Poly-γ-glutamylation of biomolecules.

Poly-γ-glutamate tails are a distinctive feature of archaeal, bacterial, and eukaryotic cofactors, including the folates and F420. Despite decades of research, key mechanistic questions remain as to how enzymes successively add glutamates to poly-γ-glutamate chains while maintaining cofactor specificity. Here, we show how poly-γ-glutamylation of folate and F420 by folylpolyglutamate synthases and γ-glutamyl ligases, non-homologous enzymes, occurs via processive addition of L-glutamate onto growing γ-glutamyl chain termini. We further reveal structural snapshots of the archaeal γ-glutamyl ligase (CofE) in action, crucially including a bulged-chain product that shows how the cofactor is retained while successive glutamates are added to the chain terminus. This bulging substrate model of processive poly-γ-glutamylation by terminal extension is arguably ubiquitous in such biopolymerisation reactions, including addition to folates, and demonstrates convergent evolution in diverse species from archaea to humans.

Sustainability assessment of biofuel and value-added product from organic fraction of municipal solid waste.

The current study aims to examine the techno-economic and environmental assessment of biorefinery development within a circular bioeconomy context by using an organic fraction of municipal solid waste (OFMSW) by extraction of lipids, carbohydrates, and proteins with 98, 51 and 62 % by mass of conversion efficiency and yield recovery, and value-added fractions production as well. Fatty acid methyl ester (biodiesel) and glycerol (biofuel) were produced by applying transesterification process, and the remaining biomass was converted into biocrude oil by thermal liquefication. The biorefinery using 613 kg of OFMSW produced biodiesel, glycerol, and bioethanol with 126 litter, 14.3 kg, and 172 litter respectively, as well as value-added fractions, such as biocrude oil with 78 kg. The environmental impact was assessed using the life cycle assessment (LCA) framework, ReCiPe2016 Mid-point (H) approach, through 18 different environmental categories. The key findings elucidate that Terrestrial ecotoxicity, Climate change, Fossil depletion and Human toxicity were the main impact categories which are potentially contributed 9.81E+02 kg 1,4-DB eq., 1.43E+03 kg CO2 eq., 2.04E+02 kg oil eq., and 8.08E+01 kg 1,4-DB eq. The normalization (person per equivalent) analysis revealed that only categories of resource reduction (fossil and metal depletion) are the key contributors to environmental degradation. The biorefinery system's total revenue was estimated at USD 6.817,509 million annually. The calculated revenue was USD 0.026 million daily in a shift of 8 h. The Net present worth (NPW) was calculated at USD 499.97 million by assuming a discount factor of 10 % and inflation rate of 5 % for 15 years. The project is considered feasible by demonstrating 7.15 payback year. This research showcased the efficient portrayal of the biorefinery system and succinctly conveyed the significant circular bioeconomy for a greener future. Thus, it could be helpful to the stakeholder's context towards techno-economic and environmental evaluation.

Green loops and clean skies: Optimizing municipal solid waste management using data science for a circular economy.

The interplay between Municipal Solid Waste (MSW) Management and data science unveils a panorama of opportunities and challenges, set against the backdrop of rising global waste and evolving technological landscapes. This article threads through the multifaceted aspects of incorporating data science into MSW management, unearthing key findings, novel knowledge, and instigating a call to action for stakeholders (e.g. policymakers, local authorities, waste management professionals, technology developers, and the general public) across the spectrum. Predominant challenges like the enigmatic nature of "black-box" models and tangible knowledge gaps in the sector are scrutinized, ushering in a narrative that emphasizes transparent, stakeholder-inclusive, and policy-adaptive approaches. Notably, a conscious shift towards "white-box" and "grey-box" data science models has been spotlighted as a pivotal response to transparency issues. Furthermore, the discourse highlights the necessity of crafting data science solutions that are specifically moulded to the nuanced challenges of MSW management, and it underscores the importance of recalibrating existing policies to be reflexive to technological advancements. A resolute call echoes for stakeholders to not just adapt but immerse themselves in a continuous learning trajectory, championing transparency, and fostering collaborations that hinge on innovative, data-driven methodologies. Thus, as the realms of data science and MSW management entwine, the article sheds light on the potential transformation awaiting waste management paradigms, contingent on the nurtured amalgamation of technological advances, policy alignment, and collaborative synergy.

