Therapeutic hyperthermia regulates complement C3 activation and suppresses tumor development through HSPA5/NFκB/CD55 pathway in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is endemic in Southern China and Southeast Asia. Hyperthermia is widely used in combination with chemotherapy and radiotherapy to enhance therapeutic efficacy in NPC treatment, but the underlying anti-tumor mechanisms of hyperthermia remain unclear. Complement C3 has been reported to participate in the activation of immune system in the tumor microenvironment, leading to tumor growth inhibition. In this study, we aimed to explore the effect and mechanisms of hyperthermia and investigate the functional role of complement C3 in NPC hyperthermia therapy (HT). The serum levels of complement C3 before and after hyperthermia therapy in patients with NPC were analyzed. NPC cell lines SUNE1 and HONE1 were used for in vitro experiment to evaluate the function of complement C3 and HT on cell proliferation and apoptosis. SUNE1 xenograft mouse model was established and tumor-bearing mice were treated in water bath at a constant temperature of 43°C. Tumor samples were collected at different time points to verify the expression of complement C3 by immunohistochemical staining and western blot. The differential expressed genes after hyperthermia were analyzed by using RNA sequencing. We found that complement could enhance hyperthermia effect on suppressing proliferation and promoting apoptosis of tumor cells in NPC. Hyperthermia decreased the mRNA expression of complement C3 in tumor cells, but promoted the aggregation and activation circulating C3 in NPC tumor tissue. By using in vitro hyperthermia-treated NPC cell lines and SUNE1 xenograft tumor-bearing mice, we found that the expression of heat shock protein 5 (HSPA5) was significantly upregulated. Knockdown of HSPA5 abrogated the anti-tumor effect of hyperthermia. Moreover, we demonstrated that hyperthermia downregulated CD55 expression via HSPA5/NFκB (P65) signaling and activated complement cascade. Our findings suggest that therapeutic hyperthermia regulates complement C3 activation and suppresses tumor development via HSPA5/NFκB/CD55 pathway in NPC.

Tissue-Specific Gamma-Flicker Light Noninvasively Ameliorates Retinal Aging.

Aging is a risk factor for multiple retinal degeneration diseases. Entraining brain gamma oscillations with gamma-flicker light (γFL) has been confirmed to coordinate pathological changes in several Alzheimer's disease mouse models and aged mice. However, the direct effect of γFL on retinal aging remains unknown. We assessed retinal senescence-associated beta-galactosidase (ß-gal) and autofluorescence in 20-month-old mice and found reduced ß-gal-positive cells in the inner retina and diminished lipofuscin accumulation around retinal vessels after 6 days of γFL. In immunofluorescence, γFL was further demonstrated to ameliorate aging-related retinal changes, including a decline in microtubule-associated protein 1 light chain 3 beta expression, an increase in complement C3 activity, and an imbalance between the anti-oxidant factor catalase and pro-oxidant factor carboxymethyl lysine. Moreover, we found that γFL can increase the expression of activating transcription factor 4 (ATF4) in the inner retina, while revealing a decrease of ATF4 expression in the inner retina and positive expression in the outer segment of photoreceptor and RPE layer for aged mice. Western blotting was then used to confirm the immunofluorescence results. After mRNA sequencing (NCBI Sequence Read Archive database: PRJNA748184), we found several main mechanistic clues, including mitochondrial function and chaperone-mediated protein folding. Furthermore, we extended γFL to aged Apoe-/- mice and showed that 1-m γFL treatment even improved the structures of retinal-pigment-epithelium basal infolding and Bruch's membrane. Overall, γFL can orchestrate various pathological characteristics of retinal aging in mice and might be a noninvasive, convenient, and tissue-specific therapeutic strategy for retinal aging.

Saponins isolated from Radix polygalae extent lifespan by modulating complement C3 and gut microbiota.

