Inhibition Of Lysis Research Articles

Hypoxic Stress Induces Complement-Mediated Lysis of Mesenchymal Stem Cells by Downregulating Factor H and CD59.

Factor H and membrane inhibitor of reactive lysis (CD59) are key regulators of complement activation. Mesenchymal stem cells (MSCs) secrete Factor H and express CD59 to protect themselves from complement-mediated damage. Severe hypoxia found to decrease the survival chances of MSCs after transplantation; however, little is known about the impact of severe hypoxia on modulating the complement system activity and its effect on MSCs survival. Our study seeks to explore the effect of severe hypoxia on modulating the complement cascade in MSCs. Human adipose tissue-derived MSCs (hAD-MSCs) were cultured under severe hypoxia using 400μM Cobalt Chloride (CoCl2) for 48h. The protein expressions of survival marker; Phosphoinositide 3-kinases (PI3K), and pro-apoptotic marker; Caspase-3 were assessed using western blotting. The level of complement system related factors; Factor H, CD59, C3b, iC3b, C5b, C9, and the complement membrane attack complex (MAC) were analyzed using Elisa assays, western blotting, and immunocytochemistry. Our results showed for the first time that severe hypoxia can significantly impair Factor H secretion and CD59 expression in MSCs. This has been associated with upregulation of MAC complex and increased level of cell lysis and apoptosis marked by downregulation of PI3K and upregulation of Annexin v and Caspase-3. The loss of Factor H and CD59 in hypoxic MSCs can initiate their lysis and apoptosis mediated by activating MAC complex. Preserving the level of Factor H and CD59 in MSCs has significant clinical implication to increase their retention rate in hypoxic conditions and prolong their survival.

Biotoxicity Evaluation of Biodegradable Polymeric Particles: Exploring the Possible Adverse Impacts on Biological Systems

ABSTRACTThis study aimed to investigate the impact of plastic particles on cell viability and tissue toxicity. The cells were subjected to incubation with plastic particles immobilized in agar gels, resulting in minimal cell death and cytotoxicity. Furthermore, direct exposure of various cell types to suspended plastic particles showed negligible effects on cell viability, with no observed lysis or growth inhibition. Staining techniques revealed minimal plastic particle‐induced cell death, with the majority of cells remaining viable. However, histological evaluations of mouse tissues demonstrated that the nondegradable low‐density poly(ethylene) (LDPE) groups exhibited severe irritation, characterized by an excessive presence of macrophages, neutrophils, fibrosis, fat infiltration, and increased blood vessel formation in subcutaneous tissue. In contrast, biodegradable neat poly(butylene succinate) (KRICT‐PBS) and cellulose nanocrystals/PBS nanocomposite (CNC‐PBS) groups induced minimal toxicity. Interestingly, the intravenous injection of KRICT‐PBS and CNC‐PBS degradate solutions in mice exhibited mild toxicity, suggesting no significant damage to normal kidney and liver function.

Investigating the role of allelochemicals in the interaction between Alexandrium monilatum and other phytoplankton species

Species of the dinoflagellate genus Alexandrium can release bioactive extracellular compounds with allelopathic effects (e.g., immobilization, inhibition of growth, photosynthesis or lysis) towards other phytoplanktonic organisms. In the lower Chesapeake Bay, Virginia, US, succession or co-occurrence of blooms of Akashiwo sanguinea, Margalefidinium polykrikoides and the goniodomin-producing A. monilatum tend to be common during summer months, however the allelopathic potential of A. monilatum, and how it may affect bloom dynamics, has not be studied. We used a rapid fluorescence-based bioassay as well as flow cytometry and an assessment of immobilization to determine the potential effects of A. monilatum culture supernatants on M. polykrikoides, A. sanguinea and the diatom Chaetoceros muelleri (included as a reference strain). In addition, we also investigated the effects of standards of known A. monilatum toxins goniodomin A (GDA) and GDA seco-acid (GDA-sa). Exposure of C. muelleri to culture supernatants from two different strains of A. monilatum resulted in an inhibition of the maximum quantum yield of the photosystem II (Fv/Fm) and cell lysis of the diatom, with strain-specific variation observed, highlighted by a 30-fold difference in the effective concentration resulting in 10 % inhibition (EC10) and 2-fold difference in the lethal concentration resulting in 50 % mortality (LC50) between the two A. monilatum strains tested. Exposure of M. polykrikoides to A. monilatum culture supernatants resulted in a decrease in motility (at ≥ 500 eq. cells mL-1) along with a decrease in Fv/Fm (at ≥ 1,500 cells mL-1) and altered cellular morphology (at ≥ 3,500 eq. cells mL-1). No effect was observed when A. sanguinea was exposed to A. monilatum culture supernatants (at the concentrations tested in these studies). When M. polykrikoides was exposed to the GDA standard, its Fv/Fm (at ≥ 1,951 nM) and motility (at ≥ 390 nM) decreased, and its morphology (at ≥ 975 nM) was modified. The study of supernatant time- and temperature-stability, and the absence of a relationship between observed effects and goniodomin concentrations suggested the presence of additional unknown allelochemicals distinct from goniodomins, as has been observed for other Alexandrium species. Immobilization of M. polykrikoides by A. monilatum culture supernatants and GDA standard in laboratory-based exposure experiments may indicate that A. monilatum has a competitive advantage when both species co-occur in the Chesapeake Bay and this warrants further testing.