Empirical analysis of cost-effective and equitable solid waste management systems: Environmental and economic perspectives.

The solid waste management (SWM) system is in a transitional phase in developing economies, and local municipalities and waste management companies are stepping toward integrating a waste treatment approach in the scheme of waste handling. However, there is an urgent need to explore cost-effective techniques, models, and potential revenue streams to sustain the state-run waste sector self-sufficiently. The proposed SWM model aims to support the local waste sector in Islamabad, the capital city of Pakistan, with 100% service area coverage to attain environmental and economic sustainability by defining dedicated waste collection streams to ensure quality material recovery under a cost-effective approach and modality. The innovative approach is applied to allocate the tonnage to various streams as per the city's current land use plan. The estimated/cost of the cleanliness services will be USD13.1 million per annum with an estimated per ton cost of USD 23. The establishment of the proposed material recovery facility (MRF) will process about 500 t/d of waste to produce 45 t/d compost and recover 130 t/d of recyclables. The environmentally friendly model saves 2.4 million tons of CO2‒eq/month from composting and recycling. The average economic potential from MRF and debris-crushing plants, including environmental benefit value, is calculated as USD 3.97 million annually. Recovery of services fee (70%) for various collection streams based on city land use and socio-economic conditions will generate revenue of USD 7.33 million annually. The total revenue will be USD 11.31 million (86% of total annual expenditures) to track the sector's self-sufficiency. To successfully reach the Sustainable Development Goals (SDGs) and Nationally Determined Contributions (NDCs), engaging the private sector from environmentally advanced economies to collaborate in the waste sector to enhance local technical capabilities is recommended.

Unveiling Scapular Pseudo-winging: A Case Report on Ventral Osteochondroma of the Scapula in a 21-year-old.

Introduction: Scapular pseudo-winging caused by ventral osteochondromas is a rare condition that has been reported in only a handful of cases. This case report describes a 21-year-old male patient with scapular pseudo-winging due to ventral osteochondromas of the scapula. This report adds to the limited literature on this topic and highlights the importance of considering ventral osteochondromas as a possible cause of scapular pseudo-winging. Case Report: A 21-year-old male presented with a swelling on his left upper back that had gradually increased in size over the past 10 years. Physical examination revealed a bony hard swelling arising from the medial border of the scapula, with associated scapular pseudo-winging. Diagnostic imaging confirmed the presence of two ventral osteochondromas located on the body of the scapula. Surgical excision was performed to address the patient's cosmetic concerns. Conclusion: This case report highlights the importance of considering ventral osteochondromas as a possible cause of scapular pseudo-winging. It contributes to the existing literature by documenting a rare presentation and providing insights into the clinical course, diagnostic imaging, and surgical management of this condition. By raising awareness among tumor surgeons and orthopedic surgeons, this report may aid in early recognition and appropriate management of scapular pseudo-winging cases caused by ventral osteochondromas. Furthermore, this report expands our understanding of the etiology and treatment options for scapular pseudo-winging, potentially benefiting patients across various clinical specialties. Categories: Oncology, orthopedics.

Isolated Tillaux Fracture in a Skeletally-Mature Patient: A Rare Presentation.

Isolated Tillaux fractures are uncommon injuries that occur due to external rotational forces acting on the ankle joint. They are more commonly seen in adolescents due to the presence of open epiphyses. In adults, isolated Tillaux fractures present as plafond fractures well described with the AO (Arbeitsgemeinschaft für Osteosynthesefragen) Classification as well as Types 1, 2, and 3 according to the degree of articular comminution present. They occur rarely and can be easily missed or misdiagnosed as other ankle injuries. A thorough clinical examination, combined with imaging studies such as X-rays and CT scans, can aid in accurate diagnosis and treatment planning. Management typically involves open reduction and internal fixation, followed by a short period of immobilization and early mobilization with non-weight bearing. We present a case report of a 27-year-old female who presented with an isolated Tillaux fracture of the ankle following a road traffic accident. This type of fracture is typically seen in teenagers and young adults due to the incomplete closure of the growth plate, which makes it more susceptible to injury. The patient underwent open reduction and internal fixation with a contoured three-hole 3.5 mm titanium T-plate, followed by immobilization in a plaster splint for one week. Early mobilization was encouraged with strict non-weight bearing for 8-10 weeks. Follow-up at 12 weeks revealed complete union at the fracture site, with the patient being asymptomatic except for minimal pain and mild restriction in dorsiflexion.