With the increase in human lifespan, population aging is one of the major problems worldwide. Aging is an irreversible progressive process that affects humans via multiple factors including genetic, immunity, cellular oxidation and inflammation. Progressive neuroinflammation contributes to aging, cognitive malfunction, and neurodegenerative diseases. However, precise mechanisms or drugs targeting age-related neuroinflammation and cognitive impairment remain un-elucidated. Traditional herbal plants have been prescribed in many Asian countries for anti-aging and the modulation of aging-related symptoms. In general, herbal plants' efficacy is attributed to their safety and polypharmacological potency via the systemic manipulation of the body system. Radix polygalae (RP) is a herbal plant prescribed for anti-aging and the relief of age-related symptoms; however, its active components and biological functions remained un-elucidated. In this study, an active methanol fraction of RP containing 17 RP saponins (RPS), was identified. RPS attenuates the elevated C3 complement protein in aged mice to a level comparable to the young control mice. The active RPS also restates the aging gut microbiota by enhancing beneficial bacteria and suppressing harmful bacteria. In addition, RPS treatment improve spatial reference memory in aged mice, with the attenuation of multiple molecular markers related to neuroinflammation and aging. Finally, the RPS improves the behavior and extends the lifespan of C. elegans, confirming the herbal plant's anti-aging ability. In conclusion, through the mouse and C. elegas models, we have identified the beneficial RPS that can modulate the aging process, gut microbiota diversity and rectify several aging-related phenotypes.

Evaluation of electroacupuncture for symptom modification in a rodent model of spontaneous osteoarthritis.

OBJECTIVE: Faced with the frustration of chronic discomfort and restricted mobility due to osteoarthritis (OA), many individuals have turned to acupuncture for relief. However, the efficacy of acupuncture for OA is uncertain, as much of the evidence is inconclusive. The purpose of this study was to evaluate electroacupuncture (EA) in a rodent model of OA such that conclusions regarding its effectiveness for symptom or disease modification could be drawn. METHODS: Ten 12-month-old male Hartley guinea pigs-which characteristically have moderate to advanced OA at this age-were randomly assigned to receive EA for knee OA (n = 5) or anesthesia only (control group, n = 5). Treatments were performed three times weekly for 3 weeks, followed by euthanasia 2 weeks later. Gait analysis and enclosure monitoring were performed weekly to evaluate changes in movement. Serum was collected for inflammatory biomarker testing. Knee joints were collected for histology and gene expression. RESULTS: Animals receiving EA had significantly greater changes in movement parameters compared to those receiving anesthesia only. There was a tendency toward decreased serum protein concentrations of complement component 3 (C3) in the EA group compared to the control group. Structural and antioxidant gene transcripts in articular cartilage were increased by EA. There was no significant difference in total joint histology scores between groups. CONCLUSION: This study provides evidence that EA has a positive effect on symptom, but not disease, modification in a rodent model of OA. Further investigations into mechanistic pathways that may explain the efficacy of EA in this animal model are needed.

Synthesis and identification of a novel derivative of salidroside as a selective, competitive inhibitor of monoamine oxidase B with enhanced neuroprotective properties.

Salidroside [(2R,3S,4S,5R,6R)-2-(hydroxymethyl)-6-(4-hydroxyphenethoxy)tetrahy-dro-2H-pyran-3,4,5-triol] is an antioxidant, anti-inflammatory and neuroprotective agent, but its drug-like properties are unoptimized and its mechanism of actions is uncertain. We synthesized twenty-six novel derivatives of salidroside and examined them in CoCl2-treated PC12 cells using MTT assay. pOBz, synthesized by esterifying the phenolic hydroxyl group of salidroside with benzoyl chloride, was one of five derivatives that were more cytoprotective than salidroside, with an EC50 of 0.038 µM versus 0.30 µM for salidroside. pOBz was also more lipophilic, with log P of 1.44 versus -0.89 for salidroside. Reverse virtual docking predicted that pOBz would bind strongly with monoamine oxidase (MAO) B by occupying its entrance and substrate cavities, and by interacting with the inter-cavity gating residue Ile199 and Tyr435 of the substrate cavity. Enzymatic studies confirmed that pOBz competitively inhibited the activity of purified human MAO-B (Ki = 0.041 µM versus Ki = 0.92 µM for salidroside), and pOBz was highly selective for MAO-B over MAO-A. In vivo, pOBz inhibited cerebral MAO activity after middle cerebral artery occlusion with reperfusion in rats, and it reduced cerebral infarct volume, improved neurological function and NeuN expression, and inhibited complement C3 expression and apoptosis. Our results suggest that pOBz is a structurally novel type of competitive and selective MAO-B inhibitor, with potent neuroprotective properties after cerebral ischemia-reperfusion injury in rats.

Effectiveness of Bailing capsules in the treatment of lupus nephritis: A meta­analysis.