Competitive advantages of T-even phage lysis inhibition in response to secondary infection.

T-even bacteriophages are known to employ lysis inhibition (LIN), where the lysis of an infected host is delayed in response to secondary adsorptions. Upon the eventual burst of the host, significantly more phage progenies are released. Here, we analysed the competitive advantage of LIN using a mathematical model. In batch culture, LIN provides a bigger phage yield at the end of the growth where all the hosts are infected due to an exceeding number of phage particles and, in addition, gives a competitive advantage against LIN mutants with rapid lysis by letting them adsorb to already infected hosts in the LIN state. By simulating plaque formation in a spatially structured environment, we show that, while LIN phages will produce a smaller zone of clearance, the area over which the phages spread is actually comparable to those without LIN. The analysis suggests that LIN induced by secondary adsorption is favourable in terms of competition, both in spatially homogeneous and inhomogeneous environments.

Lysis Physiology of Pseudomonas aeruginosa Infected with ssRNA Phage PRR1.

The phage PRR1 belongs to the Leviviridae family, a group of ssRNA bacteriophages that infect Gram-negative bacteria. The variety of host cells is determined by the specificity of PRR1 to a pilus encoded by a broad host range of IncP-type plasmids that confer multiple types of antibiotic resistance to the host. Using P. aeruginosa strain PAO1 as a host, we analyzed the PRR1 infection cycle, focusing on cell lysis. PRR1 infection renders P. aeruginosa cells sensitive to lysozyme approximately 20 min before the start of a drop in suspension turbidity. At the same time, infected cells start to accumulate lipophilic anions. The on-line monitoring of the entire infection cycle showed that single-gene-mediated lysis strongly depends on the host cells' physiological state. The blockage of respiration or a reduction in the intracellular ATP concentration during the infection resulted in the inhibition of lysis. The same effect was observed when the synthesis of PRR1 lysis protein was induced in an E. coli expression system. In addition, lysis was strongly dependent on the level of aeration. Dissolved oxygen concentrations sufficient to support cell growth did not ensure efficient lysis, and a coupling between cell lysis initiation and aeration level was observed. However, the duration of the drop in suspension turbidity did not depend on the level of aeration.

MICROWAVE AIDED SYNTHESIS OF ZINC OXIDE NANOCERAMICS: A POTENT BIOACTIVE AGENTS WITH THERAPEUTIC CHARACTERS

Nanoparticles have been inexhaustibly utilized in Nanochemistry cohere with Bio-Technology to upgrade the immobilization and movement of catalysts in pharmaceutical nanoengineering for conveyance of helpful specialists in incessant illness diagnostics and in sensors.Herein communication we report microwave irradiation technique implemented for the preparation of novel Zinc oxide nanoparticles with beneficial bio-efficacy wherein the method employed has the advantages of producing small particle size metal oxide with high purity owing to short reaction time. We successfully synthesized and characterized two Zinc oxide based Cs2 doped ZnO2 and Li2 doped ZnO2 nanoparticles and were subjected to in vitro free radical scavenging assay wherein all the particles could scavenge DPPH and Nitric oxide radicals with less IC50 value over the positive control. Microbicidal properties of nanoparticles were proved by its effectiveness against both Gram positive and Gram negative bacteria tested besides its action against Candida albicans. Both nanoparticles synthesized showed a prominent inhibition of erythrocyte lysis by PhospholipaseA2 in a concentration dependent manner indicating better anti-inflammatory agent with possibility of preventing cancer. In this study we demonstrate by shell less CAM assay that synthesized nanoparticles could inhibit in vivo angiogenesis which would otherwise under abnormal conditions causes the acceleration of several inflammatory diseases in spite of its role in normal growth and wound healing process. Inhibition of such process is the promising methodology to hinder the progression of diseases. Present characteristic features make use of ZnO2 based doped nanoparticles to be a possible approach as a therapeutic molecule with favourable biological performance.

