Development and characterization of SARS-CoV-2 variant-neutralizing monoclonal antibodies.

Vaccination and administration of monoclonal antibody cocktails are effective tools to control the progression of infectious diseases and to terminate pandemics such as COVID-19. However, the emergence of SARS-CoV-2 mutants with enhanced transmissibility and altered antigenicity requires broad-spectrum therapies. Here we developed a panel of SARS-CoV-2 specific mouse monoclonal antibodies (mAbs), and characterized them based on ELISA, Western immunoblot, isotyping, and virus neutralization. Six neutralizing mAbs that exhibited high-affinity binding to SARS-CoV-2 spike protein were identified, and their amino acid sequences were determined by mass spectrometry. Functional assays confirmed that three mAbs, F461G11, F461G15, and F461G16 neutralized four variants of concern (VOC): B.1.1.7 (alpha), B.1.351 (beta), P.1 (gamma) and B.1.617.2 (delta) These mAbs are promising candidates for COVID-19 therapy, and understanding their interactions with virus spike protein should support further vaccine and antibody development.

Novel anoplin-based (lipo)-peptide models show potent antimicrobial activity.

The subject of this study is the synthesis and biological evaluation of anoplin-based (Gly-Leu-Leu3 -Lys-Arg5 -Ile-Lys-Thr8 -Leu-Leu-NH2 )-designed (lipo)-peptides, aiming at the development of new antibiotic substances. The design of synthetic compounds based on natural bioactive molecules is an optimistic strategy for the development of new pharmaceutics. Antimicrobial peptides (AMPs) and (lipo)-peptides are two classes of promising compounds, with characteristics that allow them to express their activity by differentiated mechanisms of action. On this basis, anoplin, a natural AMP, was used as a scaffold to design five peptides and seven lipopeptide analogs of them. Substitutions were made on residues Leu3 and Arg5 of the interphase and on Thr8 of the polar phase, as well as N-terminus conjunctions with octanoic and decanoic acid. The outcome of the biological evaluation revealed that some analogs might have substantial clinical potential. Specifically, Ano 1-F, Ano 3-F, Ano 4-C10 , and Ano 5-F are strongly active against Gram-negative bacteria at minimum inhibitory concentration (MIC) values of 3 µg/ml, while Ano 4-F is active against Gram-positive bacteria at 1 µg/ml. Ano 2-C10 , C10 -Gly-Leu-Lys3 -Lys-Ile5 -Ile-Lys-Lys8 -Leu-Leu-NH2 , is the most promising compound (MIC = 0.5 µg/ml) for the development of new pharmaceutics. The conformational features of the synthetic peptides were investigated by circular dichroism spectroscopy, and their physicochemical parameters were calculated. Our study shows that appropriate substitutions in the anoplin sequence in combination with Nα -acylation may lead to new effective AMPs.

Efficacy and Safety of TurboHawk Plaque Rotation Combined With Drug-coated Balloon in Treating Diabetic Patients With Lower Extremity Arterial Disease.

To evaluate the efficacy and safety of TurboHawk plaque rotation system combined with drug-coated balloon in treating lower extremity arterial disease (LEAD) of diabetes patients, a total of 145 diabetic patients with LEAD from March 2015 to September 2016 were recruited in our study. Lower extremity arterial disease was diagnosed by ultrasound and CT angiography (CTA). According to the surgical method, 65 cases underwent TurboHawk plaque rotation combined with drug-coated balloon (group A), 80 cases underwent simple drug-coated balloon expansion (group B). The characteristics of lesion, function test, ankle-brachial index (ABI), and postoperative complications were analyzed. All the patients were followed up at 1, 3, 6, 12, and 24 months after operation. At baseline, there was no difference in all the characteristics between the 2 groups. The early postoperation minimum lumen diameter (MLD), lumen stenosis rate, and ABI in 2 groups both improved. As the follow-up time increased, patients in group A had significantly higher MLD and ABL value, as well as lower level of lumen stenosis rate, restenosis rate, late lumen loss, and target lesion revascularization (all P < .05). Accordingly, functional testing revealed the 6-minute walk distance, 6-minute claudication distance, resting ABI, and post-exercise ABI in group A were significantly higher than those in group B (all P < .05). Besides, major graft reintervention (4.62% vs 11.25%) and major adverse limb events (6.15% vs 12.5%) in group A occurred less frequently than group B (all P < .05). In conclusion, the long-term effect of the combined approach was better than only drug-coated balloon in LEAD in Chinese diabetes patients.