Previous studies have explored the treatment of lupus nephritis with Bailing capsules; however, due to limited sample sizes and inconsistent results across these studies, no definitive conclusions have been drawn. Thus, the present study aimed to provide evidence for the effectiveness of Bailing capsules in the treatment of lupus nephritis. To obtain relevant clinical studies (published before 20 July 2019), PubMed, Embase, Cochrane Library, China National Knowledge Infrastructure, WanFang and the Chinese Biomedical Literature Database were searched, and relevant studies concerning the use of Bailing capsules for treating lupus nephritis were selected. The extracted data were general characteristics such as the first author, publication year, study year, follow­up time, age, sex, course of the disease and a number of outcome indicators. These included systemic lupus erythematosus disease activity index (SLEDAI) score, serum albumin (Alb), 24­h urinary protein, serum creatinine, anti­ds­DNAIgM, complement component 3 (C3), and the number of effective treatments and complications. Meta­analysis was performed using R­3.12 software. Publication bias was assessed using Egger's test. A total of 14 studies comprising 1,301 participants were combined for analysis in the present study. The results demonstrated that with the exception of anti­ds­DNAIgM and complement C3, other indicators, such as SLEDAI score, Alb, 24­h urinary protein, serum creatinine, and the number of effective treatments and complications) in the Bailing capsule treatment group were improved compared with those in the control group. The results of the present meta­analysis suggested that Bailing capsules may be effective in the treatment of lupus nephritis.

Effect of Hyperbaric Oxygen on Tissue Damage and Expression of Adhesion Molecules and C3 in a Rat Model of Renal Ischemia-Reperfusion Injury After Kidney Transplantation.

BACKGROUND The aim of this study was to investigate the protective effect and mechanism of hyperbaric oxygen (HBO) in a rat model of renal ischemia-reperfusion injury following kidney transplantation. MATERIAL AND METHODS Sprague Dawley rats were randomly divided into 3 groups (n=18): sham group, kidney transplantation group, and HBO treatment group. Six rats in each group were sacrificed at 1, 3, and 5 hours after reperfusion, and serum and renal tissue were then collected. The serum creatinine levels and histopathological changes of the renal tissue were detected. ICAM-1, VCAM-1, and C3 expression levels were also detected by immunohistochemical staining or real-time polymerase chain reaction. RESULTS Renal function was damaged in the kidney transplantation group and the HBO treatment group compared with sham group (P<0.05). Renal histopathological changes, including tubular cell swelling, tubular dilatation, and hyaline casts, were remarkably reduced in the HBO treatment group compared to the kidney transplantation group. In the immunohistochemical examination, the expression levels of ICAM-1, VCAM-1, and C3 were obviously increased in the kidney transplantation group and the HBO treatment group; moreover, the levels in the HBO treatment group were significantly lower than in the kidney transplantation group (P<0.05). In addition, the ICAM-1 and C3 mRNA levels were increased in the kidney transplantation group and HBO treatment group, but the levels of in the HBO treatment group them were significantly decreased compared to the kidney transplantation group that at 3 and 5 hours after reperfusion (P<0.05). CONCLUSIONS HBO treatment exerted a protective effect on renal function through inhibition of adhesion molecule overexpression and complement system activation in a rat model of renal ischemia-reperfusion injury after kidney transplantation.

Uncovering the protective mechanism of Huoxue Anxin Recipe against coronary heart disease by network analysis and experimental validation.

Coronary heart disease (CHD) is a leading cause of death and disability worldwide. Huoxue Anxin Recipe (HAR) is a novel Chinese Herbal Medicine formula of that has been used to treat CHD for several decades. Our previous study found that HAR had anti-oxidative effects, and could promote myocardial angiogenesis and improve cardiac function following myocardial infarction (MI) in rats. However, the active compounds, potential targets, and biological processes related to HAR have not been systematically investigated. Here, network pharmacology and experimental validation were used to study the protective mechanisms of HAR against CHD. We identified 124 active components, 124 verified targets, and 111 predictive targets. A total of 1192 genes related to CHD were identified by cDNA microarray and database analysis. A total of 47 putative targets of HAR against CHD were identified, including 32 verified targets and 15 predictive targets. ClueGo enrichment analysis identified 49 biological processes involved in the anti-CHD effects of HAR. Among them, the negative regulation of blood coagulation and regulation of collagen biosynthetic process were experimentally validated. After constructing a protein-protein interaction network and clustering with MECODE and ClusterONE, 162 key proteins (from ClueGo and clustering) were used to construct an internal interaction network. Complement C3 (C3), Fibrinogen alpha (FGA), Fibrinogen gamma (FGG), interleukin-6 (IL6), and Apolipoprotein A1 (APOA1) were the top 5 hub proteins identified by cytoHubber analysis. HAR limited the concentrations of C3, FGA, FGG, and IL6 and increased APOA1 levels. The results indicated that HAR could down-regulate blood coagulation, regulate collagen biosynthesis, inhibit peroxidation and inflammation injury, and promote cholesterol efflux. HAR could be a potential source of novel and effective drugs for CHD.