Abstract 39: Preclinical in vitro and in vivo evaluation of CD8αβ+ CD19 CAR iPSCs T cells generated in a TCR signal-independent manner using DLL4/VCAM-coupled microbeads

Abstract The use of clonally-derived, induced pluripotent stem cells (iPSCs) as starting material for therapeutic T cell manufacturing would overcome many limitations of autologous Chimeric Antigen Receptor T cell (CAR-T) therapies. Complex, multi-stage genome engineering, large batch production and rigorous lot testing could provide a consistent source of off-the-shelf, functionally enhanced T cell products. However, this vision cannot be realized using existing T cell differentiation platforms, which fail to present Notch ligands with the precision and intensity required to control T cell differentiation in scalable, suspension culture. We have previously shown that VCAM1 synergizes with DLL4 to enhance Notch signaling and progenitor T cell differentiation. We have extended the utility of this discovery through the creation of DLL4/VCAM1-coated magnetic microbeads, enabling precise and temporal control of Notch signaling in suspension culture. iPSC-derived CD34+ cells cultured with DLL4/VCAM1 beads demonstrate dose-responsive activation of Notch gene expression (e.g., Notch1, DTX1, TCF7), resultant T lineage commitment (e.g. CD5, CD7 expression) and maturation to CD4-CD8+ (single positive) cells in a TCR signal-independent manner. Removal of the endogenous TCR is necessary to prevent graft-versus-host-disease. Therefore, differentiation and maturation in a TCR-independent process presents substantial advantage in the development of safe, off the shelf iPSC T cell products. Using a clonal iPSC line with a CD19 CAR knocked in at the TRAC locus, we apply this approach to generate CAR-expressing, CD8αβ+ functional T cells, capable of multiple rounds of in vitro tumor cell lysis and sustained tumor growth inhibition in vivo comparable to primary T cells. This advancement demonstrates proof-of-concept for generation of highly efficacious, off-the-shelf CAR-T cells using novel, small-footprint manufacturing in order to broaden the applicability and accessibility of T cell therapies for cancer patients. Citation Format: Muluken S. Belew, Richard Carpenedo, Alessia Pallaoro, Hiofan Hoi, Avisek Deyati, Daniel C. Kirouac, Siddarth Chandrasekaran, Valerie Wall, Libin Abraham, Sommer Apelu, Michael H. Cadell, Amanda AuYeung, Omar Subedar, Elisa Martinez, Shri Joshi, Elizabeth Csaszar, Steven Woodside, Emily Titus, Christopher Bond. Preclinical in vitro and in vivo evaluation of CD8αβ+ CD19 CAR iPSCs T cells generated in a TCR signal-independent manner using DLL4/VCAM-coupled microbeads [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 39.

A dimeric holin/antiholin complex controls lysis by phage T4.

Lytic phages control the timepoint of host cell lysis by timing the holin-mediated release of cell wall-degrading endolysins. In phage T4, the antiholin RI inhibits the holin T, thereby preventing the early release of the T4 endolysin and lysis. The antiholin achieves lysis inhibition (LIN) in response to phage superinfections, thereby increasing the chance for lysis in an environment with a lower phage concentration. The holin T consists of a small N-terminal cytoplasmic domain, a transmembrane helix, and a periplasmic C-terminal domain. The antiholin is targeted to the periplasm by a cleavable signal peptide. Recently, the periplasmic soluble domains of the holin and the antiholin were found to form T2/RI2 tetramers in crystals. To investigate the functional relevance of this complex, we reconstituted LIN in a phage-free system, using only RI, T, and endolysin, and combined targeted mutagenesis with functional analyses. Inactivation of the RI signal peptide cleavage site did not abolish LIN, indicating that RI can function in a membrane-bound state, which argued against the tetramer. This led to analyses showing that only one of the two T/RI interfaces in the tetramer is physiologically relevant, which is also the only interaction site predicted by AlphaFold2. Some holin mutations at this interaction site prevented lysis, suggesting that the RI interaction likely acts by blocking the holin oligomerization required for hole formation. We conclude that LIN is mediated by a dimeric T/RI complex that, unlike the tetramer, can be easily formed when both partners are membrane-anchored.

