Jiraprapa Wipasa

Long-Lived Antibody and B Cell Memory Responses to the Human Malaria Parasites, Plasmodium falciparum and Plasmodium vivax

Antibodies constitute a critical component of the naturally acquired immunity that develops following frequent exposure to malaria. However, specific antibody titres have been reported to decline rapidly in the absence of reinfection, supporting the widely perceived notion that malaria infections fail to induce durable immunological memory responses. Currently, direct evidence for the presence or absence of immune memory to malaria is limited. In this study, we analysed the longevity of both antibody and B cell memory responses to malaria antigens among individuals who were living in an area of extremely low malaria transmission in northern Thailand, and who were known either to be malaria naïve or to have had a documented clinical attack of P. falciparum and/or P. vivax in the past 6 years. We found that exposure to malaria results in the generation of relatively avid antigen-specific antibodies and the establishment of populations of antigen-specific memory B cells in a significant proportion of malaria-exposed individuals. Both antibody and memory B cell responses to malaria antigens were stably maintained over time in the absence of reinfection. In a number of cases where antigen-specific antibodies were not detected in plasma, stable frequencies of antigen-specific memory B cells were nonetheless observed, suggesting that circulating memory B cells may be maintained independently of long-lived plasma cells. We conclude that infrequent malaria infections are capable of inducing long-lived antibody and memory B cell responses.

Investigation of the anti-inflammatory effect of Curcuma longa in Helicobacter pylori-infected patients.

Helicobacter pylori infection of the lining of the stomach induces an array of inflammatory cytokine production leading to gastritis and peptic ulcer disease. The aim of this study was to investigate the effect of curcumin on the production of interleukin (IL)-8, IL-1beta, tumor necrosis factor (TNF)-alpha and cyclooxygenase (COX)-2 in gastric mucosa from H. pylori-infected gastritis patients. Patients were randomly assigned to receive either OAM (Omeprazole, Amoxicillin and Metronidazole) treatment or a course of curcumin. Gastric biopsies were collected before and after treatment and were examined for the level of inflammatory cytokines mRNA by semi-quantitative reverse transcription polymerase chain reaction. The eradication rate of H. pylori in patients that received OAM treatment was significantly higher than the patients that received curcumin (78.9% versus 5.9%). The levels of IL-8 mRNA expression in the OAM group significantly decreased after treatment, but no changes of other cytokines were found. This emphasizes an important role of IL-8 in H. pylori infection. The decreases of cytokine production were not found in the curcumin group. We concluded that curcumin alone may have limited anti-bactericidal effect on H. pylori, and on the production of inflammatory cytokines. Nevertheless, other studies have reported that patients treated with curcumin had relieved symptoms. Further investigation should be carried out as the use of curcumin in combination with therapeutic regimens may be beneficial as an alternative treatment.

CpG oligodeoxynucleotide enhances immunity against blood-stage malaria infection in mice parenterally immunized with a yeast-expressed 19 kDa carboxyl-terminal fragment of Plasmodium yoelii merozoite surface protein-1 (MSP119) formulated in oil-based Montanides