GSPE alleviates renal fibrosis by inhibiting the activation of C3/ HMGB1/ TGF-ß1 pathway.

Grape seed proanthocyanidin extract (GSPE) has been reported to exhibit a variety of protective effects, such as antioxidant, anti-atherosclerosis and other pharmacological effects. As a member of the complement system, complement component 3 (C3) deposition in the glomerulus is recognized as an important causative mediator of various kidney diseases. In this study, we aimed to identify the effect of GSPE on C3 in the chronic kidney fibrosis and evaluate the possible mechanism. We observed that administration of GSPE relieves inflammation and chronic renal fibrosis in mouse models of UUO. GSPE inhibited C3 secreted by macrophages to relieve renal interstitial inflammation. In vitro, we found that C3 stimulated HMGB1 translocation form nucleus to cytoplasm and promote the expression of pro-inflammatory cytokines including TGF-ß1 in primary renal tubular epithelial cells (PTEC), which could be inhibited by GSPE. Meanwhile, GSPE could also decreased HMGB1-induced EMT of PTEC through suppresses the HMGB1/TLR4/p65/TGF-ß1 pathway. In addition, the myofibroblast activation was inhibited by GSPE via TGF-ß1/Smad2/3 signaling pathways in normal rat kidney fibroblast (NRK-49F) cells. Overall, these observations provide that GSPE alleviates renal fibrosis by inhibiting the C3/HMGB1/TGF-ß1 pathway and could thus lead to find the potential therapy for the suppression of renal fibrosis.

Omega-3 Fatty Acids Supplementation: Therapeutic Potential in a Mouse Model of Stargardt Disease.

Purpose: To evaluate the therapeutic effects of omega-3 (ω3) fatty acids on retinal degeneration in the ABCA4-/- model of Stargardt disease when the blood level of arachidonic acid (AA)/eicosapentaenoic acid (EPA) ratio is between 1 and 1.5. Methods: Eight-month-old mice were allocated to three groups: wild type (129S1), ABCA4-/- untreated, and ABCA4-/- ω3 treated. ω3 treatment lasted 3 months and comprised daily gavage administration of EPA and docosahexaenoic acid (DHA). Blood and retinal fatty acid analysis was performed using gas chromatography to adjust the blood AA/EPA ∼1 to 1.5. Eyecups were histologically examined using transmission electron microscopy and confocal microscopy to evaluate lipofuscin granules and the photoreceptor layer. Retinal N-retinylidene-N-retinylethanolamine (A2E), a major component of retinal pigment epithelium lipofuscin, was quantified using liquid chromatography and tandem mass spectrometry, in addition to retinal proteomic analysis to determine changes in inflammatory proteins. Results: EPA levels increased and AA levels decreased in the blood and retinas of the treatment group. Significantly less A2E and lipofuscin granules were observed in the treatment group. The thickness of the outer nuclear layer was significantly greater in the treatment group (75.66 ± 4.80 µm) than in the wild-type (61.40 ± 1.84 µm) or untreated ABCA4-/- (56.50 ± 3.24 µm) groups. Proteomic analysis indicated lower levels of complement component 3 (C3) in the treatment group, indicative of lower complement-induced inflammatory response. Conclusions: Three months of ω3 supplementation (AA/EPA ∼1-1.5) reduces A2E levels, lipofuscin granules, and C3 levels in the ABCA4-/- mouse model of Stargardt disease, consistent with slowing of the disease.

The MFHR1 Fusion Protein Is a Novel Synthetic Multitarget Complement Inhibitor with Therapeutic Potential.