Inhibition of oenocytoid lysis of the common cutworm Spodoptera litura (Lepidoptera: Noctuidae) by carbon dioxide gas

Inhibition of oenocytoid lysis of the common cutworm Spodoptera litura (Lepidoptera: Noctuidae) by carbon dioxide gas

Synergism of red blood cells and tranexamic acid in the inhibition of fibrinolysis

BackgroundPostpartum hemorrhage (PPH) is the leading cause of maternal death worldwide. The World Maternal Antifibrinolytic trial showed that antifibrinolytic tranexamic acid (TXA) reduces PPH deaths. Maternal anemia increases the risk of PPH. The World Maternal Antifibrinolytic-2 trial is now assessing whether TXA can prevent PPH in women with anemia. Low red blood cell (RBC) counts promote fibrinolysis by altering fibrin structure and plasminogen activation. ObjectivesWe explored interactions between RBCs and TXA in inhibiting fibrinolysis. MethodsWe used global fibrinolytic assays (ball sedimentation and viscoelasticity) to monitor the lysis of fibrin containing plasminogen and tissue-type plasminogen activator. We applied a fluorogenic kinetic assay to measure plasmin generation in fibrin clots and scanning electron microscopy to study fibrin structure. ResultsAccording to parallel-line bioassay analysis of the fibrin lysis-time data, the antifibrinolytic potency of 4-128 μM TXA was increased in the presence of 10% to 40% (v/v) RBCs. Global fibrinolysis assays showed that the joint effect of RBCs and TXA was about 15% larger than the sum of their individual effects in the inhibition of fibrinolysis. In plasminogen activation, TXA added the same increment of inhibition to the effect of RBCs at any cell count in the fibrin clot. Regarding fibrin structure, TXA thickened fibrin fibers, which impaired plasminogen activation, whereas RBCs promoted fine fibers that were more resistant to plasmin. ConclusionsThe antifibrinolytic potency of TXA is enhanced in fibrin formed in the presence of RBCs through inhibition of plasminogen activation and fibrin lysis, which correlates with modifications of fibrin structures.

Muscimol inhibits plasma membrane rupture and ninjurin-1(NINJ1) oligomerization during pyroptosis

Pyroptosis is a cell death process that causes inflammation and contributes to numerous diseases. Pyroptosis is mediated by caspase-1 family proteases that cleave the pore-forming protein gasdermin D, causing plasma membrane rupture and release of pathogenic cellular contents. We previously identified muscimol as a small molecule that prevents plasma membrane rupture during pyroptosis via an unidentified mechanism. Here, we show that muscimol has reversible activity to prevent cellular lysis without affecting earlier pyroptotic events. Although muscimol is a well-characterized agonist for neuronal GABAA receptors, muscimol protection is not altered by GABAA receptor antagonists or recapitulated by other GABAA agonists, suggesting that muscimol acts via a novel mechanism. We find that muscimol blocks oligomerization of ninjurin-1, which is required for plasma membrane rupture downstream of gasdermin D pore formation. Our structure-activity relationship studies reveal distinct molecular determinants defining inhibition of pyroptotic lysis compared to GABAA binding. In addition, we demonstrate that muscimol reduces lethality during LPS-induced septic shock. Together, these findings demonstrate that ninjurin-1-mediated plasma membrane rupture can be pharmacologically modulated and pave the way toward identification of therapeutic strategies for pathologic conditions associated with pyroptosis.

