Production, purification, and functional characterization of glucan exopolysaccharide produced by Enterococcus hirae strain OL616073 of fermented food origin.

Microbial exopolysaccharides (EPS) are high molecular weight polymeric substances with great diversity and variety of applications in the food and pharma industry. In this study, we report the extraction of an EPS from Enterococcus hirae OL616073 strain originally isolated from Indian fermented food and its purification by ion exchange and size exclusion chromatography for physical-functional analyses. The EPS showed two prominent fractions (EPS F1 and EPS F2) with molecular mass 7.7 × 104 and 6.5 × 104 Da respectively by gel permeation chromatography. These fractions were further characterized by FTIR, HPTLC, GC-MS, and NMR as a homopolysaccharide of glucose linked with α-(1 â†’ 6) and α-(1 â†’ 3) glycosidic linkages. The porous, spongy, granular morphology of EPS was observed under scanning electron microscopy. EPS has revealed strong physico-functional properties like water solubility index (76.75 %), water contact angle (65.74°), water activity (0.35), hygroscopicity (3.05 %), water holding capacity (296.19 %), oil holding capacity (379.91 %), foaming capacity (19.58 %), and emulsifying activity (EA1-72.22 %). Rheological analysis showed that aqueous solution of EPS exhibited a non-Newtonian fluid behavior and shear-thinning characteristics. Overall, EPS exhibits techno functional properties with potential applications as a functional biopolymer in food and pharma industry.

Multifunctional role of the ubiquitin proteasome pathway in phagocytosis.

Phagocytosis is a specialized form of endocytosis where large cells and particles (>0.5µm) are engulfed by the phagocytic cells, and ultimately digested in the phagolysosomes. This process not only eliminates unwanted particles and pathogens from the extracellular sources, but also eliminates apoptotic cells within the body, and is critical for maintenance of tissue homeostasis. It is believed that both endocytosis and phagocytosis share common pathways after particle internalization, but specialized features and differences between these two routes of internalization are also likely. The recruitment and removal of each protein/particle during the maturation of endocytic/phagocytic vesicles has to be tightly regulated to ensure their timely action. Ubiquitin proteasome pathway (UPP), degrades unwanted proteins by post-translational modification of proteins with chains of conserved protein Ubiquitin (Ub), with subsequent recognition of Ub chains by the 26S proteasomes and substrate degradation by this protease. This pathway utilizes different Ub linkages to modify proteins to regulate protein-protein interaction, localization, and activity. Due to its vast number of targets, it is involved in many cellular pathways, including phagocytosis. This chapters describes the basic steps and signaling in phagocytosis and different roles that UPP plays at multiple steps in regulating phagocytosis directly, or through its interaction with other phagosomal proteins. How aberrations in UPP function affect phagocytosis and their association with human diseases, and how pathogens exploit this pathway for their own benefit is also discussed.

The characterization and quantification of structures of Cajanus scarabaeoides phytochemicals and their seasonal variation analysis using ultra-performance liquid chromatography-tandem mass spectrometry.

