Decline of FoxP3+ Regulatory CD4 T Cells in Peripheral Blood of Children Heavily Exposed to Malaria

FoxP3+ regulatory CD4 T cells (T_regs) help to maintain the delicate balance between pathogen-specific immunity and immune-mediated pathology. Prior studies suggest that T_regs are induced by P. falciparum both in vivo and in vitro; however, the factors influencing T_reg homeostasis during acute and chronic infections, and their role in malaria immunopathogenesis, remain unclear. We assessed the frequency and phenotype of T_regs in well-characterized cohorts of children residing in a region of high malaria endemicity in Uganda. We found that both the frequency and absolute numbers of FoxP3+ T_regs in peripheral blood declined markedly with increasing prior malaria incidence. Longitudinal measurements confirmed that this decline occurred only among highly malaria-exposed children. The decline of T_regs from peripheral blood was accompanied by reduced in vitro induction of T_regs by parasite antigen and decreased expression of TNFR2 on T_regs among children who had intense prior exposure to malaria. While T_reg frequencies were not associated with protection from malaria, there was a trend toward reduced risk of symptomatic malaria once infected with P. falciparum among children with lower T_reg frequencies. These data demonstrate that chronic malaria exposure results in altered T_reg homeostasis, which may impact the development of antimalarial immunity in naturally exposed populations.     

Increased incidence of pertussis and parapertussis in HIV-1-positive adolescents vaccinated previously with whole-cell pertussis vaccine

We investigated 296 adolescents (11–18 years), who had been immunized previously with the three doses of DPT vaccines. 48 were diagnosed positive for HIV-1. Nasopharyngeal swabs were obtained from 296 adolescents who presented with persistent cough and nasopharyngeal secretions. Nasopharyngeal swabs (calcium alginate) specimens were collected by passing the swabs through the nares into the posterior nasopharynx and rotating the swabs for a few seconds. The swabs were plated for culture of Bordetella organisms in charcoal cephalexin blood agar (CCBA). The CCBA plates were incubated for 2–6 days at 35 °C in a humid aerobic atmosphere. The suspected, shiny (mercury-like) colonies were tested by slide agglutination with antisera to B. pertussis and B. parapertussis, and urease, oxidase activities were performed. Results indicate that out of 48 HIV-1-positive adolescents, 18 had positive cultures for Bordetella organisms (14, Bordetella pertussis, and 4, Bordetella parapertussis). Of 248 HIV-1-negative subjects, 3 had Bordetella organisms (2, Bordetella pertussis, 1, Bordetella bronchiseptica). One of the subjects, a boy, aged 14 years, with Bordetella bronchiseptica had a dog as pet, which was found to be infected. The results indicate that adolescents with HIV-1 infection, despite being vaccinated against pertussis have a higher rate of infection when exposed to pertussis bacteria than HIV-1-negative adolescents. This revised version was published online in August 2006 with corrections to the Cover Date.     

Synthesis of (aminoalkyl)cycleanine analogues: cytotoxicity,cellular uptake,and apoptosis induction in ovarian cancer cells

Our previous studies demonstrated that cycleanine, a macrocyclic bisbenzylisoquinoline (BBIQ) alkaloid, showed potent anti-ovarian cancer activity via apoptosis induction. Here, we synthesized two novel (aminoalkyl)cycleanine analogues (2 and 3) through a simple and efficient two-step reaction starting from cycleanine isolated from Triclisia subcordata Oliv. These analogues showed greater potency than the unmodified cycleanine in three human ovarian cancer cell lines. Both 2 and 3 induced apoptosis in ovarian cancer cells by activations of caspases 3/7, cleavage of PARP, increase in subG₁ cell cycle phase and in the percentage of apoptotic cells. Further confocal fluorescence microscopy analysis confirmed the cellular uptake of alkaloids in ovarian cancer cells by using the unique (alkynyl)cycleanine (3) via click chemistry reaction. Our results suggest that cycleanine could be a hit compound for the future development in attacking ovarian cancer.     

Sensitivity to antimicrobial agents of some mercury-sensitive and mercury-resistant strains of Gram-negative bacteria

The sensitivity and resistance of some Gram-negative mercury (Hg²⁺)-sensitive and-resistant strains to chemotherapeutic agents and to disinfectants and preservatives are described.Escherichia coli andPseudomonas aeruginosa strains harboring plasmid pUB 1351 [pUB 367:Tn 501] andE. coli bearing R100-1 were resistant to inorganic mercury and to various antibiotics, but were not more resistant to organic mercury and other preservatives and disinfectants than plasmidless strains.     

