Drug-induced death of the asexual blood stages of Plasmodium falciparum occurs without typical signs of apoptosis

There is clear evidence that most antimalarial drugs, while acting through different mechanisms, are associated with parasite growth/development inhibition and eventual parasite death. However, the exact mode of parasite death remains unclear. In the present study, we investigated the ability of various drugs, including two antimalarial drugs (chloroquine and atovaquone), a topoisemerase II inhibitor (etoposide) and a nitric oxide donor (S-nitro-N-acetyl-D, L-penicillamine), to induce apoptosis in a laboratory strain of Plasmodium falciparum. Results obtained from flow cytometric analysis showed a significant reduction in the percent of parasitemia and parasite growth in all drug-treated parasite cultures, including those treated with etoposide and S-nitro-N-acetyl-D, L-penicillamine. For further investigation, we used various biochemical approaches including the terminal dUTP nick-end labeling assay, determination of mitochondrial membrane integrity and DNA degradation/fragmentation, to analyze the changes occurring during parasite-drug interactions and eventual death. We observed that loss of membrane potential was induced in parasite cultures treated with atovaquone, while S-nitro-N-acetyl-D, L-penicillamine induced abnormal parasite forms, "crisis forms", and minor DNA degradation. However, these features were not observed in the parasite cultures treated with chloroquine nor were other features of apoptosis-like death associated with any of the drugs used in this study. The death resulting from the various drug treatments is atypical of apotosis. More studies will be needed to define the precise mode of death exhibited by P. falciparum.     

Evaluation of Electronic Medical Record (EMR) at Large Urban Primary Care Sexual Health Centre

Objective

Despite substantial investment in Electronic Medical Record (EMR) systems there has been little research to evaluate them. Our aim was to evaluate changes in efficiency and quality of services after the introduction of a purpose built EMR system, and to assess its acceptability by the doctors, nurses and patients using it.

Methods

We compared a nine month period before and after the introduction of an EMR system in a large sexual health service, audited a sample of records in both periods and undertook anonymous surveys of both staff and patients.

Results

There were 9,752 doctor consultations (in 5,512 consulting hours) in the Paper Medical Record (PMR) period and 9,145 doctor consultations (in 5,176 consulting hours in the EMR period eligible for inclusion in the analysis. There were 5% more consultations per hour seen by doctors in the EMR period compared to the PMR period (rate ratio = 1.05; 95% confidence interval, 1.02, 1.08) after adjusting for type of consultation. The qualitative evaluation of 300 records for each period showed no difference in quality (P>0.17). A survey of clinicians demonstrated that doctors and nurses preferred the EMR system (P<0.01) and a patient survey in each period showed no difference in satisfaction of their care (97% for PMR, 95% for EMR, P = 0.61).

Conclusion

The introduction of an integrated EMR improved efficiency while maintaining the quality of the patient record. The EMR was popular with staff and was not associated with a decline in patient satisfaction in the clinical care provided.     

Culture-expanded human dermal stem cells exhibit donor to donor differences in cAMP generation

Stem cell techniques have facilitated a number of potential uses for such cells in cell therapy and drug development. Studies of the cAMP/protein kinase A (PKA) pathway are widely employed to investigate the effects of a large variety of substances. We assayed the cAMP pathway in human skin-derived mesenchymal stem cells (S-MSC) to evaluate donor to donor variations in response to pharmacological manipulations in vitro. Immunophenotyping of S-MSC revealed that, in general, 95% of S-MSCs were positive for CD90, CD73 and CD105 and negative for the expression of haemopoetic markers CD14, CD45 and human leukocyte antigen-DR (HLA-DR). Nevertheless, fluctuations occurred in basal cAMP levels from 5 pmol/mg to 18 pmol/mg. Total cAMP response element binding protein (CREB) concentrations ranged from 0.8 ng/ml to 1 ng/ml, whereas the proportions of phospho-CREB versus total CREB differed between the cell lines. Basic fibroblast growth factor (FGF-2) and epidermal growth factor (EGF) stimulated cAMP generation, whereas leukaemia inhibiting factor reduced some of their effects. Forskolin (0.05 and 1 mM) acted in synergy with FGF-2 and EGF; however, it caused pronounced donor to donor differences in the increase of cAMP and phospho-CREB levels. Additionally, dibutyryl-cAMP caused significant donor to donor variations in cell proliferation, possibly indicating a change of cell differentiation status. We speculate that similar donor diversity might be observed after cell stimulation with various G_s-protein-coupled receptor ligands. Heterogeneity of donor cell responses to stimulation of the cAMP pathway indicates the need for wide safety margins for S-MSC use in drug screening; nevertheless, knowledge of this heterogeneity might be useful for the design of donor-specific cell therapy.     