The 19kDa carboxyl-terminal fragment of Plasmodium yoelii merozoite surface protein-1 (MSP1(19)), an analog of the leading falciparum malaria vaccine candidate, induces protective immunity to challenge infection when formulated with complete/incomplete Freunds adjuvant (CFA/IFA), an adjuvant unsuitable for use in humans. In this study, we investigate Montanide ISA51 and Montanide ISA720 as well as CpG oligodeoxynucleotide (ODN) as adjuvants for induction of immunity to MSP1(19). Mice immunized with MSP1(19) adjuvanted with Montanide ISA51 were protected even though some mice experienced low-grade parasitemia before resolving the infection. Mice immunized with MSP1(19) adjuvanted with Montanide ISA720 showed delayed patent parasitemia with all mice ultimately succumbing to infection. Interestingly, when the synthetic CpG ODN 1826 was included in either Montanide formulation, mice were completely protected with no parasites detected in the blood. MSP1(19)-specific antibodies in MSP1(19)-immunized mice adjuvanted with Montanide ISA51 or Montanide ISA720 showed predominantly IgG1 antibody and low levels of IgG2a. CpG ODN 1826 significantly enhanced both IgG1 and IgG2a antibody responses in Montanide ISA51-adjuvanted mice but significantly enhanced only the IgG2a antibody response in Montanide ISA720-adjuvanted mice. To investigate the relative roles of antibody and CD4(+) T cells in protection, MSP1(19)-immunized mice adjuvanted with Montanide ISA720 and CpG ODN 1826 were depleted of CD4(+) T cells just prior to challenge. Results showed that three of nine immunized/T cell depleted mice died following infection. These results suggest that antibody and CD4(+) T cells are critical for protection following immunization with MSP1(19) adjuvanted with Montanide and CpG ODN and that the formulation of a human malaria vaccine candidate in Montanide ISA720 or ISA51 together with human compatible CpG ODN would be useful for improving efficacy.

Short-lived IFN-γ effector responses, but long-lived IL-10 memory responses, to malaria in an area of low malaria endemicity.

Immunity to malaria is widely believed to wane in the absence of reinfection, but direct evidence for the presence or absence of durable immunological memory to malaria is limited. Here, we analysed malaria-specific CD4+ T cell responses of individuals living in an area of low malaria transmission in northern Thailand, who had had a documented clinical attack of P. falciparum and/or P. vivax in the past 6 years. CD4+ T cell effector memory (CD45RO+) IFN-γ (24 hours ex vivo restimulation) and cultured IL-10 (6 day secretion into culture supernatant) responses to malaria schizont antigens were detected only in malaria-exposed subjects and were more prominent in subjects with long-lived antibodies or memory B cells specific to malaria antigens. The number of IFN-γ-producing effector memory T cells declined significantly over the 12 months of the study, and with time since last documented malaria infection, with an estimated half life of the response of 3.3 (95% CI 1.9–10.3) years. In sharp contrast, IL-10 responses were sustained for many years after last known malaria infection with no significant decline over at least 6 years. The observations have clear implications for understanding the immunoepidemiology of naturally acquired malaria infections and for malaria vaccine development.

Comparison of Immunogenicity and Safety of Four Doses and Four Double Doses vs. Standard Doses of Hepatitis B Vaccination in HIV-Infected Adults: A Randomized, Controlled Trial

Background HBV vaccination is recommended in HIV-infected adults with CD4+ cell count >200/mm3 although the efficacy is only 33.3% -65%. We conducted a randomized, controlled trial to evaluate the efficacy and safety of three regimens of HBV vaccination at Chiang Mai University Hospital, Thailand. Methods From February 4, 2011 to May 4, 2012, 132 HIV-infected adults with CD4+ cell counts >200 cells/mm3, undetectable plasma HIV-1 RNA, and negative for all HBV markers were randomly assigned to receive one of three recombinant vaccine (Hepavax-Gene® Berna, Korea) regimens: 20 μg IM at months 0, 1, and 6 (Standard doses group, n=44), 20 μg IM at months 0, 1, 2, 6 (four doses group, n=44), or 40 μg IM at months 0, 1, 2, and 6 (four double doses group, n=44). The primary outcomes were to compare the immunogenicity and safety between the four-doses groups with the Standard doses group. Results At months 7 and 12, the percentages of responders (anti-HBs ≥10 mIU/mL) were 88.6% and 70.4% in the Standard doses group, 93.2% and 86.4% in the four doses group, (P=0.713 and 0.119), and 95.4% and 88.6% in the four double doses group, (P=0.434 and 0.062), respectively. Factors associated with a high titer level (anti-HBs ≥100 mIU/mL) were vaccination schedule and younger age. The most common adverse event was pain at the injection site (42.4%); this was significantly more frequent in the four double doses group compared to the Standard doses group. No serious adverse events were observed. Conclusions In Northern Thailand, the standard three-doses HBV vaccination in HIV-infected adults with CD4+ cell counts >200 cells/mm3 and undetectable plasma HIV-1 RNA is highly effective. Although regimens of four injections of either standard or double doses could not significantly increase the response rate, these regimens may induce higher levels of antibody to the virus. Trial registration information: ClinicalTrials.gov; NCT1289106; http://clinicaltrials.gov/ct2/show/NCT01289106