Slow growing bacteria survive bacteriophage in isolation

The interactions between bacteria and bacteriophage have important roles in the global ecosystem; in turn changes in environmental parameters affect the interactions between bacteria and phage. However, there is a lack of knowledge on whether clonal bacterial populations harbour different phenotypes that respond to phage in distinct ways and whether the abundance of such phenotypes within bacterial populations is affected by variations in environmental parameters. Here we study the impact of variations in nutrient availability, bacterial growth rate and phage abundance on the interactions between the phage T4 and individual Escherichia coli cells confined in spatial refuges. Surprisingly, we found that fast growing bacteria survive together with all of their clonal kin cells, whereas slow growing bacteria survive in isolation. We also discovered that the number of bacteria that survive in isolation decreases at increasing phage doses possibly due to lysis inhibition in the presence of secondary adsorptions. We further show that these changes in the phenotypic composition of the E. coli population have important consequences on the bacterial and phage population dynamics and should therefore be considered when investigating bacteria-phage interactions in ecological, health or food production settings in structured environments.

Relative Free Radical Scavenging, Tyrosinase Inhibitory, and Anti-Inflammatory Potential of the Polar Peel decoctures of distinct Malus Species indigenous to Kashmir

Apple, (Malus domestica, Brokh), is a vital source of phenolic and polyphenolic compounds whose accumulation in the different parts of the fruit depends upon various intrinsic and extrinsic factors. This study aimed to perform a comparative evaluation of phenolics of peels of three red-skinned (“Delicious”, “Maharaji”, “Royal delicious”), and one yellow-skinned (“Golden Delicious”) apple varieties incorporating preliminary secondary metabolite perusal followed by estimation of Total phenol and total flavonoid contents. Secondary phytocompounds being usually polar in nature were extracted from peels via the Soxhlet and hot maceration processes using different menstruum systems of varied polarity. Comparative assessment of Free radical scavenging, anti-pigmentation, and anti-inflammatory properties of all the decoctures was analyzed thereafter.Free radical scavenging capacity was analyzed via 2, 2-diphenyl-1-picrylhydrazyl assay.Tyrosinase inhibitory activity was assessed by studying the in vitro impact of decoctures on the monophenolase and diphenolase reactions of the enzyme tyrosinase. The anti-inflammatory potential of the samples was assessed by inhibition of hypotonicity-induced HRBC membrane lysis. TPC/TFC estimation was analyzed via spectrophotometry. Methanol crude peel extracts of all apple species revealed rich phytochemical content as per TPC/TFC analysis. In a dose-dependent manner, all the crude peel extracts demonstrated considerable free radical scavenging capacity with DPPH assay. Maharaji methanolic decocture revealed potent free radical scavenging activity by showing the lowest IC50 value while Royal delicious acetone extract showed the highest IC50 value when collated with the potent reference antioxidant ascorbic acid with an IC50value of 114.62µg/ml. Polarextract of Malus domestica var Maharaji revealed significant tyrosinase inhibitory activity by revealing IC50 of 8460.65µg/mlon L-Tyrosine substrate and IC50 of 7830.46 µg/ml on L-DOPA substrate. Malus peel compounds turn out to be more effective in diphenolase reaction and kojic acid acts more effectively in the monophenolase one.At various concentrations, the percentage of membrane stabilization for methanolic extracts and the reference NSAID diclofenac sodium was determined.The highest level of membrane stabilization wasshown by methanolic Maharaji and delicious apple peel extracts (83.76 % and 72.32% at a dose of 200 mg/ml respectively) when juxtaposed with standard NSAID showing 98.58 % inhibition at a concentration of 25 mg/ml. Methanol extracts of Malus domestica var Maharaji out performed the other Malus peel extracts in terms of all the parameters analyzed, according to the current study. So, this study is giving the highest pharmaceutical priority to the Malus domestica var Maharaji when it comes to human-health benign bioactive compounds, oxidative stress-related diseases, pigmentation disorders, and inflammatory diseases.

Isolation, Characterization, and Comparative Genomic Analysis of Bacteriophage Ec_MI-02 from Pigeon Feces Infecting Escherichia coli O157:H7.

