Malaria diagnostic update: From conventional to advanced method.

BACKGROUND: Update diagnostic methods play essential roles in dealing with the current global malaria situation and decreasing malaria incidence. AIM: Global malaria control programs require the availability of adequate laboratory tests in the quick and convenient field. RESULTS: There are several methods to find out the existence of parasites within the blood. The oldest one is by microscopy, which is still a gold standard, although rapid diagnostic tests (RDTs) have rapidly become a primary diagnostic test in many endemic areas. Because of microscopy and RDTs limitation, novel serological and molecular methods have been developed. Many kinds of polymerase chain reaction (PCR) provide rapid results and higher specificity and sensitivity. The loop-mediated isothermal amplification (LAMP) and biosensing-based molecular techniques as point of care tests (POCT) will become a cost-effective approach to advance diagnostic testing. CONCLUSION: Despite conventional techniques are still being used in the field, the exploration and field implementation of advanced techniques for the diagnosis of malaria are still being developed rapidly.

Coelomic Fluid of Lumbricus rubellus Synergistically Enhances Cytotoxic Effect of 5-Fluorouracil through Modulation of Focal Adhesion Kinase and p21 in HT-29 Cancer Cell Line.

Coelomic fluid of Lumbricus rubellus (CFL) has attracted interest due to its pharmacological properties, including antitumor effect. Furthermore, it is necessary to evaluate the response to treatment with new cancer therapeutic agents. This study aims to investigate whether the combination of CFL and 5-fluorouracil could reduce FAK protein level and iCa2+ and enhance p21 level. Furthermore, it is necessary to evaluate the response to treatment with new cancer therapeutic agents. After 24 hours of treatment, it was necessary to assess the percentage of apoptosis, FAK, and p21 protein expression by flow cytometry. iCa2+ concentration was measured using immunofluorescence. The combination therapy of CFL with 5-fluorouracil potently suppressed six treatment groups were included in this study. HT-29 cell lines were cultured and divided into six groups: group 1 was treated with vehicle (negative control), groups 2-5 were treated with 5-fluorouracil, groups 3-5 were treated with either CFL 5, 10, or 20 µg/ml immediately after 5-fluorouracil, and group 6 was treated with CFL 20 µg/ml, the progression of colorectal cancer. Combination of CFL and 5-fluorouracil significantly decreased FAK expression (p<0.05), iCa2+ (p<0.05), and increased p21 expression (p<0.05) in HT-29 cells. Our results suggest that CFL has an anticancer potential in colorectal cancer when combined with 5-fluorouracil.

T-Bet Is Dependent on Stat-4 Inhibiting Acute Colitis but Not Stat-1 Using L4 Somatic Antigen of Heligmosomoides polygyrus.

Helminths may alter the immunoinflammatory reactions of colitis. Proteins derived from H. polygyrus have prospective therapy for colitis. The goal of this study was to interpret the protective mechanisms of L4 somatic antigen (LSA) from Heligmosomoides polygyrus against an inflammatory response to the pathogenesis of DNBS-induced colitis. Colitis was actuated in mice by rectal instillation of DNBS. The mice were randomly divided into five groups containing control, DNBS alone, and three groups, with different doses of LSA (50, 100, and 200 µg/mL), respectively. Mice initiated colitis by rectal administration of DNBS and after that were immunized with LSA for 14 days. Mice treated with LSA inhibited wasting disease compared with DNBS only group. The percentages of cells producing IFN-γ were reduced by LSA treatment. The level of T lymphocytes CD4+IFN-γ+ cells in the LPL was significantly diminished by LSA at both 100 and 200 µg/mL groups (p<0.05). The mRNA expression of T-bet was significantly declined in LSA immunized mice, but not RORγ-T mRNA, whereas GATA-3 expression tended to increase. The activation of STAT-4 significantly reduced LSA-treated mice but not STAT-1. It can be concluded that T-bet is required for optimal production of IFN -γ in colitis.

Extract from mango mistletoes Dendrophthoe pentandra ameliorates TNBS-induced colitis by regulating CD4+ T cells in mesenteric lymph nodes.