Dependence on size and shape of non-nature amino acids in the enhancement of lipopolysaccharide (LPS) neutralizing activities of antimicrobial peptides.

HYPOTHESIS: Release of lipopolysaccharides (LPS) from bacteria into bloodstream may cause serious unwanted stimulation of the host immune system. P-113 is a clinically active histidine-rich antimicrobial peptide. Nal-P-113, a ß-naphthylalanine-substituted P-113, is salt-resistant but has limited LPS neutralizing activity. We suspected the size and shape of the non-natural bulky amino acid may affect its LPS neutralizing activity. Herein, antimicrobial, LPS neutralizing, and antiproteolytic effects of phenylalanine- (Phe-P-113), ß-naphthylalanine- (Nal-P-113), ß-diphenylalanine- (Dip-P-113), and ß-(4,4'-biphenyl)alanine- (Bip-P-113) substituted P-113 were studied. EXPERIMENTS: Structure-activity relationships of P-113, Phe-P-113, Nal-P-113, Dip-P-113, and Bip-P-113 were evaluated using antimicrobial activity assays, serum proteolytic assays, peptide-induced permeabilization of large unilamellar vesicles, zeta potential measurements, dynamic light scattering measurement of LPS aggregation, and Limulus amebocyte lysate assays for measuring LPS neutralization. In vitro and in vivo LPS neutralizing activities were further confirmed by LPS-induced inflammation inhibition in an endotoxemia mouse model. FINDINGS: Bip-P-113 and Dip-P-113 had the longest and widest non-nature amino acids, respectively. Bip-P-113 enhanced salt resistance, serum proteolytic stability, peptide-induced permeabilization, zeta potential measurements, LPS aggregation, and in vitro and in vivo LPS neutralizing activities. These results could help design novel antimicrobial peptides that have enhanced stability in vivo and that can have potential therapeutic applications.

Evaluación entre dos tecnologías usando Técnicas de Aglutinación en Columna (TAC) para la titulación e identificación de aloanticuerpos anti-eritrocitarios (columnas de micro-esferas de cristal y columnas de gel) / Evaluation between two technologies using Column Agglutination Techniques (TAC) for the titration and identification of anti-erythrocytic alloantibodies (columns of glass microspheres and gel columns)

Introducción: La presencia de anticuerpos anti eritrocitarios disminuye la sobrevida de los eritrocitos mediante la presencia de enfermedad hemolítica post transfusional en los pacientes oncológicos. La selección de la tecnología más conveniente para la determinación de los anticuerpos antieritrocitarios está directamente relacionada con la prevención de las reacciones transfusionales hemolíticas que son las responsables de la morbilidad y mortalidad relacionadas con la transfusión. El objetivo del presente estudio fue valorar la asociación estadística entre la técnica tradicional del gel versus la técnica de microesferas de cristal. Métodos: El estudio fue realizado en el laboratorio Clínico del Instituto Oncológico Nacional Dr. Juan Tanca Marengo, Solca-Guayaquil el período de estudio enero 2017 a marzo del 2017. Se realizaron técnicas de titulación a partir de muestras de sangre periférica con aloanticuerpos anti-eritrocitarios de especificidad única que habían sido detectados durante la realización de las pruebas pre-transfusionales. Las muestras fueron procesadas con la tecnología conocida usando la Técnica de Aglutinación en Columnas, columnas con gel. Posterior a realizar las pruebas pretransfusionales en los casos que presentaran positiva la prueba de escrutinio de anticuerpos o test de Coombs indirecto, inmediatamente después se llevaba a cabo la identificación del anticuerpo usando la misma técnica.Simultáneamente se realizaba las mismas pruebas usando la misma muestra con la otra tecnología de microesferas cristalizadas. Las reacciones de aglutinación se clasificaron como fuertemente positivas (4+ y 3+), moderadamente positivas (2+ y 1+) y positivo débil (w+). El análisis estadístico se utilizó correlación de Spearman. Resultados: En el período de estudio se efectuaron detección e identificación en 9 muestras. Seis muestras mostraron títulos idénticos, para ambas tecnologías, en 2 muestras se mostró títulos más altos con la tecnología en gel, y en 1 muestra se observó títulos más altos con la tecnología en micro esferas de cristal. La asociación Mediante Rho de Spearman entre las dos pruebas fue de R=0.84; r2=0.72, P=0.005. Conclusión: Existen una buena asociación entre las técnicas de gel y microesferas de cristal para la identificación de anticuerpos antieritrocitarios.