The complement system is essential for host defense, but uncontrolled complement system activation leads to severe, mostly renal pathologies, such as atypical hemolytic uremic syndrome or C3 glomerulopathy. Here, we investigated a novel combinational approach to modulate complement activation by targeting C3 and the terminal pathway simultaneously. The synthetic fusion protein MFHR1 links the regulatory domains of complement factor H (FH) with the C5 convertase/C5b-9 inhibitory fragment of the FH-related protein 1. In vitro, MFHR1 showed cofactor and decay acceleration activity and inhibited C5 convertase activation and C5b-9 assembly, which prevented C3b deposition and reduced C3a/C5a and C5b-9 generation. Furthermore, this fusion protein showed the ability to escape deregulation by FH-related proteins and form multimeric complexes with increased inhibitory activity. In addition to substantially inhibiting alternative and classic pathway activation, MFHR1 blocked hemolysis mediated by serum from a patient with aHUS expressing truncated FH. In FH-/- mice, MFHR1 administration augmented serum C3 levels, reduced abnormal glomerular C3 deposition, and ameliorated C3 glomerulopathy. Taking the unique design of MFHR1 into account, we suggest that the combination of proximal and terminal cascade inhibition together with the ability to form multimeric complexes explain the strong inhibitory capacity of MFHR1, which offers a novel basis for complement therapeutics.

Effects of partial replacement of fish meal by yeast hydrolysate on complement system and stress resistance in juvenile Jian carp (Cyprinus carpio var. Jian).

A 10-week feeding trial was carried out to investigate the effects of dietary fish meal replacement by yeast hydrolysate (YH) on growth performance, complement system and stress resistance of juvenile Jian carp (Cyprinus carpio var. Jian) (initial average weight 19.44 ± 0.06 g). In the study, there were five groups: one control group was fed with a basal diet (YH0), and four treatment groups were fed with dietary fish meal replaced by 1% YH (YH1), 3% (YH3), 5% (YH5) and 7% (YH7), respectively. Each group had four replicates. At the end of feeding trial, twelve fish from each group (three fish per replicate) were randomly selected for assessing the growth and immunity. Meanwhile, 20 fish per replicate were injected by Aeromonas hydrophila. The results showed that (1) Replacement levels of YH significantly affected the growth of the fish with the highest values of weight gain (WG) occurred in fish fed YH3 diet. However, no significant difference in feed conversion ratios (FCR) was observed among all groups. (2) Pre-stressed plasma lysozyme activity, total protein and albumin contents and complement component 3 (C3) and complement component 4 (C4) levels of fish fed YH3 diet were significantly higher than those of fish fed YH0 diet. However, post-stressed immune parameters of fish in all groups were significantly lower. (3) There was a trend that the expression levels of the complement-related genes (c1r/s-A, c4-1, c3-H1, c5-1, fb/c2-A, mbl-2 and masp) initially increased and then decreased except mbl-2 and masp, with the maximum values observed in fish fed YH3 diet. Before stress, the expression levels of the inflammation-related genes (alp, il-1ß and tnf-α) in the hepatopancreas and spleen of fish fed YH1 diet and YH7 diet were significant higher than that of fish fed YH0 diet. After stress, no significant difference in the expression levels of those genes was observed among all groups. These results indicated that FM replacement by YH could improve growth performance, enhance innate immunity, and activate complement via the alternative complement pathway (ACP) and the classical complement pathway (CCP).

Targeted Intraceptor Nanoparticle for Neovascular Macular Degeneration: Preclinical Dose Optimization and Toxicology Assessment.

Neovascular age-related macular degeneration (AMD) is treated with anti-VEGF intravitreal injections, which can cause geographic atrophy, infection, and retinal fibrosis. To minimize these toxicities, we developed a nanoparticle delivery system for recombinant Flt23k intraceptor plasmid (RGD.Flt23k.NP) to suppress VEGF intracellularly within choroidal neovascular (CNV) lesions in a laser-induced CNV mouse model through intravenous administration. In the current study, we examined the efficacy and safety of RGD.Flt23k.NP in mice. The effect of various doses was determined using fluorescein angiography and optical coherence tomography to evaluate CNV leakage and volume. Efficacy was determined by the rate of inhibition of CNV volume at 2 weeks post-treatment. RGD.Flt23k.NP had peak efficacy at a dose range of 30-60 µg pFlt23k/mouse. Using the lower dose (30 µg pFlt23k/mouse), RGD.Flt23k.NP safety was determined both in single-dose groups and in repeat-dose (three times) groups by measuring body weight, organ weight, hemoglobin levels, complement C3 levels, and histological changes in vital organs. Neither toxicity nor inflammation from RGD.Flt23k.NP was detected. No side effect was detected on visual function. Thus, systemic RGD.Flt23k.NP may be an alternative to standard intravitreal anti-VEGF therapy for the treatment of neovascular AMD.