BACKGROUND: Mango mistletoes Dendrophthoe pentandra (MMDP) extract has attracted interest due to its pharmacological properties, including gastro protective effects. The aim of this study was to investigate whether MMDP extract could increase Foxp3 regulatory T cells and inhibits development of Th17 cells. METHODS: Colitis was induced in Balb/c mice by rectal administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS). The mice were randomly divided into five groups comprising group1 receiving vehicle (the negative control), group 2-5 receiving TNBS, group 3-5 orally receiving either MMDP extract 150, 300 and 600 mg/kgBW for 7 days after TNBS administration. On day 8 of the experiment, the colon tissues were removed for histological examination, cytokine and myeloperoxidase (MPO) measurement. T-cells sub-population in mesenteric lymph nodes were analyzed by flow cytometer. RESULTS: MMDP extract potently suppressed colon shortening and MPO in mice with TNBS-induced colitis. Administration of the extract significantly decreased the severity of TNBS-induced colitis in a dose-dependent manner. The extract significantly attenuated the loss of body weight (p < 0.05). These effects were associated with a remarkable amelioration of the disruption of the colonic architecture, significant reduction of the colonic MPO (p < 0.05). The extract lowered the levels of Th17-associated cytokines but increased the production of Treg-associated cytokines in mesenteric lymph node cells. CONCLUSION: Our results suggest that MMDP has the therapeutic potential to ameliorate TNBS-induced colitis symptoms revealed by histological change and inhibit IL-17 production.

Dendrophthoe pentandra (L.) Miq extract effectively inhibits inflammation, proliferation and induces p53 expression on colitis-associated colon cancer.

BACKGROUND: Indonesian mistletoe grows on various trees. Mango Mistletoes (Dendrophthoe pentandra) is one type of mistletoe that grown on mango tree (.benalu mangga in bahasa Indonesia). Our study used mistletoe as a parasitic plant that has been used for traditional medicine. It has been known that Dendrophtoe pentandra extract (DPE) anti-inflammatory and anticancer. Furthermore, it is necessary to follow-up study in vivo to evaluate the response to treatment of new cancer therapeutic agents. This research aimed to determine the levels of IL-22, myeloperoxide (MPO), proliferation and wild-type p53 expression after the administration of DPE to murine models of CAC. METHODS: Mouse colitis associated colon cancer (CAC) was induced firstly by azoxymethane (AOM) and followed by administration of drinking water containing 5 % dextran sodium sulfate (DSS) in a cycle protocol, each cycle consisted of seven days of 5 % DSS in the drinking water and followed by seven days of regular water. This study consists of five treatment groups: I was treated water only (control), II was administrated by (DSS only, without DPE), (III-V) were administrated by DPE (125 mg/kg BW, 250 mg/kg BW and 500 mg/kg BW) respectively. The administrated of DPE were started from the 8th weeks, were continued until 21 weeks. At the end of 21 weeks of the experiment, mice were sacrificed, colon tissue was removed, and then subjected to ELISA, flow cytometry, real-time PCR and histology examination. RESULTS: Administration of DPE 250 mg/kgBW significantly reduce the levels of IL-22 and MPO compared with DSS only group (p < 0.001; p < 0.001). Colonic epithelial cells proliferation of group IV (DPE 250 mg/kgBW) were significantly lower than III and V groups. There was no significant change in the S phase in mice were treated DPE 125 mg/kg BW and 500 mg/kg BW, while administration of DPE 250 mg/kg BW was able to increase the percentage of cells in S phase. The expression of mRNA p53 was up regulated in mice received DPE 125 mg/kg BW. CONCLUSION: These findings indicate that the DPE could inhibit colonic epithelial cells proliferation through p53 pathway independently. This study also showed that DPE could be potential sources of new therapy.

Treatment of low doses curcumin could modulate Th17/Treg balance specifically on CD4+ T cell cultures of systemic lupus erythematosus patients.