RATIONALE: Cajanus scarabaeoides, belonging to the Fabaceae family, is an underutilized herb and traditionally used to treat several ailments. However, it is not well explored phytochemically. Therefore, mass spectrometry (MS)-based phytochemical analysis was carried out to investigate the bioactive ingredients of the herb. METHODS: A ultra-performance liquid chromatography (UPLC) coupled to photodiode array detection (PDA) and electrospray ionization (ESI) tandem mass spectrometry (UPLC-PDA-ESI-MS/MS) system was used for the qualitative and quantitative analysis of phytochemicals. The chromatographic separation was achieved on the Acquity BEH C18 column (150 × 2.1 mm, 1.7 µm) using a gradient system consisting of three solvents, acetonitrile, methanol, and 0.1% formic acid, used at a flow rate of 0.300 ml/min. RESULTS: Sixteen bioactive ingredients (gallic acid, gallocatechin, epigallocatechin, catechin, procyanidin dimer, epicatechin, procyanidin trimer, isoorientin, orientin, vitexin, isovitexin, quercetin-mono-O-glycoside, isoquercitrin, luteolin-7-O-glucoside, quercetin, and luteolin) were identified and structurally characterized. Consequently, 12 compounds were reported for the first time from C. scarabaeoides, and 13 were quantitatively determined in different seasons. Isoorientin (10.2-7.1% w/w) and orientin (5.78-5.17% w/w) were the most abundant constituents in the dry weight of plant material, followed by vitexin and isovitexin in the rainy season. CONCLUSIONS: The phytochemical investigation has revealed that C. scarabaeoides could be a potential alternate source of bioactive ingredients, namely, isoorientin, orientin, vitexin, and isovitexin, contributing to further exploration of its biological activity. In addition, analytical methods can be used for the rapid identification and quantification of bioactive ingredients in C. scarabaeoides.

Erratum to: Double Rectangle Fascia Lata Frontalis Sling: A Rationale Approach for Ptosis.

[This corrects the article DOI: 10.1055/s-0041-1723909.].

Bacterial exopolysaccharides for improvement of technological, functional and rheological properties of yoghurt.

Exopolysaccharides (EPS) are known to have technological and functional applications in food industry including dairy based products such as yoghurt. Yoghurt is a widely consumed dairy based product due to pleasant taste and texture, as well as a source of nutrients and bioactive compounds. At the same time, structural, rheological and sensorial properties are important in the production of good quality yoghurt. Various natural hydrocolloids including EPS with stabilizing and texture enhancing properties could be useful in enhancing these desirable properties. Apart from that, EPS may enhance various other functional properties of yoghurt such as antioxidant and prebiotic potential. Based on its prebiotic property, symbiotic products could be developed by combining EPS and probiotic bacterial strains. EPS has potential to provide physical and micro structural stability, thereby enhancing the protein distribution and viscoelastic properties. Main focus of the present review is to provide an insight on the action of EPS as a functional hydrocolloid on the technological, rheological and functional properties of yoghurt and related products.

LxCxD motif of the APC/C coactivator subunit FZR1 is critical for interaction with the retinoblastoma protein.

Retinoblastoma protein (pRB) regulates cell cycle by utilizing different regions of its pocket domain for interacting with E2F family of transcription factors and with cellular and viral proteins containing an LxCxE motif. An LxCxE-like motif, LxCxD, is present in FZR1, an adaptor protein of the multi-subunit E3 ligase complex anaphase-promoting complex/cyclosome (APC/C). The APC/CFZR1 complex regulates the timely degradation of multiple cell cycle proteins for mitotic exit and maintains G1 state. We report that FZR1 interacts with pRB via its LxCxD motif. By using point mutations, we found that the cysteine residue in the FZR1 LxCxD motif is critical for direct interaction with pRb. The direct binding of the LxCxD motif of FZR1 to the pRB LxCxE binding pocket is confirmed by using human papillomavirus protein E7 as a competitor, both in vitro and in vivo. While mutation of the cysteine residue significantly disrupts FZR1 interaction with pRB, this motif does not affect FZR1 and core APC/C association. Expression of the FZR1 point mutant results in accumulation of S-phase kinase-associated protein 2 (SKP2) and Polo-like kinase 1 (PLK1), while p27Kip1 and p21Cip1 proteins are downregulated, indicating a G1 cell cycle defect. Consistently, cells containing point mutant FZR1 enter the S phase prematurely. Together our results suggest that the LxCxD motif of FZR1 is a critical determinant for the interaction between FZR1 and pRB and is important for G1 restriction.

Double Rectangle Fascia Lata Frontalis Sling: A Rationale Approach for Ptosis.