The most significant serotype of Shiga-toxigenic Escherichia coli that causes foodborne illnesses is Escherichia coli O157:H7. Elimination of E. coli O157:H7 during food processing and storage is a possible solution. Bacteriophages have a significant impact on bacterial populations in nature due to their ability to lyse their bacterial host. In the current study, a virulent bacteriophage, Ec_MI-02, was isolated from the feces of a wild pigeon in the United Arab Emirates (UAE) for potential future use as a bio-preservative or in phage therapy. Using a spot test and an efficiency of plating analysis, Ec_MI-02 was found to infect in addition to the propagation host, E. coli O157:H7 NCTC 12900, five different serotypes of E. coli O157:H7 (three clinical samples from infected patients, one from contaminated green salad, and one from contaminated ground beef). Based on morphology and genome analysis, Ec_MI-02 belongs to the genus Tequatrovirus under the order Caudovirales. The adsorption rate constant (K) of Ec_MI-02 was found to be 1.55 × 10-8 mL/min. The latent period was 50 min with a burst size of almost 10 plaque forming units (pfu)/host cell in the one-step growth curve when the phage Ec_MI-02 was cultivated using the propagation host E. coli O157:H7 NCTC 12900. Ec_MI-02 was found to be stable at a wide range of pH, temperature, and commonly used laboratory disinfectants. Its genome is 165,454 bp long with a GC content of 35.5% and encodes 266 protein coding genes. Ec_MI-02 has genes encoding for rI, rII, and rIII lysis inhibition proteins, which supports the observation of delayed lysis in the one-step growth curve. The current study provides additional evidence that wild birds could also be a good natural reservoir for bacteriophages that do not carry antibiotic resistance genes and could be good candidates for phage therapy. In addition, studying the genetic makeup of bacteriophages that infect human pathogens is crucial for ensuring their safe usage in the food industry.

Autoinduction AND Gate Inhibits Cell Lysis to Enhance Protein Production in Bacillus subtilis Controlled by Population Density and Cell Physiological State.

The extracellular protease-deficient strain Bacillus subtilis WB600 is commonly used as a chassis cell for the production of industrial proteins. However, B.subtilis WB600 exhibits an increased susceptibility to cell lysis and a reduction in biomass. Inhibition of cell lysis by knocking out lytic genes will impair physiological function. Here, we dynamically inhibited cell lysis in B.subtilis WB600 to balance the impairment of physiological function with the accumulation of biomass. First, the inducible protein degradation systems (IPDSs) were constructed and used to investigate the effects of inhibiting cell lysis on biomass, cell morphology, and protein production at different times (using pullulanase as a test). The highest pullulanase activity was obtained at 20 h of inhibiting cell lysis, 184.8 U/mL, which was 44% higher than the activity of B.subtilis WB600. Then, to avoid addition of inducers, we introduced orthogonal quorum sensing and constructed autoinduction protein degradation systems (AIPDSs). The optimized AIPDS showed similar pullulanase activity to the optimal IPDS (20 h), 181.3 U/mL. Next, we constructed dual-signal input autoinduction protein degradation systems (DSI-AIPDSs) via AND gate to further address two deficiencies of AIPDS, one-time activation and damage to new cells. These DSI-AIPDSs were controlled by quorum sensing and stationary phase promoters that respond to population density and single-cell physiological state, respectively. Finally, the OD600 and pullulanase activity of the strain with optimal DSI-AIPDS were 51% and 115% higher than those of B.subtilis WB600 in pullulanase production, respectively. We provided a B.subtilis chassis strain with considerable potential for biomass accumulation and enhanced protein production.

Anti-inflammatory, Antinociceptive, and Toxicological Properties of Uvaria comperei Stem Crude Extract and Fractions.

