Higher rate of long-term serologic response of four double doses vs. standard doses of hepatitis B vaccination in HIV-infected adults: 4-year follow-up of a randomised controlled trial.

BACKGROUND: We previously reported that four doses or four double doses of hepatitis B vaccination regimens could not significantly increase a response rate compared with standard doses. However, the antibody levels were higher in the four doses and four double doses groups. This study followed those patients for at least 3 years and aimed to evaluate the immunogenicity of the three vaccination regimens. METHODS: HIV-infected adults who had CD4+ cell counts > 200 cells/mm3, undetectable plasma HIV-1 RNA, and negative for all hepatitis B virus markers were randomly assigned to receive one of three recombinant vaccines (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) between February 2011 and May 4, 2012. Of 132 participants, 126 were evaluated from August 2015 to January 2016; 42 in the standard doses, 43 in the four doses, and 41 in the four double doses groups. RESULTS: At a median duration of 49.7 months (range 46.7-53.7) after completion of the primary vaccination schedule, the percentages of responders with anti-HBs ≥ 10 mIU/mL were 57.1% (95% CI 41.5-72.8%) in the standard doses group; 76.7% (95% CI 63.6-89.9%) in the four doses group (P = 0.067 vs. the standard doses group); and 80.5% (95% CI 67.8-93.2%) in the four double doses group (P = 0.033 vs. the standard doses group). Factors associated with a responder were the vaccination schedule (either four doses or four double doses groups) and a younger age. CONCLUSIONS: Despite the highly effectiveness of the standard hepatitis B vaccination regimen at 6 months after completion, the long-term immunogenicity was lower than the four double doses regimen among HIV-infected adults with CD4+ cell counts > 200 cells/mm3 and undetectable plasma HIV-1 RNA. The standard vaccination regimen may not be the best strategy to provide long-term immune response against hepatitis B virus among HIV-infected individuals. Trial registration NCT1289106, NCT02713620.

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.

BACKGROUND: 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. METHODS: Our experimental study exploited a dot enzyme-linked immunosorbent assay (Dot-ELISA) strip to detect autoantibodies to IFN-γ. Sera from 66 HIV-negative patients having autoantibodies to IFN-γ as determined by indirect ELISA were tested. RESULTS: Dot enzyme-linked immunosorbent assay was sensitive (100%) and specific (94.5%), with a positive predictive value of 97.6% and a negative predictive value of 100%. CONCLUSION: This simple method provides prompt qualitative results that can be read visually and used in facilities with limited testing capabilities.

Proteomic analysis of human glutathione transferase omega (hGSTO1) stable transfection in a 6-hydroxydopamine-induced neuronal cells.

Parkinson's disease is the second most common neurodegenerative disorder after Alzheimer's disease. The disease is associated with dopaminergic neuron losses in the substantia nigra area of the brain and the formation of cytoplasmic inclusion bodies. Human glutathione transferase omega 1 (hGSTO1) appears to have a role in modulating stress response. The study was aimed to elucidate differentially expressed proteins caused by oxidative stress induced by 6-hydroxydopamine (6-OHDA). Human neuronal cells SH-SY5Y overexpressing hGSTO1 were used to investigate protein glutathionylation and the modulation of cellular protein expression. Therefore SH-SY5Y/hGSTO1 and SH-SY5Y/control lysate proteins were separated by 2D-gel electrophoresis compared with untreated conditions in both standard and non-reducing conditions. In standard conditions, the analysis of protein profiles demonstrated 25 differentially expressed spots and 10 spots were chosen for further protein identification by LC-MS analysis. Several proteins were later identified as vimentin, galectin-1, high mobility group protein B2, clathrin, tropomyosin, heterogenous nuclear ribonucleoprotein and peroxiredoxin-2. Search Tool for Interactions of Chemicals (STITCH) analysis suggested that oxidative stress induced by 6-OHDA involved carbohydrate metabolism in SH-SY5Y via a lactose metabolic pathway. Our results raise the possibility that hGSTO1 modulates the functions of many proteins that play a role in the degenerative cell response of a Parkinson's model.

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

BACKGROUND: Uvaria 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. METHODS: We 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. RESULTS: The 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. CONCLUSIONS: These 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.

Rodent blood-stage Plasmodium survive in dendritic cells that infect naive mice.