A review on properties, challenges and commercial aspects of eco-friendly biolubricants productions.

Lubricants operate as antifriction media, preserving machine reliability, facilitating smooth operation, and reducing the likelihood of frequent breakdowns. The petroleum-based reserves are decreasing globally, leading to price increases and raising concerns about environmental degradation. The researchers are concentrating their efforts on developing and commercializing an environmentally friendly lubricant produced from renewable resources. Biolubricants derived from nonedible vegetable oils are environmentally favorable because of their non-toxicity, biodegradability, and close to net zero greenhouse gas emissions. The demand for bio lubricants in industry and other sectors is increasing due to their non-toxic, renewable, and environmentally friendly nature. Good lubrication, anti-corrosion, and high flammability are characteristic properties of vegetable oils due to their unique structure. This study presents several key properties of nonedible oils that are used to produce lubricants via the transesterification process. Bibliometric analysis is also performed, which provides us with a better understanding of previous studies related to the production of bio lubricants from the transesterification process. Only 371 published documents in the Scopus database were found to relate to the production of bio lubricants using the transesterification process. The published work was mostly dominated by research articles (286; 77.088%). Significant development can be seen in recent years, with the highest occurrence in 2021, reaching 68 publications accounting for 18.38% of the total documents. In the second step, (i) the authors with the most number of publications; (ii) journals with the most productions; (iii) most productive countries; and (iv) the authors' most frequently used keywords were evaluated. These results will provide a pathway for researchers interested in this field. Lastly, recommendation is made on research gaps to device possible strategies for its commercialization.

Construction of Adipogenic ceRNA Network Based on lncRNA Expression Profile of Adipogenic Differentiation of Human MSC Cells.

The Long non-coding RNA (lncRNA) expression profile data of ten samples including human Mesenchymal Stem Cell (MSC) adipogenic differentiation 0, 3, and 6 days from the GEO database, and then perform gene ID conversion, BLAST comparison, and annotation marking. Finally, group A (treatment group on day 3 of differentiation and control group on day 0 of differentiation) obtained a total of 1180 mRNA and 185 lncRNA; group B (treatment group on day 6 of differentiation and control group on day 0 of differentiation). A total of 1376 mRNA and 206 lncRNA were obtained. Finally, we processed the differential lncRNAs and mRNAs obtained in the two groups, and obtained 113 shared differential lncRNAs to further predict the targeted miRNA, a total of 815 lncRNA-miRNA pairs. The targeted mRNA was further predicted, and the grouped differential mRNAs were combined to obtain 64 differential mRNAs. In the end, we obtained 216 ceRNAs containing 26 lncRNAs, 27 miRNAs and 64 mRNAs. We found that the mRNAs in the ceRNA network were mainly enriched with 45 Gene Ontology (GO) terms, mainly including glucose homeostasis mechanism and insulin stimulation response. 69 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were mainly enriched. It mainly includes many pathways related to lipid metabolism such as Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK), Rap1, cAMP, mitogen-activated protein kinase (MAPK), Ras, hypoxia inducible factor-1 (HIF-1), PI3K-Akt, insulin signaling and so on. In the end, we identified 216 ceRNA regulatory relationships related to obesity research. Our research provides a clearer direction for understanding the molecular mechanism of obesity, the screening and determination of drug targets biomarkers in the future.

Liver infarct masquerading as intrahepatic cholangiocarcinoma.

Cholangiocarcinoma is one of the most lethal tumors because of its complex location and lack of good chemoradiotherapy options. When it is diagnosed, urgent intervention is needed, often involving radical surgical resection. It generally presents as a liver mass with biliary obstruction. We discuss the case of a young patient presenting with liver dysfunction and imaging mimicking a liver mass concerning for cholangiocarcinoma, where he actually had a liver infarct from splanchnic venous thrombosis from primary myelofibrosis.