Background The purpose of this study was to evaluate the functional outcomes of a modified technique of double rectangle pattern for correction of severe ptosis. Methods This is a retrospective study over a period of 8 years including patients who underwent correction of ptosis by double rectangle using autologous fascia lata sling. Surgical outcomes were assessed postoperatively by distance from the corneal light reflex to the upper eyelid margin (MRD1) and levator function. Results Twenty-six eyelids were operated in 20 patients. There were 9 males and 11 females, with age ranging from 4 to 35 years. Preoperatively, all patients had poor MRD1 and poor levator function. Postoperative MRD1 was good in 13 patients (17 eyelids), fair in 5 (7eyelids), and poor in 2 patients (2 eyelids). Postoperative levator function was excellent in 12 patients (15 eyelids), good in 6 (9 eyelids), and fair in 2 patients (2 eyelids). At a mean follow-up of 12 months, adequate correction was achieved in 24 eyelids, and 2 eyelids had undercorrection. Conclusion Frontalis sling with a double rectangle is simple and more efficient, as it provides a straight line of pull to the eyelid for correction of severe ptosis.

Artificial intelligence to deep learning: machine intelligence approach for drug discovery.

Drug designing and development is an important area of research for pharmaceutical companies and chemical scientists. However, low efficacy, off-target delivery, time consumption, and high cost impose a hurdle and challenges that impact drug design and discovery. Further, complex and big data from genomics, proteomics, microarray data, and clinical trials also impose an obstacle in the drug discovery pipeline. Artificial intelligence and machine learning technology play a crucial role in drug discovery and development. In other words, artificial neural networks and deep learning algorithms have modernized the area. Machine learning and deep learning algorithms have been implemented in several drug discovery processes such as peptide synthesis, structure-based virtual screening, ligand-based virtual screening, toxicity prediction, drug monitoring and release, pharmacophore modeling, quantitative structure-activity relationship, drug repositioning, polypharmacology, and physiochemical activity. Evidence from the past strengthens the implementation of artificial intelligence and deep learning in this field. Moreover, novel data mining, curation, and management techniques provided critical support to recently developed modeling algorithms. In summary, artificial intelligence and deep learning advancements provide an excellent opportunity for rational drug design and discovery process, which will eventually impact mankind. The primary concern associated with drug design and development is time consumption and production cost. Further, inefficiency, inaccurate target delivery, and inappropriate dosage are other hurdles that inhibit the process of drug delivery and development. With advancements in technology, computer-aided drug design integrating artificial intelligence algorithms can eliminate the challenges and hurdles of traditional drug design and development. Artificial intelligence is referred to as superset comprising machine learning, whereas machine learning comprises supervised learning, unsupervised learning, and reinforcement learning. Further, deep learning, a subset of machine learning, has been extensively implemented in drug design and development. The artificial neural network, deep neural network, support vector machines, classification and regression, generative adversarial networks, symbolic learning, and meta-learning are examples of the algorithms applied to the drug design and discovery process. Artificial intelligence has been applied to different areas of drug design and development process, such as from peptide synthesis to molecule design, virtual screening to molecular docking, quantitative structure-activity relationship to drug repositioning, protein misfolding to protein-protein interactions, and molecular pathway identification to polypharmacology. Artificial intelligence principles have been applied to the classification of active and inactive, monitoring drug release, pre-clinical and clinical development, primary and secondary drug screening, biomarker development, pharmaceutical manufacturing, bioactivity identification and physiochemical properties, prediction of toxicity, and identification of mode of action.

Post-translational modifications: Regulators of neurodegenerative proteinopathies.