Plasmodium spp. parasites cause malaria in 300 to 500 million individuals each year. Disease occurs during the blood-stage of the parasite's life cycle, where the parasite is thought to replicate exclusively within erythrocytes. Infected individuals can also suffer relapses after several years, from Plasmodium vivax and Plasmodium ovale surviving in hepatocytes. Plasmodium falciparum and Plasmodium malariae can also persist after the original bout of infection has apparently cleared in the blood, suggesting that host cells other than erythrocytes (but not hepatocytes) may harbor these blood-stage parasites, thereby assisting their escape from host immunity. Using blood stage transgenic Plasmodium berghei-expressing GFP (PbGFP) to track parasites in host cells, we found that the parasite had a tropism for CD317(+) dendritic cells. Other studies using confocal microscopy, in vitro cultures, and cell transfer studies showed that blood-stage parasites could infect, survive, and replicate within CD317(+) dendritic cells, and that small numbers of these cells released parasites infectious for erythrocytes in vivo. These data have identified a unique survival strategy for blood-stage Plasmodium, which has significant implications for understanding the escape of Plasmodium spp. from immune-surveillance and for vaccine development.

Safety and immunogenicity of CoronaVac and ChAdOx1 heterologous prime-boost vaccines in an overweight population in Chiang Mai, Thailand.

Background: In early 2021, the Ministry of Public Health of Thailand announced heterologous regimens for COVID-19 vaccines using CoronaVac as the first dose followed by ChAdOx1 nCoV-19 at 3 weeks apart. Priority was given to individuals above 60 years old and those who had seven underlying conditions, including obesity. The vaccine regimen was evaluated for safety and immunogenicity in overweight populations in Chiang Mai, Thailand. Methods: Participants who had a COVID-19 vaccination appointment for the heterologous prime-boost regimen were enrolled. Before each immunization and on day 28 following the second dosage, blood samples were taken, and were examined for anti-spike and neutralizing antibodies by using an indirect ELISA and virus neutralization assays. Safety profile of the vaccine regimen was assessed via a self-recorded diary of adverse events after each vaccination. Results: No serious adverse events related to vaccination were reported during study period and the majority of adverse reactions were fatigue and pain at the injection site. The levels of anti-spike IgG were 26.3, 56.4 and 1752.1 BAU/mL at baseline, 21 days after first dose and 28 days after second dose, respectively. At 4 weeks after complete vaccination, the median inhibition rates of neutralizing antibody determined by surrogate neutralization assay against wild type, Delta and Omicron variants were 95.2, 85.0 and 3.8, respectively. Moreover, the NT50 level against wild type and Delta variants determined by pseudotyped virus neutralization assay were 133.3 and 41.7, respectively. The neutralizing activity against Omicron variant was almost lower than cutoff level for detection. Conclusions: The heterologous CoronaVac-ChAdOx1vaccination was safe, well-tolerated and able to induce humoral immunity against wild-type and Delta variants but not against the Omicron variant in overweight population.

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.

A preliminary study on the effectiveness of maternal to neonatal transfer of antibodies against SARS-CoV-2 in the women vaccinated with heterologous CoronaVac-ChAdOx1.

The objective is to evaluate the effectiveness of placental transfer of maternally derived SARS-CoV-2 IgG antibodies after the vaccination of pregnant women with heterologous CoronaVac-ChAdOx1. Thirty pregnant women were vaccinated with CoronaVac as the first dose, followed by ChAdOx1 3 weeks later. The antibody levels in the maternal blood and in the umbilical cord blood at the time of delivery were determined. The results showed that the vaccination effectively increased antibody levels in both mothers and newborns. The antibody levels in the mothers were strongly correlated with those in the newborns (P < .001). The high levels of passive immunity in the newborns were achieved when the first and second doses of vaccination were given more than 40 and 20 d before delivery, respectively. After 1 month of the second dose, the immune levels seemed to decline in the mothers but increase in the newborns. The antibody levels in the newborns appear to be higher than those in the mothers in cases of delivery after 20 d of the second dose (1419 ± 699 vs 1222 ± 593 BAU/L; p < .05). In conclusion, heterologous CoronaVac-ChAdOx1-S schedule can increase antibody levels in a short time during pregnancy. Also, the regimen effectively increases immunity in the newborns. The antibody levels in the newborns appear to be higher than that in the mothers in most cases, if receiving the second dose more than 3 weeks before delivery. Therefore, the regimen should be considered as an effective regimen for pregnant women, especially in settings where mRNA vaccine is not available.

