Potent antiviral activity of amprenavir in primary macrophages infected by human immunodeficiency virus

Objective of the present study was then to assess the antiviral activity of the protease inhibitor amprenavir in macrophages (M/M), and to compare it with its efficacy in peripheral blood lymphocytes (PBL). M/M were obtained from blood of sero-negative healthy donors and infected with M-tropic HIV-1 strain (HIV-1(Ba-L)). The stabilized infection was assessed by monitoring the HIV-1 p24 gag antigen production in the supernatants of M/M cultures. In the setting of acute infection (treatment before HIV-1 challenge), amprenavir showed substantial activity both in M/M and PBL at similar concentrations (EC(50): 0.011 and 0.031 microM, respectively); complete inhibition of HIV-1 replication was achieved in both cell types at concentration of about 2 microM. In the setting of chronical infection (i.e. antiviral treatment several days after established infection), an antiviral effect of amprenavir was achieved in M/M, but at concentrations higher than those active in acutely infected M/M (EC(50): 0.72 microM, EC(90): 18.2 microM). The antiviral effect in chronically infected M/M was sustained for at least 2 weeks of continuous treatment. These findings suggest that amprenavir (at relatively high concentrations) has a clinically relevant antiviral effect in persistently infected reservoirs of HIV.     

Macrophage depletion induced by clodronate-loaded erythrocytes

Given the important role of macrophages in various disorders, the transient and organ specific suppression of their functions may benefit some patients. Until now, liposome-encapsulated bisphosphonate clodronate has been extensively proposed to this end. In this paper, we demonstrate that erythrocytes loaded with clodronate can also be effective in macrophage depletion. Here, clodronate was encapsulated in erythrocytes through hypotonic dialysis, isotonic resealing and reannealing to final concentrations of 4.1 +/- 0.4 and 10.1 +/- 0.8 micromol/ml of human and murine erythrocytes, respectively. The ability of clodronate-loaded erythrocytes to deplete macrophages was evaluated both in vitro and in vivo. In vitro studies on human macrophages showed that a single administration of engineered erythrocytes was able to reduce cell adherence capacity in a time-dependent manner, reaching 50 +/- 4% reduction, 13 days post treatment. The administration of loaded erythrocytes to cultures of murine peritoneal macrophages was able to reduce macrophage adhesion 67 +/- 3%, 48 h post treatment. In vivo, the ability of clodronate-loaded erythrocytes to deplete macrophages was evaluated both in Swiss and C57BL/6 mice. Swiss mice received 125 microg of clodronate through erythrocytes and 6 days post treatment 69 +/- 7% reduction in the number of adherent peritoneal macrophages and 75 +/- 5% reduction in number of spleen macrophages were observed. C57BL/6 mice received 220 microg clodronate by RBC and 3 and 8 days post treatment 65 +/- 7% reduction in the number of spleen macrophages and the complete depletion of liver macrophages were obtained. In summary, our results indicate that clodronate selectively targeted to the phagocytic cells by a single administration of engineered erythrocytes is able to deplete macrophages, even if not completely. The transient suppression of macrophage functions through clodronate-loaded erythrocytes can be used in many biomedical phenomena and research applications.     

The role of white cells in the transmission of Yersinia enterocolitica in blood components

The mechanism for the transmission of Yersinia enterocolitica in blood components has been studied experimentally. One hypothesis is that, during a Yersinia infection in the blood donor, bacteria are phagocytosed by white cells (WBCs), but are not killed. After collection of blood from such a donor and component production, the bacteria are present in WBCs for some time, during which the unit appears sterile. Later, when the WBCs disintegrate, the bacteria are released and multiply in the unit. Aliquots of whole blood and buffy coat were inoculated with 100 colony-forming units (CFU) per mL of a Y. enterocolitica strain of type O:3 and left at room temperature for 5 hours. Some aliquots were then WBC-reduced by filtration, while others retained their WBC contents. All aliquots were kept at 4 degrees C for 6 weeks. Meat extract broth culture medium was used as a control. Growth in the range of 2000 CFU per mL was obtained in the broth control by 24 hours, whereas the whole blood and buffy coat units appeared sterile for the first days of storage. After 1 week, a trace of bacteria and, after 4 weeks, massive growth were found in the WBC-containing units but not in the WBC-reduced units. The likely explanation is that the bacteria had been phagocytosed by the WBCs and were thereby hidden and not available for bacterial culture during the first phase of storage. When the WBCs spontaneously disintegrated, bacteria were released and multiplied in the blood units.(ABSTRACT TRUNCATED AT 250 WORDS)     