The present study was carried out to investigate the anti-inflammatory activity of a methanolic extract and fractions of Uvaria comperei stems. The crude extract was obtained by maceration of the powder in methanol and fractions by vacuum chromatography from the methanolic extract. To study the anti-inflammatory activity in vitro, red blood cell lysis inhibition assay and albumin denaturation inhibition were performed, while in vivo measurements of carrageenan-induced paw oedema and formalin-induced pain in albino mice were performed. Acute toxicity and cytotoxicity studies of the fraction F2 were performed, as well as its HPLC, and some biochemical parameters were quantified. Uvaria comperei crude extract (UCCE) at 250 and 500 μg/mL completely inhibited albumin denaturation, while decreasing 75.5% of heat blood cell lysis at 500 μg/mL. The fractions 128-136 (F3), 10-11 (F1), and 56-62 (F2) at 500 μg/mL displayed a significant anti-inflammatory activity with percentages of inhibition of 60.5, 67.4, and 100%, respectively. Administration of fraction F2 (25, 50, and 100 mg/kg, p.o.) produced a dose-dependent inhibition of formalin-induced pain of 60.2% at 50 mg/kg in the neurogenic phase (p < 0.05) and 70.2% at 25 mg/kg in the inflammatory phase (p < 0.05). Similarly, the time-dependent increase in carrageenan-induced paw circumference induced by carrageenan was inhibited by pretreatment with F2: 50% of inhibition at 25 mg/kg after 30 min (p < 0.05) and 96.5% inhibition at 25 mg/kg after 6 h (p < 0.05). In this research, the fraction F2 presented its maximum analgesic property at 50 mg/kg, while it presented the highest anti-inflammatory property at 25 mg/kg. The oral lethal median dose (LD50) of F2 was determined to be greater than 2000 mg/kg; further low cytotoxicity in RAW cells was also observed. Overall, this work shows that the methanolic crude extract and fractions, mainly F2, of Uvaria comperei stem have interesting anti-inflammatory properties.

An Evaluation of the Complement-Regulating Activities of Human Complement Factor H (FH) Variants Associated With Age-Related Macular Degeneration.

Factor H (FH, encoded by CFH) prevents activation of the complement system's alternative pathway (AP) on host tissues. FH impedes C3 convertase (C3bBb) formation, accelerates C3bBb decay, and is a cofactor for factor I (FI)-catalyzed C3b cleavage. Numerous CFH variants are associated with age-related macular degeneration (AMD), but their functional consequences frequently remain undetermined. Here, we conduct functional comparisons between a control version of FH (not AMD linked) and 21 AMD-linked FH variants. Recombinantly produced, untagged, full-length FH versions were assayed for binding to C3b and decay acceleration of C3bBb using surface-plasmon resonance, FI-cofactor activity using a fluorescent probe of C3b integrity, suppression of C5b-9 assembly on an AP-activating surface, and inhibition of human AP-mediated lysis of sheep erythrocytes. All versions were successfully purified despite below-average yields for Arg2Thr, Arg53Cys, Arg175Pro, Arg175Gln, Ile221Val, Tyr402His, Pro503Ala, Arg567Gly, Gly1194Asp, and Arg1210Cys. Compared to control FH, Arg2Thr, Leu3Val, Ser58Ala, Asp90Gly, Asp130Asn, Gln400Lys, Tyr402His, Gly650Val, Ser890Ile, and Thr965Met showed minimal functional differences. Arg1210C, Arg53His, Arg175Gln, Gly1194Asp, Pro503Ala, Arg53Cys, Arg576Gly, and Arg175Pro (in order of decreasing efficacy) underperformed, while Ile221Val, Arg303Gln, and Arg303Trp were "marginal." We newly identified variants toward the center of the molecule, Pro503Ala and Arg567Gly, as potentially pathogenic. Our approach could be extended to other variants of uncertain significance and to assays for noncanonical FH activities, aiming to facilitate selection of cohorts most likely to benefit from therapeutic FH. This is timely as recombinant therapeutic FH is in development for intravitreal treatment of AMD in patients with reduced FH functionality.

In Vitro Metabolite Profiling and Anti-Inflammatory Activities of Rhodomyrtus Tomentosa with Red Blood Cell Membrane Stabilization Methods.

Rhodomyrtus Tomentosa (Karamunting) is one of South Kalimantan's biodiversity. In previous research on asthma and coal dust exposure models, nebulization with an ethanol extract of R. tomentosa leaves has been shown to reduce inflammatory cells. This study aimed to investigate the anti-inflammatory activity on the stabilization of red blood cell membranes and to identify the chemical compounds present in extracts of R. tomentosa leaves. Anti-inflammatory effect of R. tomentosa leaves was evaluated by in vitro red blood cell membrane stabilization methods. Nonsteroidal anti-inflammatory sodium diclofenac was used as the reference drug. The content of chemical compounds in the ethanol extract of R. tomentosa leaves was identified using Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS). The results of inhibition of red blood cells membrane lysis showed the n-hexane fraction (concentration 25 ppm), ethyl acetate fraction (concentration 50 ppm), and a fraction of water (concentration 50 ppm) with an inhibition level of 54.5%, 81.8%, 63.6%, respectively. The ethyl acetate fraction showed the highest inhibition of hemolysis. This result is not significantly different from the standard anti-inflammatory sodium diclofenac (90.09%). Oleanonic acid and ursonic acid were two similar metabolites in subfractions ethyl acetate 1, 2, and 3, which may have anti-inflammatory properties. R. Tomentosa leaves can have potency as an anti-inflammatory by increasing the red blood cell membrane stability equal to lysosome cells, depending on the concentration.