In-vitro antimalarial activity of methanolic leaf- and stem-derived extracts from four Annonaceae plants.

OBJECTIVE: Plants in the Annonaceae family are known for having abundant biologically active secondary metabolites. They have been used in alternative drugs for various diseases in several countries, for instance, the bark of Cananga odorata (Lam.) Hook and Thomson is used for Ophthalmic inflammation and wound healing in Malaysia. Extracts from the leaves and stems of four Annonaceae plants, namely Uvaria longipes (Craib) L.L.Zhou, Y.C.F.Su & R.M.K.Saunders, Dasymaschalon sp., Artabotrys burmanicus A.DC, and Marsypopetalum modestum (Pierre) B.Xue & R.M.K.Saunders were investigated for growth inhibitory activity against blood-stage Plasmodium falciparum growth in vitro and for non-specific cytotoxicity against normal peripheral blood mononuclear cells (PBMCs). Antimalarial activity was assessed by invasion inhibition assay and the percentage of infected red blood cells on blood smears were determined. Cytotoxicity was tested by culturing PBMCs with the extracts, and viabilities were determined by Annexin V/propidium iodide staining. RESULTS: A. burmanicus stem extract and M. modestum leaf extract were capable of inhibiting growth of P. falciparum when used at 200 µg/mL compared to chloroquine. The extracts at effective concentrations, did not affect the viability of PBMCs. These results support further need for characterization of active compounds from specific Annonaceae plants in order to exploit their components for potential malaria treatment.

The anti-leukemic activity of a luteolin-apigenin enriched fraction from an edible and ethnomedicinal plant, Elsholtzia stachyodes, is exerted through an ER stress/autophagy/cell cycle arrest/ apoptotic cell death signaling axis.

BACKGROUND: Elsholtzia is a genus in the family Lamiaceae, and some species in this genus are commonly used for food and in ethnomedicinal formulations by some ethnic groups of China and Thailand. Despite their apparent utility, few studies have been conducted to evaluate their potential as sources of medicinally active agents. PURPOSE: We aimed to investigate the cytotoxicity of ethanolic extracts from three selected edible plant species of the genus Elsholtzia and the most promising extract was further characterized for the bioactive constituents and signaling mechanisms associated with the anti-leukemic activity. MATERIALS AND METHODS: Ethanolic extracts were screened for cytotoxicity using flow cytometry. HPLC and LC-MS were used to analyze the chemical constituents of the most potent fraction from E. stachyodes. The relevant mechanism of action was assessed by western blot and multispectral imaging flow cytometry (MIFC). RESULTS: The most potent anti-leukemic activity was observed with the ethanolic extract from E. stachyodes. Luteolin and apigenin were characterized as the major constituents in the fraction from E. stachyodes. Mechanistically, the luteolin-apigenin enriched fraction (LAEF) induced the UPR, increased autophagic flux, induced cell cycle arrest and apoptotic cell death. LAEF showed significantly less cytotoxicity towards peripheral blood mononuclear cells (PBMCs) as compared to leukemia cell lines. CONCLUSION: This study is the first to report E. stachyodes as a new source of luteolin and apigenin which are capable of triggering leukemic cell death. This could lead to a novel strategy against leukemia using ethnomedicinal plant extracts as an alternative or supplemental anti-cancer agent.

Diagnostic performance between in-house and commercial SARS-CoV-2 serological immunoassays including binding-specific antibody and surrogate virus neutralization test (sVNT).

This study aimed to evaluate the correlation between in-house and commercial binding-specific IgG antibodies and between in-house and commercial SARS-CoV-2 surrogate virus neutralization tests (sVNT). Samples from healthcare workers who received vaccines against SARS-CoV-2 were tested for RBD-specific antibody, S-specific antibody, and in-house ELISA, commercial sVNT, and in-house sVNT, against wild-type SARS-CoV-2. Three hundred and five samples were included in the analysis. The correlation between S-specific binding antibodies and in-house ELISA was 0.96 (95% CI 0.96-0.97) and between RBD-specific antibodies and in-house ELISA was 0.96 (95% CI 0.95-0.97). The Cohen's kappa between in-house sVNT and the commercial test was 0.90 (95% CI 0.80, 1.00). If using 90% inhibition of sVNT as the reference standard, the optimal cut-off value of RBD-specific antibodies was 442.7 BAU/mL, the kappa, sensitivity, and specificity being 0.99, 99%, and 100%, respectively. The optimal cut-off value of S-specific antibodies was 1155.9 BAU/mL, the kappa, sensitivity, and specificity being 0.99, 100%, and 99%, respectively. This study demonstrated a very strong correlation between in-house ELISA and 2 commercial assays. There was also a very strong correlation between in-house and commercial SARS-CoV-2 sVNT, a finding of particular interest which will inform future research.