INTRODUCTION: The balance between T helper 17 (Th17) and regulatory T cells (Treg) is a new paradigm in the pathogenesis of systemic lupus erythematosus (SLE). Currently, there are no drugs that able to modulate Th17/Treg balance specifically in SLE. Curcumin is a bioactive agent that has a specific action against hyperproliferative cells. However, its role in modulating Th17/Treg balance in SLE is still unknown. This research aimed to investigate the role of curcumin in modulating Th17/Treg balance on CD4+ T cell cultures of SLE patients. MATERIAL AND METHODS: CD4+ T cells from SLE 6 untreated patients and 6 healthy subjects were collected, stimulated with Th17 differentiating factors, and curcumin 0.1 and 1 µg/ml was added on cultures. After 72 hours incubation, cells were harvested and measured for Th17 and Treg percentages using flow cytometry and interleukin-17A (IL-17A) and transforming growth factor-ß1 (TGF-ß1) levels using ELISA. RESULTS: Administration of low doses of curcumin (0.1 and 1 µg/ml) could decrease Th17 percentages (p = 0.000 and p = 0.000 compared to control), reduce IL-17A productions (p = 0.000 and p = 0.000 compared to control), increase Treg percentages (p = 0.001 and p = 0.000 compared to control), and increase TGF-ß1 productions (p = 0.001 and p = 0.000 compared to control) on CD4+ T cells of SLE patients. Interestingly, these effects were not reproduced on CD4+ T cells cultures of healthy subjects. CONCLUSIONS: These data suggest that curcumin can modulate Th17/Treg balance specifically on CD4+ T cells of SLE patients without affecting healthy subjects.

STAT3 Expression and Its Correlation with PD-L1 Expression in Non-Hodgkin's Lymphoma and Hodgkin's Lymphoma at Dr. Saiful Anwar Regional Public Hospital in Malang, Indonesian Population.

Background: Lymphomas are malignant lymphocyte neoplasms that globally account for 10% of cancers in individuals aged <20 years. Malignant lymphomas are divided into Hodgkin's lymphoma (HL) and non-Hodgkin's lymphoma (NHL). Despite the availability of many therapeutic modalities for lymphoma, such as Brentuximab vedotin, Nivolumab, and Pembrolizumab, it is still necessary to identify appropriate strategies with minimal side effects. Immunotherapy is a promising approach, exemplified by targeting JAK/STAT3 signaling, which can inhibit tumor growth and enhance antitumor immune responses. Hence, STAT3 (signal transducer and activator of transcription 3) is a promising therapeutic target. PD-L1 (programmed death-ligand 1), an immune checkpoint molecule, is used as a frontline treatment for various cancers. This study aims to determine STAT3 expression and its correlation with PD-L1 expression in NHL and HL to serve as a basis for further research on anti-STAT3 and its combination with other therapy targets. Methods: Samples were obtained from paraffin blocks of patients with confirmed diagnoses of NHL and HL, and then immunohistochemical staining was carried out with PD-L1 and STAT3 antibodies. The collected data were then analyzed using SPSS. Results: Among the 10 HL patients, no patients (0%) expressed STAT3, while nine patients (90%) expressed PD-L1. Among the 10 NHL patients, 1 patient (10%) expressed STAT3, while six patients (60%) expressed PD-L1. There were no significant differences in STAT3 expression and PD-L1 expression between HL patients and NHL patients. There was no correlation between STAT3 and PD-L1 expression in HL and NHL because almost all STAT3 expressions were negative. Conclusion: Although this study revealed no differences between STAT3 and PD-L1 expression in HL and NHL and no significant correlation between STAT3 and PD-L1 expression in HL and NHL, this may serve as the basis for understanding the role of STAT3 and PD-L1 in the regulation of HL and NHL, which may be useful for further research targeting STAT3 and PD-L1 immunotherapy in HL and NHL.

PD-L1, STAT3, IL6, and EGFR Immunoexpressions in High-Grade Osteosarcoma.

Introduction: Immunotherapy has been widely used in the treatment of various malignancies with satisfactory results. One of the agents for immunotherapy is an inhibitor of programmed cell death-1 and its ligands (PD-1 and PD-L1). However, attempts at utilizing PD-1/PD-L1 immunotherapy in osteosarcoma have not yielded favorable results. This may be due to differences in PD-L1 regulation and the immune landscape in osteosarcoma, as the mechanism is still poorly understood. Therefore, elucidating PD-L1 regulation in osteosarcoma is paramount in order to improve treatment results using immunotherapy. Methods: This is a cross-sectional study conducted in the Department of Anatomical Pathology of Saiful Anwar Hospital using 33 paraffin blocks of confirmed cases of osteosarcoma. Immunohistochemical staining using PD-L1, STAT3, IL6, and EGFR was performed. Statistical analyses were subsequently performed on the immunoexpression data of these antibodies. Results: PD-L1, STAT3, IL6, and EGFR expressions were found in 6 (18.2%), 6 (18.2%), 28 (84.8%), and 30 (90.9%) cases, respectively. There were significant correlations between PD-L1 and STAT3 (r = 0.620, p=<0.001), PD-L1 and EGFR (r = 0.449, p=0.009), as well as STAT3 and EGFR (r = 0.351, p=0.045). Conclusion: The existence of a correlation between PD-L1, STAT3, and EGFR indicates the potential role of STAT3 and EGFR in PD-L1 regulation in osteosarcoma, which may become the basis for targeted therapy.