Introduction: The presence of anti-erythrocyte antibodies reduces the survival of erythrocytes by the presence of post-transfusion hemolytic disease in cancer patients. The selection of the most convenient technology for the determination of antierithrocytic antibodies is directly related to the prevention of hemolytic transfusion reactions that are responsible for the morbidity and mortality related to transfusion. The aim of the present study was to evaluate the statistical association between the traditional gel technique versus the crystal microsphere technique. Methods: The study was conducted in the Clinical Laboratory of the Dr. Juan Tanca Marengo National Oncological Institute, Solca-Guayaquil during the study period January 2017 to March 2017. Titration techniques were performed from peripheral blood samples with anti-alloantibodies-erythrocytes of unique specificity that had been detected during the pre-transfusion tests. The samples were processed with the known technology using the Agglutination Technique in Columns, columns with gel. After carrying out the pretransfusion tests in cases that presented a positive antibody test or indirect Coombs test, the antibody was immediately identified using the same technique. Simultaneously, the same tests were performed using the same sample with the other crystallized microsphere technology. The agglutination reactions were classified as strongly positive (4+ and 3+), moderately positive (2+ and 1+) and weak positive (w +). The statistical analysis was used Spearman's correlation. Results: In the study period, detection and identification were made in 9 samples. Six samples showed identical titers, for both technologies, 2 samples showed higher titers with gel technology, and in 1 sample higher titers were observed with the technology in crystal micro spheres. The association by Rho of Spearman between the two tests was R = 0.84; r2 = 0.72, P = 0.005. Conclusion: There is a good association between gel techniques and crystal microspheres for the identification of antierithrocytic antibodies.

Effects of topical exposure to a mixture of chlorpyrifos, cypermethrin and captan on the hematological and immunological systems in male Wistar rats.

Most Thai orchid farmers heavily used pesticide mixtures, and were shown to have various hematologic/immunologic alterations. The present study investigated the effect of exposure of male Wistar rats to a mixture of three pesticides (chlorpyrifos, cypermethrin and captan) that are most often used by the farmers. Three groups of 10 rats were dermally exposed to three different doses (high, middle and low) for 28 consecutive days. The rats showed significant changes in body, liver, kidneys and adrenals weights. Significant changes were observed in various biological parameters including hematotoxicity (increased leukocyte and platelet counts, percent neutrophil, decreased RBC count, percent lymphocyte and eosinophil), hepatotoxicity (increased serum AST, decreased serum ALP, cholesterol, triglyceride, serum protein and albumin), and immunotoxicity (decrease in numbers of NK cells, decrease splenic proliferative response to LPS, and increase in serum IgG). These results confirm the potential health danger of exposure to these pesticide mixtures in orchid farmers.

Developmental immunotoxicity is not associated with the consumption of transgenic Bt rice TT51 in rats.

TT51 is a transgenic strain of Bt rice generated by fusing a synthetic CryAb/Ac gene into MingHui rice. In this study, rats from F0, F1, and F2 generations were fed a diet with 60% TT51 rice, MingHui rice, or nominal-origin rice. The study focused on developmental immunotoxicity in F1 and F2 offspring after long-term consumption of TT51. A wide range of immunological parameters was monitored in this two-generation study on reproductive toxicity. The experiments were performed on F1 and F2 offspring at postnatal days 21 and 42. No adverse clinical effects were observed in any of the experimental groups. In addition, histopathology observations and immunotoxicity tests, including hematological indicators, spleen lymphocyte subsets, natural killer cell activity, lymphoproliferative response, and plaque-forming cell assay, revealed no significant difference between the groups. These results indicated that developmental immunotoxicity was not associated with a diet of transgenic Bt rice TT51, compared to the parental MingHui rice.

ß Cell Aging Markers Have Heterogeneous Distribution and Are Induced by Insulin Resistance.