Trichinella spiralis: Knockdown of gamma interferon inducible lysosomal thiol reductase (GILT) results in the reduction of worm burden.

Trichinella spiralis is mammalian skeletal muscles parasite which may cause trichinellosis in animals and humans. Gamma interferon inducible lysosomal thiol reductase (GILT) is a widespread superfamily which plays key role in processing and presentation of MHC class II restricted antigen by catalyzing disulfide bond reduction. There are no reports about GILT in T. spiralis. In present study, GILT from T. spiralis (Tsp-GILT) was cloned, analyzed by multiple-sequence alignment, and predicted by 3D structure model. Recombinant Tsp-GILT (about 46 kDa) was efficiently expressed in Escherichia coli and thiol reductase activity suggested that in acidic environment the addition of a reducing agent is needed. Soaking method was used to knockdown expression of Tsp-GILT using small interference RNA (siRNA). Immunofluorescence assay confirmed the transformation of siRNA into muscle larva (ML) and new born larva (NBL). Quantitative real time-PCR (QRT-PCR) analysis revealed that transcription level of Tsp-GILT mRNA can be up-regulated by stimulation of mouse IFN-γ and down-regulated by siRNA2 in vitro. NBLs soaked with siRNA2 showed 32.3% reduction in the generation of MLs. MLs soaked with siRNA2 showed 26.2% reduction in the next generation of MLs, but no significant effect was observed on adult worms or NBLs. These findings concluded that GILT may play important roles in the development of T. spiralis parasite.

Characteristics of Biotin lipoyl attachment and 2-oxoacid dehydrogenase acyltransferase of the parasitic nematode Haemonchus contortus and its modulatory functions on goat PBMCs in vitro.

Biotin lipoyl attachment and 2-oxoacid dehydrogenase acyltransferase (BLAODA), as an essential excretion of Haemonchus contortus (HcESPs), was identified to have antigenic functions. T helper-9 (Th9) cells secrete interleukin (IL)-9, a signature cytokine associated with tumour immunology, allergy and autoimmunity. Nonetheless, the understanding of modulatory functions of BLAODA on Th9 and other immune cells is limited. In this study, the BLAODA gene was cloned, and the recombinant (r) protein of BLAODA (rHcBLAODA) was expressed and immunoblotting was performed. The results revealed that HcBLAODA gene was successfully cloned and rHcBLAODA protein was expressed. The localization of rHcBLAODA was confirmed on the surface of gut sections from adult H. contortus. The rHcBLAODA protein capability to react precisely with anti-H. contortus antibodies were confirmed by immunoblotting and immunofluorescence assay (IFA). Further functional analysis showed that interaction of rHcBLAODA with host cells significantly enhanced Th9 cells generation, IL-9 expression, nitric oxide production and cell apoptosis while suppressing the cells proliferation and cells migration depending on the concentration. Overall, these findings suggest that rHcBLAODA protein could modulate the host immune response by inducing Th9 cells to secrete IL-9 cytokine in vitro.

Ontology Driven Smart Health Service Integration.

BACKGROUND AND OBJECTIVE: The massive increase, in the Internet of Things applications, has greatly evolved technological aspects of human life. The drastic development of IoT based smart healthcare services have layout the smart process models to facilitate all stakeholders (e.g. patients, doctors, hospitals etc.) and made it an important social-economic concern. There are variety of smart healthcare services like remote patient monitoring, diagnostic, disease specific remote treatments and telemedicine. Many trending Internet of Health Things research and development are done in a very disjoint and independent fashion providing solutions and guidelines for variant diseases, medical resources and remote services management. These expositions work over many shared resources such as health facilities for patient and human in healthcare system. METHODS: This research discusses the ontology for merging methods to form an integrated platform with shared knowledge of smart healthcare services. The proposed process model creates an ontological framework of integrated healthcare services, which are firstly defined using ontologies and lately integrated over similarities, differences, dependencies and other semantic relations. The data and process requirements for service integration facility is derived from various smart healthcare services. RESULTS: The proposed model is evaluated using two-step ontological modeling testing method, applied at the ontological framework of integrated smart health services. First evaluation step has targeted the model consistency validation using reasoning tool while querying tools are used to validate the retrieved data entities and relations among them for predefined use-cases. CONCLUSIONS: The research concluded with a novel approach for smart health service integration using ontological modeling and merging techniques. The model efficiency enhancement and query optimization methods are listed in future tasks of the research.