One of the hallmark features in the neurodegenerative disorders (NDDs) is the accumulation of aggregated and/or non-functional protein in the cellular milieu. Post-translational modifications (PTMs) are an essential regulator of non-functional protein aggregation in the pathogenesis of NDDs. Any alteration in the post-translational mechanism and the protein quality control system, for instance, molecular chaperone, ubiquitin-proteasome system, autophagy-lysosomal degradation pathway, enhances the accumulation of misfolded protein, which causes neuronal dysfunction. Post-translational modification plays many roles in protein turnover rate, accumulation of aggregate and can also help in the degradation of disease-causing toxic metabolites. PTMs such as acetylation, glycosylation, phosphorylation, ubiquitination, palmitoylation, SUMOylation, nitration, oxidation, and many others regulate protein homeostasis, which includes protein structure, functions and aggregation propensity. Different studies demonstrated the involvement of PTMs in the regulation of signaling cascades such as PI3K/Akt/GSK3ß, MAPK cascade, AMPK pathway, and Wnt signaling pathway in the pathogenesis of NDDs. Further, mounting evidence suggests that targeting different PTMs with small chemical molecules, which acts as an inhibitor or activator, reverse misfolded protein accumulation and thus enhances the neuroprotection. Herein, we briefly discuss the protein aggregation and various domain structures of different proteins involved in the NDDs, indicating critical amino acid residues where PTMs occur. We also describe the implementation and involvement of various PTMs on signaling cascade and cellular processes in NDDs. Lastly, we implement our current understanding of the therapeutic importance of PTMs in neurodegeneration, along with emerging techniques targeting various PTMs.

Silicon Interpositional Arthroplasty for Temporo-mandibular Joint Ankylosis.

Temporo-mandibular joint (TMJ) ankylosis is characterized by a decreased mouth opening which affects mastication, speech, and facial aesthetic. Interpositional arthroplasty using autologous tissue is accepted treatment for TMJ Ankylosis; however, harvesting autologous tissue is associated with additional morbidity. In this article we report our results of silicon interpositional arthroplasty for TMJ ankylosis. 20 patients with TMJ ankylosis were included in the study. All patients underwent standard operative procedure using preauricular incision for release of TMJ ankylosis by excision of at least 1 cm of bony block and insertion of 2 cm thick silicon block in the joint space. Postoperatively early mobilization of TMJ was advised from 3rd day onwards. Post operative result was evaluated by assessing the mouth opening as inter incisor distance (IID). 20 patients (27 joints) of TMJ ankylosis were included in the study. There were 8 male and 12 female patients with age ranged from 3-35 years. According to Sawhney classification bony ankylosis was present as Type-IV (n = 13 joints), Type-III (n = 12 joints) and Type-II (2 joints). Preoperative mean IID was 7.15 mm and post operative mean IID was 43.5. There was no facial nerve paresis, malocclusion or recurrence but infection and extrusion of implant occurred in 1 case each. Silicon interpositional arthroplasty is an effective procedure for the treatment of TMJ Ankylosis; as it restores mouth opening and function, maintains the vertical ramus height, and prevents re-ankylosis without any donor site morbidity.

Covaxin: An overview of its immunogenicity and safety trials in India.

The spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to a global coronavirus disease-19 (COVID-19) pandemic. Several vaccine types, such as inactivated, viral vector-, or mRNA-based, have received approval against SARS-CoV-2. The ability to induceT-helper-1 cell (Th1) responses is desirable from an effective vaccine against this virus. Covaxin (BBV152) is a wholevirion inactivated SARS-CoV-2 vaccine adjuvanted with Algel-Imidazoquinoline (IMDG) molecule, a toll-like receptor (TLR) 7/8 agonist. The mRNA-based vaccine use is hindered because of cold storage requirement, whereas covaxin is stored between 2°C and 8°C, making it suitable for countries with limited resources. The Drug Controller General of India (DCGI) has approved the BBV152 vaccine. Therefore, it is of interest to document known data on BBV152 vaccine phase I, phase II and phase III human clinical trials to evaluate the safety, reactogenicity, tolerance, and immunogenicity of the whole-virion inactivated SARS-CoV-2 vaccine (BBV152).

Three way ANOVA for emulsion of carotenoids extracted in flaxseed oil from carrot bio-waste.