Response of RP1+ and RP1− strains ofEscherichia coli to antibacterial agents and transfer of resistance toPseudomonas aeruginosa

The sensitivity of strains ofEscherichia coli, with and without the RP1 R-factor, to antibiotics and other antibacterial agents has been studied. RP1⁺ strains ofE. coli were resistant to kanamycin, carbenicillin, and tetracycline, resistance to the first two antibiotics being produced by destruction of the drugs. This resistance could be transferred to two strains ofPseudomonas aeruginosa. The parent strain ofE. coli UB 1005, its two mutant strains (DC2 and DC3), and two of the strains with the RP1 R-factor showed a similar order of sensitivity to phenylmercuric nitrate, chlorhexidine, thiomersal, and mercuric chloride.E. coli strains DC2 and DC2 (RP1⁺) were the most sensitive to benzalkonium chloride and cetrimide. RP1⁺ strains were more resistant than RP1⁻ strains to lysozyme-ethylenediaminetetraacetic acid, but treatment of the former strains with acriflavine rendered the cells more sensitive to the lytic system. There was no evidence thatP. aeruginosa (RP1⁺) strains possessed increased resistance to polymyxin or to disinfectants, although they became somewhat less sensitive to lysozyme-ethylenediaminetetraacetic acid.     

Carbohydrate-based micelle clusters which enhance hydrophobic drug bioavailability by up to 1 order of magnitude

Amphiphilic chitosan-based polymers (Mw < 20 kDa) self-assemble in aqueous media at low micromolar concentrations to give previously unknown micellar clusters of 100-300 nm in size. Micellar clusters comprise smaller 10-30 nm aggregates, and the nanopolarity/drug incorporation efficiency of their hydrophobic domains can be tailored by varying the degree of lipidic derivatization and molecular weight of the carbohydrate. The extent of drug incorporation by these novel micellar clusters is 1 order of magnitude higher than is seen with triblock copolymers, with molar polymer/drug ratios of 1:48 to 1:67. On intravenous injection, the pharmacodynamic activity of a carbohydrate propofol formulation is increased by 1 order of magnitude when compared to a commercial emulsion formulation, and on topical ocular application of a carbohydrate prednisolone formulation, initial drug aqueous humor levels are similar to those found with a 10-fold dose of prednisolone suspension.     

Assessment of Lactobacillus species colonizing the vagina of apparently healthy Nigerian women, using PCR-DGGE and 16S rRNA gene sequencing

The identity of lactobacilli colonizing Africa-American women, have previously been based on culture-dependent methods. This led to some misleading speculations that black women lack lactobacilli in their vagina and are therefore highly susceptible to Bacterial vaginosis and STDs including HIV. In this study we used culture-independent procedures.About 241 vaginal swabs were obtained from ‘apparently’ healthy premenopausal women, between 18 and 48 years. Samples were Gram stained for the Nugent score evaluation. DNA was extracted from the bacteria on the vaginal swabs and amplified using polymerase chain reaction (PCR), with Lactobacillus primers. Samples with PCR products were separated with denaturing gradient gel electrophoresis (DGGE) and cut bands were re-amplified with Lactobacillus primers without the GC clamp. The re-amplified product was purified and sequenced with ABI PRISM Big-Dye Terminator. The sequence was identified with BLAST algorithm having the highest Gene Bank Accession number.Out of the 241 vaginal swab samples, 207 (85.8%) had PCR products, indicating the presence of lactobacilli, while 34 (14.2%) showed absence of lactobacilli and the Nugent scores were synonymous to either intermediate bacterial vaginosis or bacterial vaginosis (BV). Out of the 207 samples that had PCR product for Lactobacillus, 149 (72%) had sequence results as revealed by the BLAST algorithm. Most of the women (64%) were colonized by Lactobacillus iners as the predominant strain. Lactobacillus gasseri had 7.3%, followed by L. plantarum and L suntoryeus (6.0%) each. Others were colonized by Lactobacillus crispatus (3.0%), Lactobacillus rhamnosus (2.7%), Lactobacillus vaginalis (2.7%), Lactobacillus rennanqilfy (2.7%), followed by Lactobacillus fermentum (1.3%), Lactobacillus helveticus (1.3%), Lactobacillus johnsonii (1.3%) and Lactobacillus salivarus (1.3%). To our knowledge, this is the first report of Lactobacillus iners being the predominant species colonizing the vagina of the Nigerian women examined. This finding is in line with those from a recent study conducted among Canadian and Swedish women.     