A Case to Overcome the Stigma of Testicular Cancer.

Testicular cancer is seen commonly in young males and has a high cure rate if diagnosed and treated early. It clinically presents with painless testicular swelling. We discuss the case of a young previously healthy male with an aggressive testicular cancer which started with a testicular swelling but the primary site underwent necrosis secondary to its own vascular demand thus giving the false impression of resolution, but not before metastasizing to the rest of the body. With this case, we aim to highlight the importance of increasing awareness of testicular cancer and its presenting symptoms in young males and the need to overcome the stigma around the evaluation of testicular swellings.

Nanoparticles (PLGA and Chitosan)-Entrapped ADP-Ribosylation Factor 1 of Haemonchus contortus Enhances the Immune Responses in ICR Mice.

ADP-ribosylation factor 1 (HcARF1) is one of the Haemonchus contortus (H. contortus) excretory/secretory proteins involved in modulating the immune response of goat peripheral blood mononuclear cells (PBMC). Here, we evaluated the immunogenic potential of recombinant HcARF1 (rHcARF1) against H. contortus infection in Institute of Cancer Research (ICR) mice. Briefly, rHcARF1 was entrapped in poly (D, L-lactide-co-glycolide) (PLGA) and chitosan (CS) nanoparticles (NP) and injected into mice as a vaccine. Fifty-six ICR mice were assigned randomly into seven groups, with eight animals in each group, and they were vaccinated subcutaneously. At the end of the experiment (14th day), the blood and the spleen were collected from euthanized mice to detect lymphocyte proliferation, cytokine analysis, and the production of antigen-specific antibodies. Scanning electron microscope was used to determine the size, morphology, and zeta potential of nanoparticles. Flow cytometry was performed, which presented the increase percentages of CD4+ T cells (CD3e+CD4+), CD8+ T cells (CD3e+CD8+) and dendritic cells (CD11c+CD83+, CD11c+CD86+) in mice vaccinated with rHcARF1+PLGA NP. Immunoassay analysis show raised humoral (Immunoglobulin (Ig)G1, IgG2a, IgM) and cell-mediated immune response (Interleukin (IL)-4, IL-12, and IL-17, and Interferon (IFN)-γ) induced by rHcARF1+PLGA NP. Experimental groups that were treated with the antigen-loaded NP yield higher lymphocyte proliferation than the control groups. Based on these results, we could propose that the rHcARF1 encapsulated in NP could stimulate a strong immune response in mice rather than administering alone against the infection of H. contortus.

In vitro effects of 5 recombinant antigens of Eimeria maxima on maturation, differentiation, and immunogenic functions of dendritic cells derived from chicken spleen.

Eimeria maxima possesses integral families of immunogenic constituents that promote differentiation of immune cells during host-parasite interactions. Dendritic cells (DCs) have an irreplaceable role in the modulation of the host immunity. However, the selection of superlative antigen with immune stimulatory efficacies on host DCs is lacking. In this study, 5 recombinant proteins of E. maxima (Em), including Em14-3-3, rhomboid family domain containing proteins (ROM) EmROM1 and EmROM2, microneme protein 2 (EmMIC2), and Em8 were identified to stimulate chicken splenic derived DCs in vitro. The cultured populations were incubated with recombinant proteins, and typical morphologies of stimulated DCs were obtained. DC-associated markers major histocompatibility complex class II, CD86, CD11c, and CD1.1, showed upregulatory expressions by flow cytometry assay. Immunofluorescence assay revealed that recombinant proteins could bind with the surface of chicken splenic derived DCs. Moreover, quantitative real-time PCR results showed that distinct gene expressions of Toll-like receptors and Wnt signaling pathway were upregulated after the coincubation of recombinant proteins with DCs. The ELISA results indicated that the DCs produced a significant higher level of interleukin (IL)-12 and interferon-γ secretions after incubation with recombinant proteins. While transforming growth factor-ß was significantly increased with rEmROM1, rEmROM2, and rEmMIC2 as compared to control groups, and IL-10 did not show significant alteration. Taken together, these results concluded that among 5 potential recombinant antigens, rEm14-3-3 could promote immunogenic functions of chicken splenic derived DCs more efficiently, which might represent an effective molecule for inducing the host Th1-mediated immune response against Eimeria infection.