Comparison of antibody responses following natural infection with Severe Acute Respiratory Syndrome Coronavirus 2 or receipt of CoronaVac or ChAdOx1 (AZD1222) vaccination in Chiang Mai, Thailand.

Background: In Thailand, early vaccination initiatives for SARS-CoV-2 relied on CoronaVac (Sinovac Life Sciences) and ChAdOx1 (Oxford-AstraZeneca) vaccines. However, the data of immunogenicity of these two vaccines in Thai populations is limited. This real time, head-to-head comparative study was conducted to investigate antibody (Ab) responses to SARS-CoV-2 following infection or receipt of either CoronaVac or ChAdOx1 vaccination in Chiang Mai, Thailand. Methods: Sera was collected within two months from participants having a history of documented SARS-CoV-2 infection or at one month after the second dose of CoronaVac vaccine. Sera from participants with a history of receiving one dose of ChAdOx1 vaccination was collected twice, at one month following each vaccine dose. Neutralizing antibodies (NAbs) were assessed using the surrogate neutralization test and anti-spike protein antibodies were assessed using an in-house enzyme-linked immunosorbent assay. Results: The prevalence of NAbs against SARS-CoV-2 was 92.1 %, 95.7 %, 64.1 % and 100 % in the infection group, CoronaVac group, ChAdOx1 group after 1st dose, and ChAdOx1 group after 2nd dose, respectively. The inhibition rate in individuals receiving two doses of ChAdOx1 vaccine (90.8%) was significantly higher than individuals who had recovered from natural infection (71.7%) or individuals who had received two doses of CoronaVac vaccine (66.7%). The prevalence of anti-spike Abs was 97.4 %, 97.8 %, 97.4 % and 100 % in the infection group, CoronaVac group, ChAdOx1 group after 1st dose, and ChAdOx1 group after 2nd dose, respectively. Significantly higher levels of anti-spike Abs were observed in the ChAdOx1 group after two doses of vaccination (1975 AU/mL) compared to those who had recovered from natural infection (468.5 AU/mL) and individuals who had received CoronaVac (554.4 AU/mL). Neutralizing activity had a statistically significant positive correlation with levels of anti-spike Abs. Conclusions: ChAdOx1 vaccine may provide superior immunogenicity than CoronaVac and natural infection.

Comparison of Immunogenicity and Reactogenicity of Five Primary Series of COVID-19 Vaccine Regimens against Circulating SARS-CoV-2 Variants of Concern among Healthy Thai Populations.

To compare immunogenicity and reactogenicity of five COVID-19 vaccine regimens against wild-type SARS-CoV-2 and variants of concern (VoCs) among Thai populations, a prospective cohort study was conducted among healthy participants aged ≥18 years who had never been infected with COVID-19 and were scheduled to get one of the five primary series of COVID-19 vaccine regimens, including CoronaVac/CoronaVac, AZD1222/AZD1222, CoronaVac/AZD1222, AZD1222/BNT162b2, and BNT162b2/BNT162b2. Anti-receptor binding domain (anti-RBD-WT) IgG and neutralizing antibody (NAb-WT) against wild-type SARS-CoV-2 were measured at pre-prime, post-prime, and post-boost visits. NAb against VoCs (NAb-Alpha, NAb-Beta, NAb-Delta, and NAb-Omicron) were assessed at the post-boost visit. Adverse events (AEs) following vaccination were recorded. A total of 901 participants (CoronaVac/CoronaVac: 332, AZD1222/AZD1222: 221, CoronaVac/AZD1222: 110, AZD1222/BNT162b2: 128, and BNT162b2/BNT162b2: 110) were enrolled. Anti-RBD-WT IgG and NAb-WT levels increased substantially after each vaccine dose. At the post-boost visit, BNT162b2/BNT162b2 induced the highest GMC of anti-RBD-WT IgG level (1698 BAU/mL), whereas AZD1222/BNT162b2 induced the highest median NAb-WT level (99% inhibition). NAb levels against VoCs, particularly the Omicron strain, were markedly attenuated for all vaccine regimens (p < 0.001). Overall, no serious AEs following vaccination were observed. All five primary series of COVID-19 vaccine regimens were well-tolerated and elicited robust antibody responses against wild-type SARS-CoV-2 but had attenuated responses against VoCs, particularly the Omicron strain, among healthy Thai populations.