Lutein and atherosclerosis: Belfast versus Toulouse revisited.

In 1995 we reported that mean plasma lutein concentrations in salaried men and women from Toulouse in Southern France were double those in subjects recruited from general practitioner lists in Belfast, Northern Ireland. At the time incidence of coronary heart disease (CHD) in Southern France was among the lowest in Europe and was much higher in Northern Ireland. Plasma lutein is a biomarker of vegetable and fruit intake and evidence suggests that high concentrations are generally associated with better cardiometabolic health. At the time we speculated like others that role of the carotenoids may well have been to prevent oxidation of lipid in the lipoproteins and so reduce the uptake of oxidised lipid by macrophages and its deposition within the intimal layers of the major arteries as plaque. It is now widely accepted that CHD is an inflammatory disease and that macrophages within plaque together with tissue damage contribute to this inflammation. Stimulated macrophages release cytokines to activate the immune system both locally and systemically. Precursor complement proteins in the blood are activated to assist immune cells in phagocytosis and cell repair. Individuals with a history of arteriosclerosis display significantly higher concentrations of complement factors C3 and C3a than subjects without such a history. Metabolism of C3 via the alternate complement pathway can give rise to the membrane attack complex (MAC) which creates a hole or pore in pathogens or host cells, killing the cell. Recent studies in patients with early age related macular disease (AMD) who also exhibit similar elevated concentrations of complement proteins in their blood, showed supplementation with lutein progressively decreased the amount of the MAC and other complement factors in the blood. Lutein was used in the supplementation experiments because it is an important constituent of macular pigment. Thus the healthier cardiometabolic features displayed by the people in Toulouse may have been due to the effects of concurrent high concentrations of plasma lutein on the immune system and complement in particular. Other carotenoids may exert similar antioxidant effects but we and others found no differences in antioxidant nutrients between subjects in Toulouse and Belfast or between subjects with asymptomatic markers of atherosclerosis and controls.

From orphan drugs to adopted therapies: Advancing C3-targeted intervention to the clinical stage.

Complement dysregulation is increasingly recognized as an important pathogenic driver in a number of clinical disorders. Complement-triggered pathways intertwine with key inflammatory and tissue destructive processes that can either increase the risk of disease or exacerbate pathology in acute or chronic conditions. The launch of the first complement-targeted drugs in the clinic has undeniably stirred the field of complement therapeutic design, providing new insights into complement's contribution to disease pathogenesis and also helping to leverage a more personalized, comprehensive approach to patient management. In this regard, a rapidly expanding toolbox of complement therapeutics is being developed to address unmet clinical needs in several immune-mediated and inflammatory diseases. Elegant approaches employing both surface-directed and fluid-phase inhibitors have exploited diverse components of the complement cascade as putative points of therapeutic intervention. Targeting C3, the central hub of the system, has proven to be a promising strategy for developing biologics as well as small-molecule inhibitors with clinical potential. Complement modulation at the level of C3 has recently shown promise in preclinical primate models, opening up new avenues for therapeutic intervention in both acute and chronic indications fueled by uncontrolled C3 turnover. This review highlights recent developments in the field of complement therapeutics, focusing on C3-directed inhibitors and alternative pathway (AP) regulator-based approaches. Translational perspectives and considerations are discussed, particularly with regard to the structure-guided drug optimization and clinical advancement of a new generation of C3-targeted peptidic inhibitors.

Complement inhibition in pre-clinical models of periodontitis and prospects for clinical application.

Periodontitis is a dysbiotic inflammatory disease leading to the destruction of the tooth-supporting tissues. Current therapies are not always effective and this prevalent oral disease continues to be a significant health and economic burden. Early clinical studies have associated periodontitis with elevated complement activity. Consistently, subsequent genetic and pharmacological studies in rodents have implicated the central complement component C3 and downstream signaling pathways in periodontal host-microbe interactions that promote dysbiosis and inflammatory bone loss. This review discusses these mechanistic advances and moreover focuses on the compstatin family of C3 inhibitors as a novel approach to treat periodontitis. In this regard, local application of the current lead analog Cp40 was recently shown to block both inducible and naturally occurring periodontitis in non-human primates. These promising results from non-human primate studies and the parallel development of Cp40 for clinical use highlight the feasibility for developing an adjunctive, C3-targeted therapy for human periodontitis.

Modulation of immune response, physical barrier and related signaling factors in the gills of juvenile grass carp (Ctenopharyngodon idella) fed supplemented diet with phospholipids.