The juice expelled from carrot, a globally produced root vegetable, leavesbehind carrot pomace (a bio- and horticultural waste) which is potentially rich source of micro-nutrients and carotenoids.However, it is discarded as waste or used as animal feed. It holds potential to be channelized to food chain by a couple of technological interventions. In this regard, present work was aimed at preparing stable emulsion based delivery system for 'green' carotenoids extracted from carrot-pomace in flaxseed oil (a green solvent), and at maximizing the amount of core material so that the resultant emulsion can potentially be used as a source of both carotenoids and omega-3 fatty acid of flaxseed oil origin. The study used natural emulsifier. Preparation of oil-in-water emulsion was optimized using 33 factorial experiment by varying levels of extract containing carotenoid (30-40%), whey protein concentrates (WPC-80) and lactose. The optimized emulsion (CREm) was selected on the basis of particle size, zeta potential, color values (L*, a*, b*) and viscosity statistically analyzed via three-way ANOVA using Proc GLM of SAS 9.3 (described in detail in this paper); the respective values of these parameters being 120.03 ± 8.20 nm, -16.57 ± 0.49 mV, 75.11 ± 0.04, 9.66 ± 0.32, 50.29 ± 0.62, and 0.124 ± 0.0115 Pa.s for CREm. CREm contained 35% flaxseed oil, 10% WPC-80 and 5% lactose and showed good centrifugal and gravitational stability (15 days). It was analyzed for total carotenoid content, antioxidant activities (ABTS (2,2-azinobis-(3-ethylbenzthiazoline-6sulfonic acid), DPPH (2,2-Diphenyl-1-picrylhydrazyl) and FRAP (Ferric reducing antioxidant power assay)) and microstructure.

Bilaterally Variant Accessory Fibularis Muscle: Its Phylogenetic, Molecular and Clinical Perspective.

Accessory fibularis muscle is prevalent in 2.9-21.8% of the world population. Incidentally during routine dissection of a 75-year-old male cadaver, bilaterally accessory fibularis muscle was observed. On both the sides, proximal site of attachment was same but muscle displayed different distal sites of insertion in the foot. Appearance of accessory muscle in the leg is indicative towards the ongoing phylogenetic evolution operating at the molecular level. Bio-mechanical advantage of this variant muscle is the additional support provided to the subtalar joint. Also it acts as synergist to fibularis longus and brevis during eversion of the foot. Clinically this muscle may predispose to chronic ankle pain, dislocation of peroneal tendons from retromalleolar groove and post fracture dislocation in foot. Wide range of accessory fibularis muscle has been previously reported with different nomenclature, however, existence of two different variants in same cadaver has been rarely reported. The current observation is significant for clinicians to acknowledge when evaluating and operating patients with foot disorders.

Organic solvent-free extraction of carotenoids from carrot bio-waste and its physico-chemical properties.

The bio-wastes (like peels, seeds, etc.) from food industry are rich source of bio-active components, but are poorly managed. In present study, carotenoids were extracted from carrot pomace using ultrasonication and high shear dispersion techniques and flaxseed oil as green solvent (green biorefinery approach). Various combinations of time and temperature were used and final selection was made on the basis of maximum recovery of carotenoids. High shear disperser yielded maximum carotenoids (recovery 94.8 ± 0.08%). The total carotenoid content, antioxidant activity as ABTS, DDPH and FRAP and ß-carotene of carotenoid rich extract from carrot pomace (CREP) were 82.66 ± 0.06 µg/g, 1596.04 ± 69.45 µg Trolox eq./ml, 380.21 ± 39.62 µg Trolox eq./ml, 941.20 ± 19.91 µM Trolox eq./ml, 78.37 µg/g, respectively were significantly higher (p < 0.05) when compared with the extracting medium. The L*, a* and b* values of CRE were 18.65 ± 0.037, 19.42 ± 0.21, 27.947 ± 0.65 and were significantly higher than extracting medium. The CRE could be used as a natural source of ß-carotene and natural colorant for food applications.