Perception of donor breast milk and determinants of its acceptability among mothers in a developing community: a cross-sectional multi-center study in south-east Nigeria

Background

Due to the health and economic benefits of breast milk, the World Health Organization (WHO) recommends that for infants who cannot receive breast milk from their own mothers, the next preferred option is donated breast milk. This recommendation is however rarely practiced in most developing countries where donor milk is not widely accepted.

Methods

This cross-sectional multi-center study enrolled mothers attending antenatal or pediatric clinics in six tertiary institution in south-east Nigeria using purposive and convenient sampling method. Data collection was done using pretested questionnaires. The study aimed to assess the knowledge, acceptability and willingness to donate breast milk and/or use donated breast milk for their infants It also explored factors that determine this behavior.

Results

A total of 1235 mothers participated; 39% (480/1225) have heard about the concept of donor milk, while only 10% (79/759) and 7% (81/1179), respectively, had adequate knowledge of the concept and policy on donor milk. Sixty percent indicated willingness to use donor milk or donate breast milk if need arises. Respondents with lower age (p?=?0.049) and with higher occupational status (p?=?0.001) were more likely to have adequate knowledge of donor breast milk, while respondents with lower educational attainment (p?=?0.002) and those who are non-Christians (p?=?0.004) were more likely to request financial inducement for donating their breast milk. Adequate knowledge of the concept of donor milk (p?=?0.001), preference of donor milk to infant formula (p?=?0.001) and requirement of financial remuneration (p?=?0.001) were the only significant predictors of willingness to donate and/or receive donated breast milk.

Conclusion

The knowledge of the concept of donor breast milk and awareness of policies regulating its practice in Nigeria is low, but the prospect of its acceptability is high among mothers surveyed in south-east Nigeria. Targeted public education by relevant government agencies in collaboration with clinicians, community and religious leaders about the concept of donor breast milk to families may help increase the acceptance and practice of donating breast milk and/or use of donated breast milk among mothers in the region.

     

Synergistic Effect of Afatinib with Su11274 in Non-Small Cell Lung Cancer Cells Resistant to Gefitinib or Erlotinib

Epidermal growth factor receptor (EGFR) and c-MET receptors are expressed on many non-small cell lung cancer (NSCLC) cells. Current single agent therapeutic targeting of a mutant EGFR has a high efficacy in the clinic, but is not curative. Here, we investigated the combination of targeting EGFR and c-MET pathways in NSCLC cells resistant to receptor tyrosine kinase inhibitors (TKIs), using RNA interference and inhibition by TKIs. Different NSCLC cell lines with various genomic characteristics (H358, H1650 and H1975) were transfected with EGFR-specific-siRNA, T790M-specific-siRNA, c-MET siRNA or the combination. Subsequently EGFR TKIs (gefitinib, erlotinib or afatinib) or monoclonal antibody cetuximab were combined respectively with the c-MET-specific TKI su11274 in NSCLC cell lines. The cell proliferation, viability, caspase−3/7 activity and apoptotic morphology were monitored by spectrophotometry, fluorimetry and fluorescence microscopy. The combined effect of EGFR TKIs, or cetuximab and su11274, was evaluated using a combination index. The results showed that the cell lines that were relatively resistant to EGFR TKIs, especially the H1975 cell line containing the resistance T790M mutation, were found to be more sensitive to EGFR-specific-siRNA. The combination of EGFR siRNA plus c-MET siRNA enhanced cell growth inhibition, apoptosis induction and inhibition of downstream signaling in EGFR TKI resistant H358, H1650 and H1975 cells, despite the absence of activity of the c-MET siRNA alone. EGFR TKIs or cetuximab plus su11274 were also consistently superior to either agent alone. The strongest biological effect was observed when afatinib, an irreversible pan-HER blocker was combined with su11274, which achieved a synergistic effect in the T790M mutant H1975 cells. In a conclusion, our findings offer preclinical proof of principle for combined inhibition as a promising treatment strategy for NSCLC, especially for patients in whom current EGFR-targeted treatments fail due to the presence of the T790M-EGFR-mutation or high c-MET expression.