HIV-1 Specific Antibody Titers and Neutralization among Chronically Infected Patients on Long-Term Suppressive Antiretroviral Therapy (ART): A Cross-Sectional Study

The majority of potent and broadly neutralizing antibodies against HIV-1 have been isolated from untreated patients with acute or chronic infection. To assess the extent of HIV-1 specific antibody response and neutralization after many years of virologic suppression from potent combination ART, we examined antibody binding titers and neutralization of 51 patients with chronic HIV-1 infection on suppressive ART for at least three years. In this cross-sectional analysis, we found high antibody titers against gp120, gp41, and the membrane proximal external region (MPER) in 59%, 43%, and 27% of patients, respectively. We observed significantly higher endpoint binding titers for gp120 and gp41 for patients with >10 compared to ≤10 years of detectable HIV RNA. Additionally, we observed higher median gp120 and gp41 antibody titers in patients with HIV RNA <50 copies/mL for ≤5 years. 22% of patients neutralized a HIV-1 primary isolate (HIV-1_JR-FL) and 8% neutralized a HIV-2/HIV-1 MPER chimera. Significantly greater HIV-1_JR-FL neutralization was found among patients with >10 years of detectable HIV RNA (8/20 [40.0%] versus 3/31 [9.7%] for ≤10 years, p = 0.02) and a trend toward greater neutralization in patients with ≤5 years of HIV RNA <50 copies/mL (7/20 [35.0%] versus 4/31 [12.9%] for >5 years, p = 0.08). All patients with neutralizing activity mediated successful phagocytosis of VLPs by THP-1 cells after antibody opsonization. Our findings of highly specific antibodies to several structural epitopes of HIV-1 with antibody effector functions and neutralizing activity after long-term suppressive ART, suggest continuous antigenic stimulation and evolution of HIV-specific antibody response occurs before and after suppression with ART. These patients, particularly those with slower HIV progression and more time with detectable viremia prior to initiation of suppressive ART, are a promising population to identify and further study functional antibodies against HIV-1.     

Sex-specific effects of increased incubation demand on innate immunity in black guillemots

Life-history theory predicts that there should be negative fitness consequences, in terms of future reproduction and survival, for parents with increased reproductive effort. We examined whether increased incubation demand affected innate immunity and body condition by performing a clutch-size manipulation experiment in black guillemots (Cepphus grylle). We found that plasma from males incubating experimentally enlarged clutches exhibited significantly reduced lysis titers compared with plasma from males incubating control clutches, while this was not observed in females. The increased incubation demand also impacted agglutination titers differently in males and females, although the effect of treatment was not significant in either sex. Among all birds, lysis titers increased and haptoglobin concentrations decreased from mid- to late incubation. Natural antibody-mediated agglutination titers and body condition were highly repeatable within the incubation bout and between years. This suggests that agglutination titers may serve as a reliable and resilient index of the immunological character of individuals in future studies. Overall, this study demonstrates that increased incubation demand impacts indices of innate immunity differently in males and females. The potential for different components of the immune system to be impacted sex-specifically should be considered in future studies linking immune function and life-history trade-offs.     

Mitochondrial DNA origins of the Latvian clefting population

Latvia has one of the highest prevalence of isolated cleft lip with or without cleft palate (CL/P) in Europe. To clarify the genetic origins of the Latvian cleft population and establish a method for genetic mapping, mitochondrial DNA variation was studied in a population affected with clefting. One-hundred and seven subjects and 351 samples from unrelated healthy volunteers representing four anthropologically, archaeologically and ethno-linguistically different regions of Latvia were selected. The case group showed a higher frequency of haplogroups U4 (p=0.02) and U5 (p=0.0003) than in non-U haplogroups. We hypothesize that U4 and U5 mtDNA haplotype carriers may also carry susceptibility genes for clefts. Future studies will take into consideration these definitions based on mtDNA haplotypes when analyzing genetic variations and their possible contribution to CL/P susceptibility.     

Sex differences in the pattern of esterase-active antigens in microsomes from various rat strains

Esterase-active antigens present in male and female liver microsomes isolated from three different rat strains (Sprague-Dawley, Wistar and Dark Agouti) were characterized in crossed immunoelectrophoresis in combination with a zymogram method for esterase activity. No qualitative but some quantitative differencies were encountered between the sexes. Thus, two out of ten antigens were present in significantly lower and one in significantly higher concentrations in male than in female microsomes, demonstrating that although the overall esterase activity in liver may be similar for males and females, the concentration of the individual antigens does vary between the sexes. No qualitative or quantitative differences in the pattern of esterase-active antigens were found between the different strains.