We hypothesized that the known heterogeneity of pancreatic ß cells was due to subpopulations of ß cells at different stages of their life cycle with different functional capacities and that further changes occur with metabolic stress and aging. We identified new markers of aging in ß cells, including IGF1R. In ß cells IGF1R expression correlated with age, dysfunction, and expression of known age markers p16ink4a, p53BP1, and senescence-associated ß-galactosidase. The new markers showed striking heterogeneity both within and between islets in both mouse and human pancreas. Acute induction of insulin resistance with an insulin receptor antagonist or chronic ER stress resulted in increased expression of aging markers, providing insight into how metabolic stress might accelerate dysfunction and decline of ß cells. These novel findings about ß cell and islet heterogeneity, and how they change with age, open up an entirely new set of questions about the pathogenesis of type 2 diabetes.

Sickle cell disease.

Sickle cell disease is a common and life-threatening haematological disorder that affects millions of people worldwide. Abnormal sickle-shaped erythrocytes disrupt blood flow in small vessels, and this vaso-occlusion leads to distal tissue ischaemia and inflammation, with symptoms defining the acute painful sickle-cell crisis. Repeated sickling and ongoing haemolytic anaemia, even when subclinical, lead to parenchymal injury and chronic organ damage, causing substantial morbidity and early mortality. Currently available treatments are limited to transfusions and hydroxycarbamide, although stem cell transplantation might be a potentially curative therapy. Several new therapeutic options are in development, including gene therapy and gene editing. Recent advances include systematic universal screening for stroke risk, improved management of iron overload using oral chelators and non-invasive MRI measurements, and point-of-care diagnostic devices. Controversies include the role of haemolysis in sickle cell disease pathophysiology, optimal management of pregnancy, and strategies to prevent cerebrovascular disease.

Functional and structural characterization of four mouse monoclonal antibodies to complement C3 with potential therapeutic and diagnostic applications.

C3 is the central component of the complement system. Upon activation, C3 sequentially generates various proteolytic fragments, C3a, C3b, iC3b, C3dg, each of them exposing novel surfaces, which are sites of interaction with other proteins. C3 and its fragments are therapeutic targets and markers of complement activation. We report the structural and functional characterization of four monoclonal antibodies (mAbs) generated by immunizing C3-deficient mice with a mixture of human C3b, iC3b and C3dg fragments, and discuss their potential applications. This collection includes three mAbs interacting with native C3 and inhibiting AP complement activation; two of them by blocking the cleavage of C3 by the AP C3-converase and one by impeding formation of the AP C3-convertase. The interaction sites of these mAbs in the target molecules were determined by resolving the structures of Fab fragments bound to C3b and/or iC3b using electron microscopy. A fourth mAb specifically recognizes the iC3b, C3dg, and C3d fragments. It binds to an evolutionary-conserved neoepitope generated after C3b cleavage by FI, detecting iC3b/C3dg deposition over opsonized surfaces by flow cytometry and immunohistochemistry in human and other species. Because well-characterized anti-complement mAbs are uncommon, the mAbs reported here may offer interesting therapeutic and diagnostic opportunities.

The effect of ethephon on immune system in male offspring of mice.

Ethephon can liberate ethylene which could interfere the plant growth process. The aim of the present study was to determine the effect of ethephon on developing immune system of male offspring. Ethephon could enhance NK cell activity in male mice. For 4-week-old male mice, lymphocytes of peripheral blood increased while the hemolytic plaque number decreased. Delayed type hypersensitivity(DTH) was inhibited in all groups. The expression of protein Bcl11b and p-p38 in thymus of treatment groups were lower than control group. Our results indicated that cellular immunity of male offspring is more sensitive to ethephon when exposed in pregnancy and lactation period. It should be emphasized that exposure to ethephon during the in utero stage and lactation stage still could damage the immune function of animal in the period before fully mature even in the dosage that could not influence the immune function of adult animal.

Effects of chelidonic acid, a secondary plant metabolite, on mast cell degranulation and adaptive immunity in rats.

