Kriangkrai Chawansuntati

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.

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.

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.

Low expression of activation marker CD69 and chemokine receptors CCR5 and CXCR3 on memory T cells after 2009 H1N1 influenza A antigen stimulation in vitro following H1N1 vaccination of HIV-infected individuals

Unlike well-studied antibody responses to pandemic 2009 H1N1 influenza A virus vaccines in human immunodeficiency virus-infected (HIV+) individuals, less well understood are cell-mediated immune (CMI) responses to this antigen in this susceptible population. We investigated such influenza-specific CMI responses in 61 HIV+ individuals and in 20 HIV-negative (HIV-) healthy controls. Each was vaccinated with a single licensed dose of inactivated, split-virion vaccine comprised of the influenza A/California/7/2009 (H1N1) virus-like strain. Cells collected just prior to vaccination and at 1 and 3 months afterwards were stimulated in vitro with dialyzed vaccine antigen and assayed by flow cytometry for cytokines TNF-α, IFN-γ, IL-2, and IL-10, for degranulation marker CD107a, as well as phenotypes of memory T-cell subpopulations. Comparable increases of cytokine-producing and CD107a-expressing T cells were observed in both HIV+ subjects and healthy HIV-controls. However, by 3 months post-vaccination, in vitro antigen stimulation of peripheral blood mononuclear cells induced greater expansion in controls of both CD4 and CD8 central memory and effector memory T cells, as well as higher expression of the activation marker CD69 and chemokine receptors CCR5 and CXCR3 than in HIV+ subjects. We concluded CD4+ and CD8+ memory T cells produce cytokines at comparable levels in both groups, whereas the expression after in vitro stimulation of molecules critical for cell migration to infection sites are lower in the HIV+ than in comparable controls. Further immunization strategies against influenza are needed to improve the CMI responses in people living with HIV.

Dot enzyme-linked immunosorbent assay strip as a screening tool for detection of autoantibody to interferon gamma in sera of suspected cases of adult-onset immunodeficiency

Being able to detect the presence of autoantibodies to interferon (IFN)‐γ in serum is essential for evaluating patients with suspected adult‐onset immunodeficiency (AOID) with unusual intracellular infections. Most reported patients with AOID have been Asian, although the exact prevalence of this illness is unknown. To date, no standard assay exists to detect autoantibodies to IFN‐γ. An easy‐to‐use, low‐cost assay that can be performed in any laboratory would be a valuable tool for clinical management of AOID, as well as better reveal its prevalence.

Characterization of anti-interferon-γ antibodies in HIV-negative immunodeficient patients infected with unusual intracellular microorganisms

A major characteristic of immunodeficiency associated with life-threatening intracellular infection in adults is the presence of anti-interferon-γ antibodies. Although little is known about the mechanism underlying this syndrome, it is believed that the antibodies inhibit the activity of downstream signaling pathway of interferon-γ. In this study, the characteristics of these antibodies in patients who presented, or have a history of, intracellular infection and were positive to anti-interferon-γ antibodies were investigated. The antibodies exhibited mainly the IgG1 and the IgG4 subtypes and recognized the C-terminal of the interferon-γ linear epitope containing the KRKR motif, which is required for the biological activity of interferon-γ. The antibodies bound to recombinant interferon-γ with significantly lower avidity than antibodies to a recall antigen, tetanus toxoid, suggesting that the antibodies might have not undergone affinity maturation. The data from this study may provide fundamental information to better understand the properties of anti-interferon-γ antibodies, which can be useful for future studies. Impact statement An increase in the number of immunodeficient patients related to autoantibodies to interferon (IFN)-γ has been observed particularly in East Asian adults. These patients are often presented with opportunistic infections caused by intracellular pathogens, including non-tuberculous mycobacteria (NTM), Cryptococcus neoformans, Penicillium marneffei (now called Talaromyces marneffei), and non-typhoidal Salmonella spp. The mortality rate for this syndrome is relatively high with 32% patients dying at the median time of 25 months after diagnosis. Characterization of these autoantibodies may promote better understanding of the syndrome, an emerging health problem affecting East Asia populations and impeding their welfare and economic development.

Hepatitis B Vaccination Induced TNF-α- and IL-2-Producing T Cell Responses in HIV− Healthy Individuals Higher than in HIV

We investigated cytokine production and expression of degranulation marker CD107a after different strategies of hepatitis B virus (HBV) vaccination in human immunodeficiency virus-infected individuals, which were three doses of 20 μg (standard dose group), four doses of 20 μg (four doses group), or four doses of 40 μg (four double doses group), compared to standard dose vaccination in healthy controls. PBMCs collected at different time points were stimulated in vitro with recombinant hepatitis B surface antigen and analyzed by flow cytometry. There was an increase in TNF-α production of total and memory CD4+ T cells at 7 months after vaccination in healthy controls compared to the HIV+ group, which received the same standard vaccination regimen. An increase in the IL-2-producing memory CD4+ T cells in the healthy control group was also observed at 7 months after vaccination. No differences were observed between the healthy controls and both groups of four doses at any time point of study. These results suggest that the standard HBV vaccination schedule might induce better production of TNF-α and IL-2 from CD4+ T cells in healthy individuals. Modification of HBV vaccination schedule by increasing the frequency and/or dosage may improve the CMI response in HIV-infected individuals. This trial is registered with NCT1289106.

Vaccination for 2009 pandemic H1N1 influenza A did not induce conserved epitope-specific memory CD8 T cell responses in HIV+ northern Thai children

The influenza virus causes severe illness in susceptible populations, including children and people living with human immunodeficiency virus (HIV). Here, we investigated cell-mediated immune responses (CMI) against influenza CD8 T cell conserved epitopes in HIV-infected (HIV+) northern Thai children following the 2009 pandemic H1N1 influenza A vaccination. Sixty HIV+ children were vaccinated with two doses of the 2009 pandemic influenza vaccine and their CD8T cell responses were assessed. We found no significant differences in the increase of cytokines-producing and CD107a-expressing CD8+ T cells or CD8+ memory T cells in response to pooled conserved epitopes stimulation in vitro between children with different serologic responses to the vaccine at all time points of the study. Our results suggest that the 2009 pandemic H1N1 vaccine did not induce the conserved epitope-specific immune responses in HIV+ children. Vaccine design and vaccination strategy against influenza in these populations warrant further studies.