CD8+CD122+ regulatory T cells (Tregs) and CD4+ Tregs cooperatively prevent and cure CD4+ cell-induced colitis.

We identified CD8(+)CD122(+) regulatory T cells (Tregs) and demonstrated their importance in the maintenance of immune homeostasis and in the recovery from experimental autoimmune encephalomyelitis. In this paper, we show that CD8(+)CD122(+) Tregs effectively prevent and cure colitis in a mouse model. In our experiments, colitis was induced in lymphocyte-deficient RAG-2(-/-) mice by transferring CD4(+)CD45RB(high) cells that were excluded with CD4(+) Tregs. Cotransfer of CD8(+)CD122(+) cells clearly suppressed the development of colitis, and this suppressive effect was similar to that of CD4(+)CD45RB(low) cells that were mostly CD4(+) Tregs. CD8(+)CD122(+) cells obtained from IL-10(-/-) mice were unable to suppress colitis, indicating that IL-10 is an important effect-transmitting factor in the suppression of colitis. CD8(+)CD122(+) cells showed a suppressive effect when they were transferred 4 wk after CD4(+)CD45RB(high) cells, indicating the therapeutic potential of CD8(+)CD122(+) cells. A mixture of CD8(+)CD122(+) cells and CD4(+)CD45RB(low) cells was far more effective than single Tregs, indicating the synergistic effect of these Tregs. These overall findings demonstrate the potential role of CD8(+) Tregs, and possibly together with CD4(+) Tregs, in the medical care of inflammatory bowel disease patients.

Correlation Between IL-8, C-Reactive Proteins (CRP) and Neutrophil to Lymphocyte Ratio (NLR) as Predictor of Mortality in COVID-19 Patients with Diabetes Mellitus Comorbidity.

Background: COVID-19 is caused by SARS-CoV-2 and has a wide range of symptoms. While Diabetes Mellitus (DM) is a common comorbidity in COVID-19 patients, it is the main comorbidity in non-surviving COVID-19 patients. Interleukin-8 (IL-8) is a cytokine that has been correlated with severity and mortality in COVID-19 patients, but its role in COVID-19 patients with DM comorbidity and its relationship with NLR and CRP as markers of inflammation are not yet fully understood. Objective: To investigate the correlation between IL-8, NLR, and CRP in COVID-19 patients with DM comorbidity. Methods: A cross-sectional study was conducted at the Integrated Infectious Disease Installation of Dr. Saiful Anwar Malang Hospital from June to November 2021 using consecutive sampling. IL-8 was measured using the ELISA method with Legendmax® Human IL-8. NLR was measured using flow cytometry, while CRP was measured using the immunoturbidimetric method with Cobas C6000®. Patient outcomes were obtained from medical records. Results: A total of 124 research subjects participated in the study. IL-8 and CRP levels were significantly higher (p < 0.05) in COVID-19 patients with DM comorbidity, and were also significantly higher (p < 0.05) in non-surviving COVID-19 patients. Overall, there was a positive correlation between IL-8 and CRP (r = 0.58, p < 0.05). There was also a positive correlation between IL-8 (r = 0.58; p < 0.05), NLR (r = 0.45, p < 0.05), CRP (r = 0.54, p < 0.05) and mortality in COVID-19 patients with DM comorbidity. The presence of DM comorbidity increased IL-8 levels and aggravated inflammation in COVID-19 patients, thereby increasing the risk of mortality. Conclusion: IL-8, CRP and NLR levels were higher in non-surviving COVID-19 patients with DM comorbidity, indicating that they could serve as good predictors of poor outcomes in this patient population.

Fractions 14 and 36K of Metabolite Extract Streptomyces hygroscopicus subsp. Hygroscopicus Have Antimalarial Activities Against Plasmodium berghei in vitro.