This study was conducted to investigate the effects of dietary phospholipids (PL) on the gill immune response and physical barrier of juvenile grass carp (Ctenopharyngodon idella). A total of 1080 juvenile grass carp with an average initial weight of 9.34 ± 0.03 g were fed six semi-purified diets containing 0.40% (unsupplemented control group), 1.43%, 2.38%, 3.29%, 4.37% and 5.42% PL for 2 months. Compared with the control group, optimal PL supplementation increased (P < 0.05): (1) the lysozyme activity, acid phosphatase activity, complement component 3 (C3) content, liver expressed antimicrobial peptide 1 (LEAP-1) and LEAP-2 mRNA expression; (2) the relative mRNA expression of interleukin 10, transforming growth factor ß1, inhibitor factor κBα (IκBα) and target of rapamycin (TOR); (3) the activities of anti-superoxide anion (ASA), anti-hydroxyl radical (AHR), copper/zinc superoxide dismutase (SOD1), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR), glutathione content and mRNA levels of SOD1, CAT, GPx, GR and NF-E2-related factor 2 (Nrf2) genes; (4) the transcription abundance of occludin, claudin b, claudin c, claudin 12 and zonula occludens 1 genes. At the same time, appropriate PL supplementation decreased (P < 0.05): (1) tumor necrosis factor α, interleukin 1ß, nuclear factor κB p65 (NF-κB p65), IκB kinase ß (IKKß) and IκB kinase γ (IKKγ) mRNA expression; (2) malondialdehyde (MDA), protein carbonyl (PC) and reactive oxygen species (ROS) content and the relative mRNA expression of Kelch-like-ECH-associated protein 1a (Keap1a) and Keap1b; (3) the transcription abundance of myosin light chain kinase (MLCK) and p38 mitogen-activated protein kinase (p38 MAPK) genes. In conclusion, the positive effect of PL on gill health is associated with the improvement of the immunity, antioxidant status and tight junction barrier of fish gills. Finally, based on ACP activity, C3 content, PC content and ASA activity in the gills, the optimal dietary PL level for juvenile grass carp (9.34-87.50 g) was estimated to be 3.62%, 4.30%, 3.91% and 3.86%, respectively.

Protective effects of Ephedra sinica extract on blood-brain barrier integrity and neurological function correlate with complement C3 reduction after subarachnoid hemorrhage in rats.

Early brain injury, which is associated with brain cell death, blood-brain barrier disruption, brain edema, and other pathophysiological events, is thought to be the main target in the prevention of poor outcomes after subarachnoid hemorrhage (SAH). Emerging evidences indicates that complement system, especially complement C3 is detrimental to neurological outcomes of SAH patients. Recently, Ephedra sinica extract was extracted and purified, which exhibits ability to block the activity of the classical and alternative pathways of complement, and improve neurological outcomes after spinal cord injury and ischemic brain injury. However, it is still unclear whether Ephedra sinica extract could attenuate early brain injury after SAH. In the present study, a standard endovascular perforation model was used to produce the experimental SAH in Sprague-Dawley rats. Ephedra sinica extract (15mg/kg) was orally administrated daily and evaluated for effects on modified Garcia score, brain water content, Evans blue extravasation and fluorescence, cortex cell death by TUNEL staining, and the expressions of complement C3/C3b, activated C3, sonic hedgehog, osteopontin and matrix metalloproteinase-9 by western bolt and immunofluorescence staining. We founded that the Ephedra sinica extract alleviated the blood-brain barrier disruption and brain edema, eventually improved neurological functions after SAH in rats. These neuroprotective effects was associated with the inhibition of complement C3, possibly via upregulating sonic hedgehog and osteopontin signal, and reducing the expressions of matrix metalloproteinase-9. Taking together, these observations suggested complement C3 inhibition by the Ephedra sinica extract may be a protective factor against early brain injury after SAH.

[Effect of Lappaconitine on Postoperative Pain and Serum Complement 3 and 4 Levels of Cancer Patients Undergoing Rectum Surgery].