Safety and immunogenicity of the third and fourth doses of vaccine against SARS-CoV-2 following a 2-dose regimen of inactivated whole-virion SARS-CoV-2 vaccine.

This study followed healthcare personnel (HCP) who had completed a primary series of CoronaVac and then received the third and fourth doses of COVID-19 vaccine. The primary objective was to determine the seroconversion rate of neutralizing antibodies against wild-type SARS-CoV-2 and VOCs at day 28 after the third dose of vaccine and day 28 after the fourth dose of vaccine. This prospective cohort study was conducted at Maharaj Nakorn Chiang Mai Hospital, a tertiary care hospital affiliated to Chiang Mai University from July 2021 to February 2022. Two hundred and eighty-three participants were assessed for eligibility; 142 had received AZD1222 and 141 BNT162b2 as the third dose. Seroconversion rates using a 30% inhibition cutoff value against wild-type SARS-CoV-2 were 57.2%, 98.6%, 97.8%, and 98.9% at points before and after the third dose, before and after the fourth dose, respectively among those receiving AZD1222 as the third dose. Frequencies were 31.9%, 99.3%, 98.9%, and 100% among those receiving BNT162b2 as the third dose, respectively. The seroconversion rates against B.1.1.529 [Omicron] were 76.1% and 90.2% (p-value 0.010) at 4 weeks after the third dose in those receiving AZD1222 and BNT162b2 as the third dose, respectively. After a booster with the mRNA vaccine, the seroconversion rates increased from 21.7 to 91.3% and from 30.4 to 91.3% in those receiving AZD1222 and BNT162b2 as the third dose, respectively. No serious safety concerns were found in this study. In conclusion, antibody responses waned over time regardless of the vaccine regimen. The booster dose of the vaccine elicited a humoral immune response against SARS-CoV-2 including SARS-CoV-2 variants of concern, including B.1.1.529 [Omicron], which was circulating during the study period. However, the results might not be extrapolated to other Omicron sublineages.

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.

An IgM monoclonal antibody against domain 1 of CD147 induces non-canonical RIPK-independent necroptosis in a cell type specific manner in hepatocellular carcinoma cells.

CD147/Basigin/EMMPRIN is overexpressed in several cancerous tissues and it has been shown to induce matrix metalloproteinases (MMPs) whose expression is associated with cancer metastasis. Thus, targeting CD147 with monoclonal antibodies (mAbs) potentially has therapeutic applications in cancer immunotherapy. Here, we report the use of anti-CD147 mAbs targeting domain 1 of CD147, namely M6-1D4 (IgM), M6-1F3 (IgM), M6-2F9 (IgM) and M6-1E9 (IgG2a), against several human cancer cell lines. Strikingly, IgM but not IgG mAbs against CD147, especially clone M6-1D4, induced acute cellular swelling, and this phenomenon appeared to be specifically found with hepatocellular carcinoma (HCC) cells. Furthermore, molecular investigation upon treating HepG2 cells with M6-1D4 showed unfolded protein response (UPR) activation, autophagosome accumulation, and cell cycle arrest, but without classic apoptosis related features. More interestingly, prolonged M6-1D4 treatment (24 h) resulted in irreversible oncosis leading to necroptosis. Furthermore, treatment with a mixed lineage kinase domain-like psuedokinase (MLKL) inhibitor and partial knockout of MLKL resulted in reduced sensitivity to necroptosis in M6-1D4-treated HepG2 cells. Surprisingly however, the observed necroptotic signaling axis appeared to be non-canonical as it was independent of receptor-interacting serine/threonine-protein kinase (RIPK) phosphorylation. In addition, no cytotoxic effect on human dermal fibroblast (HDF) was observed after incubation with M6-1D4. Taken together, this study provides clues to target CD147 in HCC using mAbs, as well as sheds new light on a novel strategy to kill cancerous cells by the induction of necroptosis.

Minireview: Insights into anti-interferon-γ autoantibodies.