Purpose: The study was conducted to investigate the effectivity and the cytotoxicity of fractions 14 and 36K of metabolite extract of Streptomyces hygroscopicus subsp. Hygroscopicus as an antimalarial compounds against Plasmodium berghei in vitro. Methods: Fractions 14 and 36K of metabolite extract of Streptomyces hygroscopicus subsp. Hygroscopicus produced by the fractionation process utilizing the Flash Column Chromatography (FCC) BUCHI Reveleris® PREP. Plasmodium berghei culture was used to assess the antimalarial activity of fractions 14 and 36K. Parasite densities and the ability of parasite growth were determined under microscopic. The cytotoxicity of the fractions was assessed using MTT assays on the MCF-7 cell line. Results: Streptomyces hygroscopicus subsp. Hygroscopicus fractions 14 and 36K have antimalarial activity against Plasmodium berghei, with fraction 14 having the more potent activity. The percentage of Plasmodium berghei-infected erythrocytes was decreased as well as the increase of fraction concentration. Fraction 14 has the highest inhibition of parasite growth at a concentration of 156,25 µg/mL, with an inhibition percentage of 67.73% (R2 = 0.953, p = 0.000). IC50 of fractions 14 and 36K were found at 10.63 µg/mL and 135,91 µg/mL, respectively. The fractions caused morphological damage in almost all asexual stages of the parasite. Both fractions were not toxic against MCF-7, indicating that the fractions have a safe active metabolite. Conclusion: Fractions 14 and 36K of metabolite extract Streptomyces hygroscopicus subsp. Hygroscopicus contains non-toxic compounds that could damage the morphology and inhibit the growth of Plasmodium berghei in vitro.

Antimalarial Drug Resistance: A Brief History of Its Spread in Indonesia.

Malaria remains to be a national and global challenge and priority, as stated in the strategic plan of the Indonesian Ministry of Health and Sustainable Development Goals. In Indonesia, it is targeted that malaria elimination can be achieved by 2030. Unfortunately, the development and spread of antimalarial resistance inflicts a significant risk to the national malaria control programs which can lead to increased malaria morbidity and mortality. In Indonesia, resistance to widely used antimalarial drugs has been reported in two human species, Plasmodium falciparum and Plasmodium vivax. With the exception of artemisinin, resistance has surfaced towards all classes of antimalarial drugs. Initially, chloroquine, sulfadoxine-pyrimethamine, and primaquine were the most widely used antimalarial drugs. Regrettably, improper use has supported the robust spread of their resistance. Chloroquine resistance was first reported in 1974, while sulfadoxine-pyrimethamine emerged in 1979. Twenty years later, most provinces had declared treatment failures of both drugs. Molecular epidemiology suggested that variations in pfmdr1 and pfcrt genes were associated with chloroquine resistance, while dhfr and dhps genes were correlated with sulfadoxine-pyrimethamine resistance. Additionally, G453W, V454C and E455K of pfk13 genes appeared to be early warning sign to artemisinin resistance. Here, we reported mechanisms of antimalarial drugs and their development of resistance. This insight could provide awareness toward designing future treatment guidelines and control programs in Indonesia.

Inhibition Effect of Physalis angulata Leaf Extract on Viability, Collagen Type I, and Tissue Inhibitor of Metalloproteinase 1 (TIMP-1) but Not Plasminogen Activator Inhibitor-1 (PAI-1) of Keloid Fibroblast Culture.