The present study evaluated the immunomodulatory effects of chelidonic acid, a secondary plant metabolite, with therapeutic potential in allergic disorders, in experimental animals. In mast cell degranulation studies, ovalbumin immunized and challenged rats, chelidonic acid (1, 3 and 10mg/kg, i.p.) dose relatedly prevented ovalbumin challenge induced mast cell degranulation by differing degrees when compared with vehicle treated group, and these effects were comparable with prednisolone (10mg/kg). A reduction in post-challenge mortality was also observed in all treated groups. Further, there were reductions in the blood eosinophil counts and serum IgE levels after chelidonic acid treatment. Chelidonic acid also inhibited histamine release from rat peritoneal mast cells (RPMC) in vitro, in a dose related manner. In tests for adaptive immunity, in rats immunized with sheep RBC, chelidonic acid differentially suppressed the (a) plaque forming cell (PFC) count in rat splenic cells, (b) anti-SRBC antibody titre and serum IgG levels and (c) increases in foot pad thickness in the DTH assay - all of which were comparable with prednisolone. These experimental results are discussed in light of the possible therapeutic potential of chelidonic acid in allergic disorders.

Comparison of the Hemagglutination Inhibition Test and IgG ELISA in Categorizing Primary and Secondary Dengue Infections Based on the Plaque Reduction Neutralization Test.

Secondary dengue infection by heterotypic serotypes is associated with severe manifestations of disease, that is, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). The World Health Organization (WHO) has recommended criteria based on the hemagglutination inhibition (HI) test to distinguish between primary and secondary dengue infections. Since the HI test has practical limitations and disadvantages, we evaluated the accuracy of WHO HI criteria and compared it with criteria based on an IgG enzyme-linked immunosorbent assay (ELISA) using a plaque reduction neutralization test (PRNT) as the gold standard. Both WHO HI criteria and IgG ELISA criteria performed strongly (16/16) in determining primary infection. However, to determine secondary infection, the IgG ELISA criteria performed better (72/73) compared to the WHO HI criteria (23/73).

Development and Application of a Simple Plaque Assay for the Human Malaria Parasite Plasmodium falciparum.

Malaria is caused by an obligate intracellular protozoan parasite that replicates within and destroys erythrocytes. Asexual blood stages of the causative agent of the most virulent form of human malaria, Plasmodium falciparum, can be cultivated indefinitely in vitro in human erythrocytes, facilitating experimental analysis of parasite cell biology, biochemistry and genetics. However, efforts to improve understanding of the basic biology of this important pathogen and to develop urgently required new antimalarial drugs and vaccines, suffer from a paucity of basic research tools. This includes a simple means of quantifying the effects of drugs, antibodies and gene modifications on parasite fitness and replication rates. Here we describe the development and validation of an extremely simple, robust plaque assay that can be used to visualise parasite replication and resulting host erythrocyte destruction at the level of clonal parasite populations. We demonstrate applications of the plaque assay by using it for the phenotypic characterisation of two P. falciparum conditional mutants displaying reduced fitness in vitro.

Modulating Effects of Arabinogalactans from Plant Gum Exudates on Human Complement System.

Gum arabic and cashew nut tree gum exudate polysaccharide (CNTG) are plant polysaccharides composed of galactose and arabinose known as arabinogalactans (AGs). Although these fractions are used in food and pharmaceutical industry, cases of allergic reactions were described in clinical reports. As AGs were reported as modulators of the classical (CP) and alternative pathways (AP) of complement system (CS), in the present work, we investigate whether gum arabic and CNTG have an effect on both CS pathways. The complement fixation tests were performed with (CP-30 and AP-30) and without pre-incubation (CP-0 and AP-0). For CP-30, CNTG and gum arabic (833 µg/ml) showed a reduction of 28.0% (P = 0.000174) and 48.5% (P = 0.000143), respectively, on CP-induced haemolysis. However, no effect was observed for CP-0 in the CP-induced haemolysis. For AP-30, both CNTG and gum arabic (833 µg/ml) showed 87% reduction on the CP-induced haemolysis, with IC50 values of 100 and 7 µg/ml, respectively. For AP-0, a reduction of 11.3% for gum arabic and no effect for the CNTG on the CP-induced haemolysis were observed. These results suggested that gum arabic and CNTG could be acting as activators of the CS. Thus, this effect on the CS, especially on the AP, which accounts for up to 80-90% of total CS activation, indicates that both fractions may be harmful because of their potential pro-inflammatory action. Considering that CS activation induces inflammatory response, further studies confirming this immunomodulatory effect of these fractions are required to insure their safe use.

A randomised double-blind clinical trial of two yellow fever vaccines prepared with substrains 17DD and 17D-213/77 in children nine-23 months old.

