PRICKLE1 and GRAP.Finally, functional differences in the CLL fractions were analyzed in vivo using NOD/SCID/γcnull (NSG) mice. 3-5x106 PB B cells were injected with 1-1.5x105 resting, autologous T cells. Compared to BR, dBR and INT/BR fractions, DIM and dDIM cells led to more extensive T-cell growth and B-cell expansion in the spleen and BM at week 6. In contrast, INT injected mice exhibited only minimal or no CLL B or T cells. Adding INT cells suppressed DIM and dDIM induced T-cell expansion. This function, which was resistant to irradiation, was associated with greater immunological synapse impairment in vitro.Thus, in vivo kinetics analysis using D2O identified levels of divided cells based on the following ranking: DIM ≥ INT/BR> dDIM > INT> BR=dBR. Although the INT/BR fraction contains recently-divided cells based on 2H-labeling, its genetic signature is similar to fractions containing lesser numbers of recently activated cells, suggesting INT/BR cells have divided but are being inactivated in the microenvironment. Therefore, these cells may represent a transition population between proliferating and resting cells. Because PB and LN DIM fractions are very similar in proliferation rates and gene expression profiles and because little CLL B-cell proliferation occurs in the blood, the circulating DIM fraction accurate reflects LN proliferation. When combining kinetics and gene expression profiling, the DIM and dDIM fractions appear as the most activated intraclonal fractions. In xenografted mice, PB DIM and dDIM cells are better at activating T cells and consequently receiving T-cell help for engraftment and growth. The INT fraction, which constitutes the tumor bulk, inhibits T-cell activation. Finally, the BR and dBR fraction are the older, most quiescent cells in CLL clones. Overall, these results inform about trafficking and transition of CLL cells between lymphoid tissues and periphery, and provide a rationale for preferential therapeutic targeting of these fractions. Disclosures:Riches:Celgene: Research Funding.

Decreased Proportion of Cytomegalovirus Specific CD8 T-Cells but No Signs of General Immunosenescence in Alzheimer’s Disease

Cytomegalovirus (CMV) has been suggested as a contributing force behind the impaired immune responsiveness in the elderly, with decreased numbers of naïve T-cells and an increased proportion of effector T-cells. Immunological impairment is also implicated as a part of the pathogenesis in Alzheimer’s disease (AD). The aim of this study was to investigate whether AD patients present with a different CMV-specific CD8 immune profile compared to non-demented controls. Blood samples from 50 AD patients and 50 age-matched controls were analysed for HLA-type, CMV serostatus and systemic inflammatory biomarkers. Using multi-colour flow cytometry, lymphocytes from peripheral blood mononuclear cells were analysed for CMV-specific CD8 immunity with MHC-I tetramers A01, A02, A24, B07, B08 and B35 and further classified using CD27, CD28, CD45RA and CCR7 antibodies. Among CMV seropositive subjects, patients with AD had significantly lower proportions of CMV-specific CD8 T-cells compared to controls, 1.16 % vs. 4.13 % (p=0.0057). Regardless of dementia status, CMV seropositive subjects presented with a lower proportion of naïve CD8 cells and a higher proportion of effector CD8 cells compared to seronegative subjects. Interestingly, patients with AD showed a decreased proportion of CMV-specific CD8 cells but no difference in general CD8 differentiation.

Associated Cancers in Waldenström Macroglobulinemia: Clues for Common Genetic Predisposition

Several population-based and cohort studies have reported an increased risk of second cancers in lymphoproliferative disorders (LPDs). The cause of second cancers in LPDs is probably multifactorial, and the relative contribution of treatments, genetic predisposition, and immune dysfunction typical of LPDs is still unclear. We retrospectively studied 230 patients with Waldenström macroglobulinemia (WM) to assess the frequency, characteristics, and predictive factors of second cancers and to evaluate whether patients with WM are at higher risk of second cancers compared with an age- and sex-matched control population. In a competing-risk model, the cumulative incidence of solid cancers was 6% at 5 years, 11% at 10 years, and 17% at 15 years, whereas the incidence of hematologic malignancies was 4% at 5 years, 7% at 10 years, and 8% at 15 years. Compared with an age- and sex-matched population, the overall risk of second cancers was 1.7-fold higher than expected (95% confidence interval [CI], 1.22-2.38; P= .002). Patients with WM were at increased risk for diffuse large B-cell lymphoma (DLBCL) (standardized incidence ratio [SIR], 8.64; 95% CI, 3.88-19.22; P< .0001), myelodysplastic syndrome/acute myeloid leukemia (MDS/AML) (SIR 9.5; 95% CI, 3.6-25.3; P< .0001), and brain cancer (SIR, 7.59; 95% CI, 1.9-30.4; P< .0001). The risk of a second hematologic malignancy was 5-fold higher in treated than in untreated patients (P= .08). Patients with WM are at increased risk of DLBCL, MDS/AML, and brain cancers compared with the general population. Further studies are needed to clarify whether the increased incidence of second cancers is related to treatments, to the immunologic impairment associated with the disease, or to genetic predisposition.