OBJECTIVE: To explore the effect of lappaconitine on patient-controlled intravenous analgesia (PCIA) and serum complement 3 and 4 (C3 and C4) levels of cancer patients undergoing rectum surgery. METHODS: Totally 60 patients, who were scheduled for rectum carcinoma surgery, were recruited to the study and assigned in 3 groups, the blank control group, the tramadol group, and the lappaconitine group, 20 in each group. Lappaconitine (8 mg) was intravenously dripped to patients in the lappaconitine group 30 min before ending the operation. PCIA started as soon as the end of the surgery and the total dose of lappaconitine was 36 mg. Patients of the tramadol group were treated with tramadol (100 mg) intravenously within 30 min before ending the operation. The dripping was completed within 30 min. PCIA was started as soon as the end of the surgery and the total dose of lappaconitine was 36 mg. Tramadol (100 mg) was intravenously dripped to patients in the tramadol group 30 min before ending the operation. PICA was started as soon as the end of the surgery and the total dose of tramadol was 900 mg. Pethidine (50 mg) and droperidol (2. 5 mg) was intramuscularly injected to patients in the blank control group for pain relief according to their complaints. Pain degrees were assessed by visual analog scale (VAS) 12 h before surgery, 12, 24, 48, and 72 h after surgery. Blood samples were withdrawn at the same time point. Contents of serum C3 and C4 were determined by immunoturbidimetry. RESULTS: VAS scores of the blank control group were significantly higher after surgery than before surgery (P <0. 01). There was no statistical difference in VAS scores between before surgery and after surgery in the tramadol group and the lappaconitine group (P >0. 05). VAS scores were significantly lower at each post-surgery time point in the tramadol group and the lappaconitine group than in the blank control group with statistical difference (P < 0.01). There was no statistical difference in VAS scores at each post-surgery time point between the tramadol group and the lappaconitine group (P >0. 05). Compared with before surgery, contents of serum C3 and C4 significantly decreased in all of the three groups at 12, 24, and 48 h after surgery (P < 0.05, P < 0.01). They recovered to the pre-surgery level till 72 h after surgery (P > 0.05). Serum C3 and C4 contents at 48 h after surgery were higher in the tramadol group than in the blank control group (P < 0.05). Serum C3 and C4 contents at 24 and 48 h after surgery were higher in the lappaconitine group than in the blank control group (P < 0.05). There was no statistical difference in serum C3 and C4 contents at each time point between the tramadol group and the lappaconitine group (P > 0.05). VAS scores were obviously negatively correlated with serum contents of C3 and C4 (r = -0.622, r = -0.649, P < 0.01). CONCLUSIONS: Lappaconitine (used at the dose in this study) showed better pain relief effect after surgery. Besides, it could inhibit the surgic wound and pain, and elevate serum contents of C3 and C4.

Tacrolimus Protects Podocytes from Injury in Lupus Nephritis Partly by Stabilizing the Cytoskeleton and Inhibiting Podocyte Apoptosis.

OBJECTIVE: Several studies have reported that tacrolimus (TAC) significantly reduced proteinuria in lupus nephritis (LN) patients and mouse models. However, the mechanism for this effect remains undetermined. This study explored the mechanism of how TAC protects podocytes from injury to identify new targets for protecting renal function. METHODS: MRL/lpr mice were given TAC at a dosage of 0.1 mg/kg per day by intragastric administration for 8 weeks. Urine and blood samples were collected. Kidney sections (2 µm) were stained with hematoxylin-eosin (HE), periodic acid-Schiff base (PAS) and Masson's trichrome stain. Mouse podocyte cells (MPC5) were treated with TAC and/or TGF-ß1 for 48 h. The mRNA levels and protein expression of synaptopodin and Wilms' tumor 1 (WT1) were determined by real-time PCR, Western blotting and/or immunofluorescence, respectively. Flow cytometry was used to detect cell apoptosis with annexin V. Podocyte foot processes were observed under transmission electron microscopy. IgG and C3 deposition were assessed with immunofluorescence assays and confocal microscopy. RESULTS: Synaptopodin expression significantly decreased in MRL/lpr disease control mice, accompanied by increases in 24-h proteinuria, blood urea nitrogen, and serum creatinine. TAC, however, reduced proteinuria, improved renal function, attenuated renal pathology, restored synaptopodin expression and preserved podocyte numbers. In MPC5 cells, TGF-ß1 enhanced F-actin damage in podocytes and TAC stabilized it. TAC also decreased TGF-ß1-induced podocyte apoptosis in vitro and inhibited foot process fusion in MRL/lpr mice. In addition, our results also showed TAC inhibited glomerular deposition of IgG and C3. CONCLUSION: This study demonstrated that TAC reduced proteinuria and preserved renal function in LN through protecting podocytes from injury partly by stabilizing podocyte actin cytoskeleton and inhibiting podocyte apoptosis.