Autoantibody to Interferon-gamma Associated with Adult-Onset Immunodeficiency in Non-HIV Individuals in Northern Thailand

Background Autoantibody to interferon-gamma (IFN-γ) has been reported to be associated with adult-onset immunodeficiency in patients from Asian countries. This study aimed to determine the prevalence of autoantibody to IFN-γ among non-HIV patients in northern Thailand who were repeatedly infected with unusual intracellular pathogens. Methods A cross-sectional, case-control study was conducted between March 2011 and March 2012 at Chiang Mai University Hospital. 20 cases, non-HIV, aged 18–60 years, presented with at least 2 episodes of culture or histopathology proven opportunistic infections were enrolled. Controls comprised 20 HIV-infected patients and 20 healthy adults who were age- and sex-matched with cases. Enzyme-linked immunosorbent assay (ELISA) was used to detect the presence of antibody to IFN-γ. Results 11 participants in each group were female. The mean ages were 48.1±6.4, 48.3±6.3, and 47.1±6.5 years among cases, HIV-infected, and healthy controls, respectively. The opportunistic infections among 20 cases included disseminated non-tuberculous mycobacterial (NTM) infection (19 patients/24 episodes), disseminated penicilliosis marneffei (12 patients/12 episodes), and non-typhoidal Salmonella bacteremia (7 patients/8 episodes). At the cutoff level of 99 percentile of controls, the prevalence of autoantibody to IFN-γ were 100%, 0%, and 0%, among cases, HIV-infected, and healthy controls, respectively (p-value <0.001). The mean concentrations of antibody to IFN-γ were 3.279±0.662 and 0.939±0.630 O.D. among cases with and without active opportunistic infection, respectively (p-value<0.001). Conclusions In northern Thailand, autoantibody to IFN-γ was strongly associated with adult-onset immunodeficiency. The level of antibody to IFN-γ in patients who had active opportunistic infection was relatively higher than those without active infection.

The immunological challenges of malaria vaccine development

Malaria remains an important public health problem throughout the tropical world causing immense human suffering and impeding economic development. Despite extensive research for > 100 years, options for preventing malaria remain limited to vector control and chemoprophylaxis. The complexity of the organism and its life cycle have, thus far, thwarted vaccine development and exacerbated the perennial problems of drug resistance. Nevertheless, development of a vaccine against malaria that reduces morbidity and mortality, and ideally also reduces transmission, has long been seen as an essential component of a sustainable malaria control strategy. In this article the authors review the biological challenges of malaria vaccine development, summarise some of the recent advances and offer some immunological insights which might facilitate further research.

Hemoglobin E Prevalence among Ethnic Groups Residing in Malaria-Endemic Areas of Northern Thailand and Its Lack of Association with Plasmodium falciparum Invasion In Vitro

Hemoglobin E (HbE) is one of the most common hemoglobin variants caused by a mutation in the β-globin gene, and found at high frequencies in various Southeast Asian groups. We surveyed HbE prevalence among 8 ethnic groups residing in 5 villages selected for their high period malaria endemicity, and 5 for low endemicity in northern Thailand, in order to uncover factors which may affect genetic persistence of HbE in these groups. We found the overall HbE prevalence 6.7%, with differing frequencies from 0% in the Pwo Karen, the Lawa, and the Skaw Karen to 24% in the Mon. All HbE genes were heterozygous (AE). Differences in HbE prevalence among the studied ethnic groups indirectly documents that ancestries and evolutionary forces, such as drift and admixture, are the important factors in the persistence of HbE distribution in northern Thailand. Furthermore, the presence of HbE in groups of northern Thailand had no effect on the in vitro infectivity and proliferation of Plasmodium falciparum, nor the production of hemozoin, a heme crystal produced by malaria parasites, when compared to normal red-blood-cell controls. Our data may contribute to a better understanding on the persistence of HbE among ethnic groups and its association with malaria.