Glyceraldehyde-3-phosphate dehydrogenase from Eimeria acervulina modulates the functions of chicken dendritic cells to boost Th1 type immune response and stimulates autologous CD4+ T cells differentiation in-vitro.

Dendritic cells (DCs) play a pivotal role to amplify antigen-specific immune responses. Antigens that sensitize T cells via antigen-presentation by DCs could enhance the capacity of host immunity to fight infections. In this study, we tested the immunogenic profiles of chicken DCs towards Glyceraldehyde-3-phosphate dehydrogenase from Eimeria acervulina (EaGAPDH). Immunoblot analysis showed that recombinant EaGAPDH (rEaGAPDH) protein was successfully recognized by rat sera generated against rEaGAPDH. Interaction and internalisation of rEaGAPDH by chicken splenic-derived DCs (chSPDCs) was confirmed by immunofluorescence analysis. Flow cytometry revealed that chSPDCs upregulated MHCII, CD1.1, CD11c, CD80, and CD86 cell-surface markers. Moreover, mRNA expressions of DC maturation biomarkers (CCL5, CCR7, and CD83) and TLR signalling genes (TLR15 and MyD88) were also upregulated whereas those of Wnt signalling were non-significant compared to negative controls. rEaGAPDH treatment induced IL-12 and IFN-γ secretion in chSPDCs but had no effect on IL-10 and TGF-ß. Likewise, DC-T cell co-culture promoted IFN-γ secretion and the level of IL-4 was unaffected. Proliferation of T cells and their differentiation into CD3+/CD4+ T cells were triggered in chSPDCs-T cells co-culture system. Taken together, rEaGAPDH could promote Th1 polarization by activating both host DCs and T cells and sheds new light on the role of this important molecule which might contribute to the development of new DCs-based immunotherapeutic strategies against coccidiosis.

Unveiling the Immunomodulatory Characteristics of Haemonchus contortus Ephrin Domain Containing Protein in the Parasite-Host Interactions.

Ephrin domain containing protein (EPH), a significant excreted and secreted product (ESPs) of Haemonchus contortus, has been identified to have antigenic functions. Over the past years, a new subset of CD4 + T named as T helper 9 cells that secrete interleukin-9 (IL-9) as a signature cytokine is associated with tumor immunity and allergy. Nonetheless, the understanding of immunomodulatory roles of EPH on goat Th9 and other immune cells remains limited. Herein, EPH from H. contortus (HcEPH) was cloned and expressed in pET-28a. Immunofluorescence assay (IFA) was carried-out to localize rHcEPH within H. contortus adult worms and to bind with goat peripheral blood mononuclear cells (PBMCs). Besides, the impact of rHcEPH on signature cytokine IL-9 expression in goat PBMCs was evaluated. Flow cytometry was employed to examine Th9 cells production and cell apoptosis. The results revealed success in the expression and localization of rHcEPH in surface of adult H. contortus gut sections. According to IFA analysis, the rHcEPH protein was capable to react precisely with anti-H. contortus antibodies. Further functional analysis showed that correlation between rHcEPH and host PBMCs significantly enhanced Th9 cell differentiation, IL-9 expression, cell apoptosis efficiency, and cell migration, whereas cell proliferation was suppressed significantly depending on the concentration. Our observations indicated that rHcEPH protein is linked to modulate the host immune cells and could enhance protective immunity by inducing Th9 responses.

Immunomodulatory dynamics of excretory and secretory products on Th9 immune response during Haemonchus contortus infection in goat.