Introduction: Keloids are excessive fibroproliferative diseases that are caused by abnormal wound healing. The anti-proliferative activity of Physalis angulata compounds has potential as a keloid therapeutic agent. This study aimed to observe the effects of P. angulata on fibroblast viability and collagen type I, tissue inhibitor of metalloproteinase 1 (TIMP-1), and plasminogen activator inhibitor 1 (PAI-1) levels in human keloid fibroblasts. Methods: We conducted an experimental study of P. angulata ethanol extract on three primary human keloid fibroblast 3 passage cultures with four replications. Fibroblast viability was measured using the MTT assay after incubation with 3, 5, and 10 µg/mL P. angulata. Concentrations of P. angulata used to observe effects on TIMP-1, PAI-1, and collagen type I levels were 10%, 20%, 30%, and 40% of inhibitory concentration 50 (IC50). The levels of collagen type I, TIMP-1, and PAI-1 were measured by ELISA. Mean comparisons between multiple treatment groups were analyzed using one-way ANOVA followed by post-hoc analysis. Results: The 10 µg/mL P. angulata group had significantly lower fibroblast viability than the control group (p<0.05) with an IC50 6.3 µg/mL. The collagen type I level of 10% IC50 (0.63 µg/mL) P. angulata group was lower than control (12.910 vs 47.866 ng/mL) (p=0.042). Level of TIMP-1 in 40% IC50 (2.51 µg/mL) P. angulata group was lower than control (5.350 vs 9.972 ng/mL) (p=0.043). There was no significant difference in the PAI-1 levels. Conclusion: This study showed the inhibitory effect of 10 µg/mL P. angulata extract on keloid fibroblast viability, with an IC50 of 6.3 µg/mL. This study also showed collagen type-1 and TIMP-1 inhibition, but not PAI-1 inhibition, after P. angulate treatment.

Bifidobacterium longum Administration Diminishes Parasitemia and Inflammation During Plasmodium berghei Infection in Mice.

Purpose: During Plasmodium berghei (P. berghei) infection, infected erythrocytes are sequestered in gut tissues through microvascular circulation, leading to dysbiosis. This study aimed to investigate the effect of Lactobacillus casei (L. casei) and Bifidobacterium longum (B. longum) administration on the parasitemia level, gut microbiota composition, expression of cluster of differentiation 103 (CD103) in intestinal dendritic and T regulatory cells (T reg), plasma interferon gamma (IFN-γ) and tumor necrosis factor (TNF-α) levels in P. berghei infected mice. Methods: P. berghei was inoculated intraperitoneally. Infected mice were randomly divided into 5 groups and treated with either L. casei, B. longum, or the combination of both for 5 days before up to 6 days post-infection (p.i). The control group was treated with phosphate-buffered saline (PBS), while uninfected mice were used as negative control. Levels of CD103 and forkhead box P3 (FoxP3) expression were measured by direct immunofluorescense, while plasma IFN-γ and TNF-α level were determined using enzyme-linked immunosorbent assay (ELISA). Results: All treated groups showed an increase in parasitemia from day 2 to day 6 p.i, which was significant at day 2 p.i (p = 0.001), with the group receiving B. longum displaying the lowest degree of parasitemia. Significant reduction in plasma IFN-γ and TNF-α levels was observed in the group receiving B. longum (p = 0.022 and p = 0.026, respectively). The CD103 and FoxP3 expression was highest in the group receiving B. longum (p = 0.01 and p = 0.02, respectively). Conclusion: B. longum showed the best protective effect against Plasmodium infection by reducing the degree of parasitemia and modulating the gut immunity. This provides a basis for further research involving probiotic supplementation in immunity modulation of infectious diseases.

Peptide Sequence of Pili Subunit Protein 49.8 kDa Shigella flexneri as Antigenic Epitope for Shigellosis Vaccine Development.

Objectives: This study investigates the amino acid sequence and identifies antigenic epitopes of 49.8 kilodalton (kDa) pili protein Shigella flexneri, which will be used as candidates for the shigellosis vaccine. Materials and Methods: Our study is a prospectively descriptive laboratory. We used bacterial isolate of S. flexneri pili isolation was performed using a pili cutter and sodium dodecyl-sulfate polyacrylamide gel electrophoresis. The amino acid sequences were analyzed using liquid chromatography dual mass spectrometry (LC-MS/MS) method in the proteomic laboratory. The target epitope antigenicity analysis was tested using Kolaskar and Tongaonkar Antigenicity software. The Bepired Linear Epitope Prediction software is used for epitope mapping. PymOL software was used for the visualization of proteins and molecular docking. Peptides and antibodies were applied to hemagglutination test and immune response was tested using the dot blot method. Results: LC-MS/MS analysis results from the mascot server showed that the 49.8 kDa pili protein is S. flexneri similar to the flagellin protein of S. flexneri 1235-66 (ID I6H2T2). The results of antigenicity analysis and epitope mapping showed that areas of protein that has the most potential and antigenic epitopes are the regions 98-111 and 263-290 with the amino acid sequences, QSSTGTNSQSDLDS (Q-S) and DTTITKAETKTVTKNQVVDTPVTTDAAK (D-K). The results of the molecular docking interaction test between the peptide and the B-cell receptor have a low binding energy. Peptide Q-S and peptide D-K antigens are hemagglutinin molecules because they can agglutinate erythrocytes. The immune response between peptide antigens and anti-peptide antibodies can react based on color gradations in the dotblot method. Conclusion: The amino acid sequences Q-S and D-K are potentially antigenic epitopes. These peptides can be used to develop candidates for shigellosis vaccine.