Cell cycle arrest and apoptosis induction by methanolic leaves extracts of four Annonaceae plants

BackgroundUvaria longipes (Craib) L.L.Zhou, Y.C.F.Su & R.M.K.Saunders, Artabotrys burmanicus A.DC, Marsypopetalum modestum (Pierre) B.Xue & R.M.K.Saunders and Dasymaschalon sp. have been used for traditional medicine to treat cancer-like symptoms in some ethnic groups of Thailand and Laos.MethodsWe evaluated the anti-cancer activity of these Annonaceae plants against several human cancer cell lines. The apoptosis induction was detected by Annexin/propidium iodide (PI) staining. Phytochemical screening was tested by standard protocols and bioactive compounds were determined by HPLC.ResultsThe crude extracts from leaves of U. longipes, Dasymaschalon sp., A. burmanicus, and M. modestum showed particular effects that were found to vary depending on the cancer cell line, suggesting that the effect was in a cell-type specific manner. Interestingly, the induction of apoptotic cell death was prominent by the leaves-derived crude extract of M. modestum. This crude was, therefore, subjected to cell cycle analysis by PI staining. Results showed that this crude extract arrested cell cycle and increased the percentage of cells in the SubG1 phase in some cancer cell lines. The phytochemical screening tests indicated that all crude extracts contained tannins and flavonoids. HPLC of flavonoids using standards identified rutin as an active compound in U. longipes and Dasymaschalon sp., whereas quercetin was found in U. longipes and M. modestum.ConclusionsThese crude extracts provide a new source for rutin and quercetin, which might be capable of inducing cancer cell apoptotic death in a cell-type specific manner. This suggests, by analyzing the major bioactive compounds, the potential use of these crudes for chemotherapy in the future.

Cellular Immune Responses in HIV-Negative Immunodeficiency with Anti-Interferon-γ Antibodies and Opportunistic Intracellular Microorganisms

Background Cell-mediated immunity plays a crucial role in resistance to intracellular infection. We previously reported antibodies against interferon-gamma (IFN-γ) in HIV− negative (HIV−) patients with acquired immunodeficiency presenting with repeated episodes of disseminated infection caused by uncommon opportunistic intracellular fungal, bacterial, and viral pathogens. This follow-up study aimed to investigate cellular immune responses in these unusual patients. Methods Twenty HIV− patients presenting with ≥2 episodes of culture- or histopathologic-proven opportunistic infections were enrolled along with age- and sex-matched controls comprised of 20 HIV+ patients plus 20 healthy adults. Monocyte phenotyping and intracellular cytokine production were determined by staining with specific antibodies followed by flow cytometry. Anti-interferon-γ antibodies were measured by enzyme-linked immunosorbent assay, and inducible nitric oxide synthase by reverse-transcription polymerase chain reaction. Results There were no differences among cases, HIV+, and healthy controls in the percentage of monocytes, or CD68 and HLA-DR expression on their surfaces. FcR1 (CD119) expression on monocytes was significantly higher in cases than in HIV+ (p<0.05) and healthy controls (p<0.01), suggesting the presence of activated monocytes in the circulation. Interleukin (IL)-2 and tumor necrosis factor (TNF)-α production in CD4 cells were significantly lower in cases than in healthy controls (p<0.01 and p<0.001, respectively). CD8 production of TNF-α among cases was significantly lower than that of healthy controls (p<0.05). Conclusion Immunodeficiency in HIV− individuals with repeated infections with intracellular pathogens may be associated with one or more of the abnormal immune responses reflected by the reduced production of both IL-2 by CD4 T cells and TNF-α by CD4 T cells and CD8 T cells, as well as presence of anti-IFN-γ antibody, as previously reported.