CD4+ T cells play critical roles in mediating adaptive immunity to a variety of pathogens. Recently, new subset of CD4+T named as T helper 9 cells that express the prototypical interleukin-9 (IL-9) cytokine have been recognized in human and mice models during different parasitic infections. Haemonchus contortus is a gastrointestinal nematode of small ruminants which cause high mortality in young animals. During infection, Excretory and Secretary Products (ESPs) are released in the host body. No other study has reported yet on immunomodulatory dynamics of H. contortus ESPs on Th9 immune response in vitro or in vivo. In this study, immunomodulatory effects of ESPs (5, 10, 20, 40, 80; µg/mL) incubated with goat PBMCs on Th9 cells, IL-9 immune response and TGF-ß/Smad signaling regulator were evaluated in vitro. Moreover, for in vivo study, goats were infected with different doses (P-800, P-2400, and P-8000) of H. contortus infective larva (L3) and immunomodulatory effects on Th9 cells, IL-9 immune response and TGF-ß/Smad signaling regulator were evaluated at 7, 10, 14, 18, 21, 28 Days Post Infection (DPI). Flow cytometry was performed to evaluate the effects on Th9 cells and quantitative real time polymerase chain reaction was performed to evaluate the IL-9 cytokine transcription level. Additionally, fecal egg counting was also performed in parallel to confirm the infection. All goats were dewormed at 29 DPI and all experiments were also performed at 35 DPI, one week post deworming. The finding indicated that 10, 20, 40, 80 µg/mL concentration of ESPs incubated with goat PBMCs showed significant increase in the production of Th9 cells, signature cytokine IL-9 and expression of TGF-ß/Smad signaling regulator as compared to control group in vitro.All infected groups showed significant increase in production of Th9 cells and IL-9 cytokine and expression of TGF-ß/Smad key genes at 18, 21, and 28 DPI as compared to control group. Likewise, at 14 DPI, P-2400 and P-8000 groups showed significant increase in production of Th9 cells, IL-9 cytokine and expression of TGF-ß/Smad key genes. While at 10 DPI, production of Th9 cells and IL-9 was significantly increased in P-2400 & P-8000 groups, and at 7 DPI only P-8000 showed significantly increase in IL-9 production. No immunomodulatory effects were observed at 0 and 3 DPI. Additionally, significant gradually up-regulated key genes expression of TGF-ß/Smad signaling regulator in all infected groups confirmed the above results. After deworming, production of Th9 cells, associated immune response and expression of signaling regulator in each group were significantly decreased. Based on this study, it is concluded that Th9 immune response was induced during H. contortus infection in goat by up-regulation of TGF-ß/Smad signaling key genes.

Recombinant ubiquitin-conjugating enzyme of Eimeria maxima induces immunogenic maturation in chicken splenic-derived dendritic cells and drives Th1 polarization in-vitro.

Dendritic cells (DCs) are key linkages between innate immunity and acquired immunity. The antigens that promote the functions of DCs might be the effective candidates of novel vaccine. In this research, the ability of ubiquitin-conjugating enzyme (UCE), a recognized common antigens among chicken Eimeria species, to stimulate DCs of chickens were evaluated. We cloned UCE gene from Eimeria maxima (EmUCE), and its protein expression was confirmed by SDS-PAGE and western-blot. Immunofluorescence assay confirmed the binding of rEmUCE on the surface of chicken splenic-derived DCs (ChSP-DCs). Flow cytometric analysis showed that rEmUCE-treated ChSP-DCs increased MHCII, CD1.1, CD11c, CD80, and CD86 phenotypes. qRT-PCR indicated that transcript levels of maturation markers CCL5, CCR7, and CD83 in ChSP-DCs were upregulated in response to rEmUCE. Following rEmUCE treatment, chSP-DCs activated TLR signaling and inhibited Wnt signaling. Moreover, rEmUCE promoted DC-mediated T-cell proliferation in DC/T-cell co-incubation. Interestingly, CD3+/CD4+ T-cells were significantly enhanced when rEmUCE-treated chSP-DCs were co-incubated with T-cells. Cytokine secretion pattern of rEmUCE-stimulated ChSP-DCs revealed that the production of IL-12 and IFN-γ was increased whereas IL-10 and TGF-ß were unchanged. Likewise, the co-incubation of ChSP-DCs with T-cells indicated increased production of IFN-γ but not IL-4. Collectively, rEmUCE could polarize DCs to immunogenic phenotype and shift the immune cells towards Th1 response. Our observations provide valuable insight for future research aimed at vaccine development against avian coccidiosis.