Shigella flexneri vaccine development: Oral administration of peptides derived from the 49.8 kDa pili protein subunit activates the intestinal immune response in mice.

Background and Aim: The morbidity and mortality of Shigella infections remain a global challenge. Epitope-based vaccine development is an emerging strategy to prevent bacterial invasion. This study aimed to identify the ability of the 49.8 kDa pili subunit adhesin protein epitope of Shigella flexneri to induce an intestinal immune response in mice. Materials and Methods: Thirty adult male Balb/c mice were divided into a control group, cholera toxin B subunit (CTB) group, CTB+QSSTGTNSQSDLDS (pep_1) group, CTB+DTTITKAETKTVTKNQVVDTPVTTDAAK (pep_2) group, and CTB+ ATLGATLNRLDFNVNNK (pep_3). We performed immunization by orally administering 50 µg of antigen and 50 µl of adjuvant once a week over 4 weeks. We assessed the cellular immune response by quantifying T helper 2 (Th2) and Th17 using flow cytometry. In addition, we assessed the humoral immune response by quantifying interleukin (IL-4), IL-17, secretory immunoglobulin A (sIgA), and ß-defensin using enzyme-linked immunoassay. Statistical analysis was performed using one-way analysis of variance and Kruskal-Wallis test. Results: Peptide oral immunization increases the cellular immune response as reflected by the increase of Th2 (p=0.019) and Th17 (p=0.004) cell counts, particularly in the CTB_pep_1 group. Humoral immune response activation was demonstrated by increased IL-4 levels, especially in the CTB+pep_3 group (p=0.000). The IL-17 level was increased significantly in the CTB+pep_1 group (p=0.042). The mucosal immune response was demonstrated by the sIgA levels increase in the CTB+pep_3 group (p=0.042) and the ß-defensin protein levels (p=0.000). Conclusion: All selected peptides activated the cellular and humoral immune responses in the intestine of mice. Further studies are necessary to optimize antigen delivery and evaluate whether the neutralizing properties of these peptides allow them to prevent bacterial infection.

Human CD8+CXCR3+ T cells have the same function as murine CD8+CD122+ Treg.

The importance of CD8(+)CD122(+) Treg in the maintenance of immune homeostasis has been previously demonstrated in mice. Because the expression pattern of CD8 and CD122 in humans is different from that in mice, human CD8(+) Treg that correspond to the murine CD8(+)CD122(+) Treg have not been identified. In this study, we performed DNA microarray analyses to compare the gene expression profiles of CD8(+)CD122(+) cells and CD8(+)CD122(-) cells in mice and found that CXCR3 was preferentially expressed in CD8(+)CD122(+) cells. When we analyzed the expression of CD122 and CXCR3 in murine CD8(+) cells, we observed a definite population of CD122(+)CXCR3(+) cells. CD8(+)CXCR3(+) cells in mice showed similar regulatory activities to CD8(+)CD122(+) cells by in vivo and in vitro assays. While CD8(+)CD122(+)CXCR3(+) cells are present in mice, CD8(+)CXCR3(+) cells, but not CD8(+)CD122(+) cells, are present in humans. In the in vitro assay, human CD8(+)CXCR3(+) cells showed the regulatory activity of producing IL-10 and suppressing IFN-gamma production from CD8(+)CXCR3(-) cells. These results suggest that human CD8(+)CXCR3(+) T cells are the counterparts of murine CD8(+)CD122(+) Treg.

Identification of afzelin potential targets in inhibiting triple-negative breast cancer cell migration using reverse docking.