Efficacy of Trevesia palmata (Roxb. ex Lindl.) Vis. Extract on MG 63 cell lines and arthritis-induced animal models

Ethnopharmacological relevanceDespite widespread use of herbal remedies for treating arthritis and osteosarcoma, many plants are still not pharmacologically evaluated for their efficacy. Contrary to many non-steroidal, immunosuppressants, antibiotics, and antineoplastic drugs that have adverse effects, phytotherapeutic compounds have promising benefits with fewer complications. In this study the unexplored Northeastern India indigenous plant Trevesia palmata (Roxb. ex Lindl.) Vis. used in traditional medicine to cure bone fractures is chosen for studying anti-proliferative and anti-rheumatic properties. Aim of the studyThis study designed to explore the polyphenolic composition, antioxidant, anti-inflammatory and anti-arthritic potential of T. palmata leaf extracts. Further, the cellular activity was studied using MG 63 osteoblast cell lines and pharmacologically evaluated using Complete Freund's Adjuvant (CFA) induced arthritic rat model. Materials and methodsIn vitro free radical scavenging activity, anti-inflammatory and anti-arthritic activities of extracts were analyzed using standardized methods. The polyphenolic profiling and apoptosis inducing ability of T. palmata ethyl acetate (TPEA) extract on MG 63 osteoblast cell lines were analyzed. The in vivo pharmacological studies were carried out with low dose 250 mg/kg and high dose of 500 mg/kg of T. palmata. The biochemical and haematological parameters and in vivo antioxidant activity were evaluated for the control and treated groups. Radiological and histological study were done to understand the impact and penetration of inflammatory arthritis from tissues to joint bones. ResultsTPEA showed highest free radical scavenging activity (DPPH - 4.72 IC50, ABTS - 242.33 ± 6.81 mM TE/g extract), anti-inflammatory (40.04% inhibition of RBC lysis) and anti-arthritic activity (32.4% inhibition of protein denaturation) with the presence of gallic acid, catechin, caffeic acid, rutin, quercetin and naringenin. The TPEA extract inhibited cell proliferation of MG 63 osteoblast cells and induced apoptosis by arresting cell cycle at different phases. After acute toxicity studies the doses 250 mg/kg and 500 mg/kg were fixed and showed better results in CFA-induced arthritic animals. Thus, the extract phytoconstituents may have immense potential against chronic inflammation, joint ailments, bone cancer and arthritis which serves as a phytomedicine contrary to synthetic medications. ConclusionsThe potential treatment of polyphenolic compounds in the T. palmata extract on osteosarcoma and arthritis was demonstrated from this study. Thus, cellular inflammatory infiltrates are significantly reduced in bone and joint tissues as well.

Interaction between Phage T4 Protein RIII and Host Ribosomal Protein S1 Inhibits Endoribonuclease RegB Activation.

Lytic viruses of bacteria (bacteriophages, phages) are intracellular parasites that take over hosts’ biosynthetic processes for their propagation. Most of the knowledge on the host hijacking mechanisms has come from the studies of the lytic phage T4, which infects Escherichia coli. The integrity of T4 development is achieved by strict control over the host and phage processes and by adjusting them to the changing infection conditions. In this study, using in vitro and in vivo biochemical methods, we detected the direct interaction between the T4 protein RIII and ribosomal protein S1 of the host. Protein RIII is known as a cytoplasmic antiholin, which plays a role in the lysis inhibition function of T4. However, our results show that RIII also acts as a viral effector protein mainly targeting S1 RNA-binding domains that are central for all the activities of this multifunctional protein. We confirm that the S1–RIII interaction prevents the S1-dependent activation of endoribonuclease RegB. In addition, we propose that by modulating the multiple processes mediated by S1, RIII could act as a regulator of all stages of T4 infection including the lysis inhibition state.