The association between the presence of anti-interferon-γ autoantibodies and the onset of immunodeficiency with intracellular infections has been clearly established. No standard regimen to control the production of these pathogenic autoantibodies, apart from antimicrobial therapy to eliminate infections, contributes to the medical burden of this syndrome, which sometimes has a fatal outcome. In this review, we summarize the findings on anti-interferon-γ autoantibodies to facilitate further research and to provide guidance for treatment strategies.

Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay.

OBJECTIVES: The aim was to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of hepatitis C virus (HCV) in a single closed tube. METHODS: Plasma samples were collected from 200 HCV-infected patients. HCV-RNA was detected by one-step RT-LAMP processed at 65 °C for 60 min. The amplified products were detected by hydroxynaphthol blue (HNB)-dependent visual method and gel electrophoresis. Specificity was tested against other viruses. Sensitivity was determined using serial dilutions of extracted RNA. RESULTS: The RT-LAMP assay detected 97.5% of HCV-RNA genotype 1, 91.1% of genotype 3, and 100% of genotype 6. The color change was evidenced with the naked eye. The assay demonstrated a clinical sensitivity of 95.5% and specificity of 100%, as well as no cross-reactivity with other viruses (i.e., hepatitis B virus, HIV). The limit of detection was as low as 10 ng per reaction for HCV genotypes 1a and 6, while it was 100 ng for genotype 3a. The assay showed a 100% detection threshold at a viral load of 5.00 log10 IU/mL in the clinical samples tested. CONCLUSIONS: This study demonstrated the use of an RT-LAMP assay for the detection of HCV in a simple, rapid, and cost-effective manner, which will be useful in resource-limited settings to allow the identification of individuals in need of HCV treatment.

Latticed Gold Nanoparticle Conjugation via Monomeric Streptavidin in Lateral Flow Assay for Detection of Autoantibody to Interferon-Gamma.

Adult-onset immunodeficiency syndrome (AOID) patients with autoantibodies (autoAbs) against interferon-gamma (IFN-γ) generally suffer from recurrent and recalcitrant disseminated non-tuberculous mycobacterial diseases. Since the early stages of AOID do not present specific symptoms, diagnosis and treatment of the condition are not practical. A simplified diagnostic method for differentiating AOID from other immunodeficiencies, such as HIV infection, was created. Anti-IFN-γ is generally identified using enzyme-linked immunosorbent assay (ELISA), which involves an instrument and a cumbersome process. Recombinant IFN-γ indirectly conjugated to colloidal gold was used in the modified immunochromatographic (IC) strips. The biotinylated-IFN-γ was incorporated with colloidal-gold-labeled 6HIS-maltose binding protein-monomeric streptavidin (6HISMBP-mSA) and absorbed at the conjugate pad. The efficacy of the IC strip upon applying an anti-IFN-γ autoAb cut-off ELISA titer of 2500, the sensitivity and specificity were 84% and 90.24%, respectively. When a cut-off ELISA titer of 500 was applied, the sensitivity and specificity were 73.52% and 100%, respectively.

The mechanism and significance of deletion of parasite-specific CD4(+) T cells in malaria infection.

It is thought that both helper and effector functions of CD4(+) T cells contribute to protective immunity to blood stage malaria infection. However, malaria infection does not induce long-term immunity and its mechanisms are not defined. In this study, we show that protective parasite-specific CD4(+) T cells were depleted after infection with both lethal and nonlethal species of rodent PLASMODIUM: It is further shown that the depletion is confined to parasite-specific T cells because (a) ovalbumin (OVA)-specific CD4(+) T cells are not depleted after either malaria infection or direct OVA antigen challenge, and (b) the depletion of parasite-specific T cells during infection does not kill bystander OVA-specific T cells. A significant consequence of the depletion of malaria parasite-specific CD4(+) T cells is impaired immunity, demonstrated in mice that were less able to control parasitemia after depletion of transferred parasite-specific T cells. Using tumor necrosis factor (TNF)-RI knockout- and Fas-deficient mice, we demonstrate that the depletion of parasite-specific CD4(+) T cells is not via TNF or Fas pathways. However, in vivo administration of anti-interferon (IFN)-gamma antibody blocks depletion, suggesting that IFN-gamma is involved in the process. Taken together, these data suggest that long-term immunity to malaria infection may be affected by an IFN-gamma-mediated depletion of parasite-specific CD4(+) T cells during infection. This study provides further insight into the nature of immunity to malaria and may have a significant impact on approaches taken to develop a malaria vaccine.