BACKGROUND: Triple-negative breast cancer (TNBC) tends to be aggressive and metastatic, characteristics attributable to its cellular migration capabilities. Afzelin is a chemical compound with anti-metastatic potentials. This study aimed to predict proteins involved in TNBC cell migration which could be inhibited by afzelin. METHODS: The protein database was constructed from the Kyoto Encyclopedia of Genes and Genomes pathways collection which related to cell motility, then screened for druggability using SuperTarget and Therapeutic Target Database. The involvement of druggable proteins in the TNBC metastasis process was investigated through existing publications in The National Center for Biotechnology Information PubMed database. Inhibitory potential of afzelin toward target proteins was compared to the proteins' known-inhibitor, using the reverse docking method. RESULTS: Ten proteins identified as potential targets of afzelin, with the top 3 being ERK2, KRas, and FAK, respectively. Afzelin's 3-O-rhamnoside group played a dominant role in forming hydrogen bonds with the target proteins. Further analysis with STRING suggested that afzelin might be able to inhibit chemotaxis and haptotaxis of TNBC cells. CONCLUSIONS: Afzelin was predicted to inhibit TNBC cell motility, by targeting ERK2, KRas, and FAK activation.

Exposure of 1800 MHz Radiofrequency with SAR 1,6 W/kg Caused a Significant Reduction in CD4+ T Cells and Release of Cytokines In-Vitro.

BACKGROUND: Although there have been many studies investigating the effects of electromagnetic fields on humans cells and tissues, the effects of radiofrequency electromagnetic fields exposure on the cells of the immune system are still controversial. OBJECTIVE: To investigate the effects of 1800 MHz RF-EMF exposure on peripheral blood mononuclear cells by measuring T helper cells count and the cytokine profile under different conditions of durations and distances. METHODS: The peripheral blood mononuclear cells (PBMCs) from healthy human subjects were exposed to 1800 MHz RF-EMF, with durations of 15, 30, 45, and 60 minutes and distances of 5 and 25 cm. The effects of RF-EMF exposure on the number of CD4+ T cells, and the expression of IL-2, IL-10, and IL-17a after 48 hours of culture were evaluated using flow cytometry. RESULTS: Our findings indicated that closer distance and longer exposure inducedlower number of CD4+ T cells. Similarly the percentagesof IL-2, IL-10 and IL-17a expressing CD4+ T cells weredecreased significantly. The number of IL-2 expressing CD4+T cells wasincreased significantly as the duration of exposure was increased, but the number was decreased after 60 minutes exposure when compared with control group with no exposure. CONCLUSION: Exposure to RF-EMF for 60 minutes at 5 cm distance causes a significant reduction in the number of CD4+ T cells, IL-2, IL-10 and IL-17a expressing T cells.

Effect of Metabolite Extract of Streptomyces hygroscopicus subsp. hygroscopicus on Plasmodium falciparum 3D7 in Vitro.

BACKGROUND: Malaria eradication has been complicated by the repeated emergence of antimalarial drug resistances. We aimed to determine whether a metabolite extract of Streptomyces hygrocopicus subsp. hygroscopicus could decrease the viability of Plasmodium falciparum 3D7 in vitro. METHODS: S. hygroscopicus subsp. hygroscopicus isolates were inoculated and fermented on the ISP4 medium. The fermented S. hygroscopicus was mixed with ethylacetate 1:5 (v/v), and the solvent phase was evaporated. Several concentrations of isolated extract was added to the P. falciparum 3D7 culture containing trophozoite and schizont stages in 24 wells plates when the degree of parasite-infected erythrocytes reached 5%, then incubated for 8 hours. DNA parasite density was measured using flow cytometry, parasite degree and morphology were observed under microscopic by Giemsa-stained smears. RESULTS: The metabolite extract affected the morphology of almost all of parasite asexual stages. Schizonts and trophozoites failed to grow and appeared damaged with pycnotic cores and loss of cytoplasmic content. At 8 hours there was a significant decrease in DNA parasite density in culture exposed to 2.6 mg/ml and 13 mg/ml (P = 0.002; P = 0.024) of the extract. The degree of parasite-infected erythrocytes was decreased from the beginning of exposure (0.02 mg/ml of the extract). There was a significant inverse correlation between the concentration of extract and the degree of parasite-infected erythrocytes as well as the density of DNA parasite (r = -0.772, P = 0.000; r =-0.753; P =0.000). CONCLUSION: Metabolite extract of S. hygroscopicus subsp. hygroscopicus causes morphological damage, decreases the degree of parasite-infected erythrocytes and the DNA density of P. falciparum 3D7 in vitro.