Augmentation of spontaneous macrophage-mediated cytolysis by eosinophil peroxidase

Eosinophil peroxidase (EPO), a cationic protein purified from horse blood, adhered to four different types of tumor cells, markedly potentiating their lysis by preformed or enzymatically generated H(2)0(2) (up to 76-fold, as assayed in serum-containing tissue culture medium without supplemental halide). Similarly, compared with uncoated tumor cells, EPO-coated tumor cells were up to 32 times more sensitive to lysis when incubated with macrophages or granulocytes whose respiratory burst was triggered by PMA. However, EPO-coated tumor cells were also readily lysed by bacillus Calmette- Guerin-activated macrophages in the absence of exogenous triggering agents. This spontaneous cytolysis was rapid (50 percent at 2 h) and potent (50 percent lysis at macrophage/tumor cell ratios of 1.5 to 4.6), and was observed with both a peroxide-sensitive tumor (TLX9) and a peroxide-resistant tumor (NK lymphoma). Under the conditions used, neither EPO alone nor macrophages alone were spontaneously cytolytic. Neither EPO nor EPO-coated tumor cells triggered a detectable increment in H(2)0(2) release from macrophages. Nonetheless, spontaneous macrophage-mediated cytolysis of EPO- coated tumor cells was completely inhibitable by catalase (50 percent inhibition, 23 U/ml), although not by heated catalase, indicating a requirement for H(2)0(2). Cytolysis was also completely inhibitable by azide (50 percent inhibition, 2.6 X 10 -5 M), indicating a requirement for enzymatic activity of EPO. Thus, a cytophilic peroxidase from eosinophils and H(2)0(2) spontaneously released from activated macrophages interacted synergistically in a physiologic medium to destroy tumor cells.     

Continuous evidence of fast HIV disease progression related to class-wide resistance to antiretroviral drugs: a 6 year follow-up analysis of a large observational database

Class-wide resistance (CWR) was increasingly associated with a higher risk of HIV progression after 72 months of follow-up among 1392 patients genotypic-tested after failure (AIDS risk 13% for no CWR to 34% for three CWR; AIDS/death risk 21-54%). At multivariate analysis, the detection of two and three CWR was significantly associated with a two and threefold increased risk, respectively, of death and AIDS/death, suggesting that extended resistance is a marker of disease progression in long-term observation.     

Origin of the 2009 Mexico influenza virus: a comparative phylogenetic analysis of the principal external antigens and matrix protein

Triple-reassortant swine influenza A (H1) viruses, containing genes from avian, human, and swine influenza viruses, emerged and became an outbreak among humans worldwide. Over a 1,000 cases were identified within the first month, chiefly in Mexico and the United States. Here, the phylogenetic analysis of haemagglutin (HA), neuraminidase (NA), and matrix protein (MP) was carried out. The analysis showed that the H1 of this reassortant originated from American pigs, while NA and MP were more likely from European pigs. All of the 2009 isolates appear homogeneous and cluster together, although they are distinct from classical human A (H1N1) viruses. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Providing monovalent oral polio vaccine type 1 to newborns: findings from a pilot birth-dose project in Moradabad district, India

Problem

Poliovirus transmission remained a public health challenge in western Uttar Pradesh, India in late 2005 and early 2006. In 2006, the India Expert Advisory Group for Polio Eradication concluded that, given the peak incidence of polio among children 6 to 12 months of age, a targeted birth dose of oral polio vaccine may be necessary to interrupt intense poliovirus transmission in high risk areas.

Approach

The Government of Uttar Pradesh, the National Polio Surveillance Project and the United Nations Children’s Fund (UNICEF) implemented a pilot birth-dose project aimed at identifying and vaccinating all newborns with a dose of oral polio vaccine within 72 hours of birth in an effort to evaluate operational feasibility and potential impact on population immunity.

Local setting

The project was piloted in Moradabad district: zone 7 in Moradabad City (urban setting), Kunderki block (rural setting) and in select birthing hospitals.

Relevant changes

Between July 2006 and February 2007, 9740 newborns were identified, of which 6369 (65%) were vaccinated by project personnel within 72 hours of birth. Project coverage (for total newborns vaccinated) ranged from 39% (in zone 7) to 76% (in Kunderki block) of the estimated number of newborns vaccinated during previous supplemental immunization activities.

Lessons learned

Birth-dose coverage among newborns was lower than expected. Expansion costs were estimated to be high, with marginal impact. The project, however, provided opportunities to strengthen newborn tracking systems which have increased the number of newborns and young infants vaccinated during supplemental immunization activities and enrolled in routine programmes.