Mechanisms for the temporal regulation of substrate ubiquitination by the anaphase-promoting complex/cyclosome.

The anaphase-promoting complex/cyclosome (APC/C) is a multi-subunit, multifunctional ubiquitin ligase that controls the temporal degradation of numerous cell cycle regulatory proteins to direct the unidirectional cell cycle phases. Several different mechanisms contribute to ensure the correct order of substrate modification by the APC/C complex. Recent advances in biochemical, biophysical and structural studies of APC/C have provided a deep mechanistic insight into the working of this complex ubiquitin ligase. This complex displays remarkable conformational flexibility in response to various binding partners and post-translational modifications, which together regulate substrate selection and catalysis of APC/C. Apart from this, various features and modifications of the substrates also influence their recognition and affinity to APC/C complex. Ultimately, temporal degradation of substrates depends on the kind of ubiquitin modification received, the processivity of APC/C, and other extrinsic mechanisms. This review discusses our current understanding of various intrinsic and extrinsic mechanisms responsible for 'substrate ordering' by the APC/C complex.

Ubc7/Ube2g2 ortholog in Entamoeba histolytica: connection with the plasma membrane and phagocytosis.

Endoplasmic reticulum (ER)-associated degradation (ERAD) and unfolded protein response (UPR) pathways are important for quality and quantity control of membrane and secretory proteins. We have identified orthologs of ER-associated ubiquitin conjugating enzymes (E2s) Ubc6/Ube2j2 and Ubc7/Ube2g2, ubiquitin ligases (E3) Hrd1 and GP78/AMFR, and sensor of UPR, Ire1 in E. histolytica that show conservation of important features of these proteins. Biochemical characterization of the ortholog of ERAD E2, Ubc7/Ube2g2 (termed as EhUbc7), was carried out. This E2 was transcriptionally upregulated several folds upon induction of UPR with tunicamycin. Ire1 ortholog was also upregulated upon UPR induction suggesting a linked UPR and ERAD pathway in this organism. EhUbc7 showed enzymatic activity and, similar to its orthologs in higher eukaryotes, formed polyubiquitin chains in vitro and localized to both cytoplasm and membranes. However, unlike its ortholog in higher eukaryotes, it also showed localization to the plasma membrane along with calreticulin. Inactivation of EhUbc7 significantly inhibited erythrophagocytosis, suggesting a novel function that has not been reported before for this E2. No change in growth, motility, or cell-surface expression of Gal/GalNAC lectin was observed due to inactivation of EhUbc7. The protein was present in the phagocytic cups but not in the phagosomes. A significant decrease in the number of phagocytic cups in inactive EhUbc7 expressing cells was observed, suggesting altered kinetics of phagocytosis. These findings have implications for evolutionary and mechanistic understanding of connection between phagocytosis and ER-associated proteins.

A Unique Conglomeration of Variations in the Celiac, Hepatic, and Superior Mesenteric Artery: A Clinico-Embryological Perspective.

This paper highlights a rare variation in the branching pattern of the celiac trunk and superior mesenteric arteries, as observed during cadaveric dissection. It was found that the celiac trunk gave origin to the following: (1) left inferior phrenic artery, (2) hepatogastric trunk which gave three branches: the left gastric artery, esophageal branch, and a left hepatic artery, (3) splenic artery, and (4) common hepatic artery. The superior mesenteric artery gave origin to the right hepatic artery, and the common hepatic artery gave origin to a middle hepatic artery. Such rare variations must be brought to the notice of surgeons and radiologists to prevent any undue complications during any interventional procedures and surgeries.

APC/C and retinoblastoma interaction: cross-talk of retinoblastoma protein with the ubiquitin proteasome pathway.