Diabetic foot infections: state‐of‐the‐art

Foot infections are frequent and potentially devastating complications of diabetes. Unchecked, infection can progress contiguously to involve the deeper soft tissues and ultimately the bone. Foot ulcers in people with diabetes are most often the consequence of one or more of the following: peripheral sensory neuropathy, motor neuropathy and gait disorders, peripheral arterial insufficiency or immunological impairments. Infection develops in over half of foot ulcers and is the factor that most often leads to lower extremity amputation. These amputations are associated with substantial morbidity, reduced quality of life and major financial costs. Most infections can be successfully treated with optimal wound care, antibiotic therapy and surgical procedures. Employing evidence-based guidelines, multidisciplinary teams and institution-specific clinical pathways provides the best approach to guide clinicians through this multifaceted problem. All clinicians regularly seeing people with diabetes should have an understanding of how to prevent, diagnose and treat foot infections, which requires familiarity with the pathophysiology of the problem and the literature supporting currently recommended care.

Immunogenicity of Licensed Influenza A (H1N1) 2009 Monovalent Vaccines in HIV-Infected Children and Youth

With the emergence of pandemic influenza A (pH1N1) in 2009, children and youth infected with human immunodeficiency virus (HIV) were vulnerable because of immunologic impairment and the greater virulence of this infection in young persons. A multicenter study of the immunogenicity of 3 licensed influenza A (H1N1) monovalent vaccines (1 live attenuated and 2 inactivated) was conducted in children and youth with perinatal HIV infection, most of whom were receiving ≥3 antiretroviral drugs, had CD4% ≥15, and plasma HIV RNA levels <400 copies/mL. Serum hemagglutinin inhibition assay (HAI) antibody levels were measured and correlated with baseline demographic and clinical variables. One hundred forty-nine subjects were enrolled at 26 sites in the United States and Puerto Rico. Over 40% had baseline HAI titers ≥40. For subjects aged 6 months to <10 years, 79% and 68%, respectively, achieved a ≥40- and ≥4-fold rise in HAI titers after the second dose of vaccine. Three weeks after a single immunization with an inactivated vaccine, similar immunogenicity results were achieved in youth aged 10-24 years. With multivariable analysis, only Hispanic ethnicity and CD4% ≥15 were associated with achieving both HAI titer ≥40- and ≥4-fold rise in titer. Although licensed pH1N1 vaccines produced HAI titers that were considered to be protective in the majority of HIV-infected children and youth, the proportion with titers ≥40- and ≥4-fold rise in titer was lower than expected for children without HIV infection. Vaccine immunogenicity was lower in HIV-infected children and youth with evidence of immune suppression.

HBV and HCV seroprevalence and their correlation with CD4 cells and liver enzymes among HIV positive individuals at University of Gondar Teaching Hospital, Northwest Ethiopia

BackgroundThe co-existence of viral hepatitis caused by HBV and HCV become common causes of severe liver complication and immunological impairment among HIV infected individuals. The aim of this study was to assess the seroprevalence of HBV and HCV and their correlation with CD4 and liver enzyme levels among HAART naïve HIV positive individuals.MethodA Cross-sectional study was conducted from March-May, 2011 at University of Gondar Teaching Hospital, Northwest Ethiopia. HBV and HCV serological tests and liver enzymes as well as CD4 T cell level determination were assessed following the standard procedures. Socio-demographic data was collected by using structured questionnaire. The data was entered and analyzed by using SPSS version 20.0 statistical software and p < 0.05 was considered as statistically significant.ResultAmong 400 study participants, the overall prevalence of HIV-viral hepatitis co-infection was 42(11.7%). The prevalence of HIV-HBV, HIV-HCV and HIV-HBV-HCV co-infections were 20(5.6%), 18(5.0%) and 4(1.1%) respectively. Study participants who had HIV-HBV, HIV-HCV and HIV-HBV-HCV co-infection have relatively raised mean liver enzyme levels (ALT, AST and ALP) than HIV mono-infected once. Individuals with HIV-HBV, HIV-HCV and HIV-HBV-HCV co-infection also had a lower mean CD4 levels than HIV mono-infected study participants. The mean CD4 value in males was lower than females.ConclusionThe prevalence of HBV and HCV was higher than reports from general population of the country. Raised levels of liver enzymes and lowered mean CD4 counts were seen in HIV-HBV, HIV-HCV and HIV-HBV-HCV co-infections. These findings underscore the importance of screening all HIV positive individuals before initiating antiretroviral treatment.

The natural history of HIV infection

Purpose of reviewTo review recent published literature around three areas: long-term nonprogression/viral control; predictors of viral load set point/disease progression; and the potential impact of antiretroviral therapy (ART) in early HIV infection.Recent findingsThe natural course of untreated HIV infection varies widely with some HIV-positive individuals able to maintain high CD4 cell counts and/or suppressed viral load in the absence of ART. Although similar, the underlying mechanistic processes leading to long-term nonprogression and viral control are likely to differ. Concerted ongoing research efforts will hopefully identify host factors that are causally related to these phenotypes, thus providing opportunities for the development of novel treatment or preventive strategies. Although there is increasing evidence that initiation of ART during primary infection may prevent the immunological deterioration which would otherwise be seen in untreated HIV infection, recent studies do not address the longer term clinical benefits of ART at this very early stage.SummaryA better understanding of the relative influences of viral, host, and environmental factors on the natural course of HIV infection has the potential to identify novel targets for intervention to prevent and treat HIV-infected persons.