This randomised, double-blind, multicentre study with children nine-23 months old evaluated the immunogenicity of yellow fever (YF) vaccines prepared with substrains 17DD and 17D-213/77. YF antibodies were titered before and 30 or more days after vaccination. Seropositivity and seroconversion were analysed according to the maternal serological status and the collaborating centre. A total of 1,966 children were randomised in the municipalities of the states of Mato Grosso do Sul, Minas Gerais and São Paulo and blood samples were collected from 1,714 mothers. Seropositivity was observed in 78.6% of mothers and 8.9% of children before vaccination. After vaccination, seropositivity rates of 81.9% and 83.2%, seroconversion rates of 84.8% and 85.8% and rates of a four-fold increase over the pre-vaccination titre of 77.6% and 81.8% were observed in the 17D-213/77 and 17DD subgroups, respectively. There was no association with maternal immunity. Among children aged 12 months or older, the seroconversion rates of 69% were associated with concomitant vaccination against measles, mumps and rubella. The data were not conclusive regarding the interference of maternal immunity in the immune response to the YF vaccine, but they suggest interference from other vaccines. The failures in seroconversion after vaccination support the recommendation of a booster dose in children within 10 years of the first dose.

A randomised double-blind clinical trial of two yellow fever vaccines prepared with substrains 17DD and 17D-213/77 in children nine-23 months old

This randomised, double-blind, multicentre study with children nine-23 months old evaluated the immunogenicity of yellow fever (YF) vaccines prepared with substrains 17DD and 17D-213/77. YF antibodies were tittered before and 30 or more days after vaccination. Seropositivity and seroconversion were analysed according to the maternal serological status and the collaborating centre. A total of 1,966 children were randomised in the municipalities of the states of Mato Grosso do Sul, Minas Gerais and São Paulo and blood samples were collected from 1,714 mothers. Seropositivity was observed in 78.6% of mothers and 8.9% of children before vaccination. After vaccination, seropositivity rates of 81.9% and 83.2%, seroconversion rates of 84.8% and 85.8% and rates of a four-fold increase over the pre-vaccination titre of 77.6% and 81.8% were observed in the 17D-213/77 and 17DD subgroups, respectively. There was no association with maternal immunity. Among children aged 12 months or older, the seroconversion rates of 69% were associated with concomitant vaccination against measles, mumps and rubella. The data were not conclusive regarding the interference of maternal immunity in the immune response to the YF vaccine, but they suggest interference from other vaccines. The failures in seroconversion after vaccination support the recommendation of a booster dose in children within 10 years of the first dose.

Validation of Single Radial Haemolysis assay: A reliable method to measure antibodies against influenza viruses.

The Single Radial Haemolysis (SRH) assay is a serological method widely used for measuring antibodies against influenza viruses. Despite the broad application and recommendation by licensing authorities, the SRH assay has not been standardized. The aim of this study is to demonstrate how the SRH assay satisfies validation parameters of regulatory agencies in terms of specificity, precision, repeatability, intermediate precision, linearity, accuracy and robustness. This study shows that the SRH is a rapid, simple, reliable and reproducible assay, which requires only small volumes of serum samples and can be easily standardized.

Trikatu, an herbal compound as immunomodulatory and anti-inflammatory agent in the treatment of rheumatoid arthritis--an experimental study.

In the present study, trikatu, an herbal compound was evaluated for its immunomodulatory and anti-inflammatory properties with reference to cell mediated immune responses (delayed type hypersensitivity reaction), humoral immune response (haemagglutination titer and plaque forming assay), macrophage phagocytic index, circulating immune complex and inflammatory mediators in rats. For comparison purposes, indomethacin was used as a reference drug for anti-inflammatory studies. The results obtained in our study showed a significant decrease in cell mediated immune responses, humoral immune responses (haemagglutination titre and plaque forming assay) and macrophage phagocytic index in trikatu treated rats (1000 mg/kg/b.wt.) compared to control animals implying its immunosuppressive property. In addition, significant anti-inflammatory effects were observed in trikatu treated adjuvant induced arthritic rats by a reduction in the levels of circulating immune complexes and inflammatory mediators (TNF-alpha and Interleukin-1beta). Thus, in conclusion, our data suggest that trikatu could be considered as a potential anti-inflammatory agent for treating autoimmune inflammatory disorders like rheumatoid arthritis with immunosuppressive property.