The ubiquitin (Ub) ligase anaphase promoting complex/cyclosome (APC/C) and the tumour suppressor retinoblastoma protein (pRB) play key roles in cell cycle regulation. APC/C is a critical regulator of mitosis and G1-phase of the cell cycle whereas pRB keeps a check on proliferation by inhibiting transition to the S-phase. APC/C and pRB interact with each other via the co-activator of APC/C, FZR1, providing an alternative pathway of regulation of G1 to S transition by pRB using a post-translational mechanism. Both pRB and FZR1 have complex roles and are implicated not only in regulation of cell proliferation but also in differentiation, quiescence, apoptosis, maintenance of chromosomal integrity and metabolism. Both are also targeted by transforming viruses. We discuss recent advances in our understanding of the involvement of APC/C and pRB in cell cycle based decisions and how these insights will be useful for development of anti-cancer and anti-viral drugs.

High Level of Perforin Expression Is Required for Effective Correction of Hemophagocytic Lymphohistiocytosis.

Perforin-1 mutations result in a potentially fatal hemophagocytic lymphohistiocytosis (HLH) with heightened immune activation, hypercytokinemia, pancytopenia, and end-organ damage. At present, hematopoietic stem cell (HSC) transplantation is curative, but limited by donor availability and associated mortality, making gene therapy an attractive alternative approach for HLH. We reported that perforin expression driven by cellular promoters in lentiviral (LV) vectors resulted in significant, albeit partial, correction of the inflammatory features in a murine model of HLH. We hypothesized that the level of perforin expression achieved per cell from ectopic moderate-strength cellular promoters (phosphoglycerate kinase gene/perforin-1 gene) is inadequate and thus engineered an LV vector using a viral promoter (MND; a modified Moloney murine leukemia virus long terminal repeat with myeloproliferative sarcoma virus enhancer) containing microRNA126 target sequences to restrict perforin expression in HSCs. We show here that the MND-LV vector restored perforin expression to normal levels in a perforin-deficient human natural killer cell line and perforin gene-corrected Perforin1-/- transplant recipients, whereas cellular promoters drove only partial correction. On lymphocytic choriomeningitis virus challenge, the clinical scores and survival improved only with the MND-LV vector, but inflammatory markers and cytotoxicity were improved with all LV vectors. Our studies suggest that although moderate levels of expression can result in partial amelioration of the HLH phenotype, high levels of perforin expression per cell are required for complete correction of HLH.

Placenta growth factor augments airway hyperresponsiveness via leukotrienes and IL-13.

Airway hyperresponsiveness (AHR) affects 55%-77% of children with sickle cell disease (SCD) and occurs even in the absence of asthma. While asthma increases SCD morbidity and mortality, the mechanisms underlying the high AHR prevalence in a hemoglobinopathy remain unknown. We hypothesized that placenta growth factor (PlGF), an erythroblast-secreted factor that is elevated in SCD, mediates AHR. In allergen-exposed mice, loss of Plgf dampened AHR, reduced inflammation and eosinophilia, and decreased expression of the Th2 cytokine IL-13 and the leukotriene-synthesizing enzymes 5-lipoxygenase and leukotriene-C4-synthase. Plgf-/- mice treated with leukotrienes phenocopied the WT response to allergen exposure; conversely, anti-PlGF Ab administration in WT animals blunted the AHR. Notably, Th2-mediated STAT6 activation further increased PlGF expression from lung epithelium, eosinophils, and macrophages, creating a PlGF/leukotriene/Th2-response positive feedback loop. Similarly, we found that the Th2 response in asthma patients is associated with increased expression of PlGF and its downstream genes in respiratory epithelial cells. In an SCD mouse model, we observed increased AHR and higher leukotriene levels that were abrogated by anti-PlGF Ab or the 5-lipoxygenase inhibitor zileuton. Overall, our findings indicate that PlGF exacerbates AHR and uniquely links the leukotriene and Th2 pathways in asthma. These data also suggest that zileuton and anti-PlGF Ab could be promising therapies to reduce pulmonary morbidity in SCD.