Truncated Complement Factor H Y402 Gene Therapy Cures C3 Glomerulonephritis.

Patients with both age-related macular degeneration (AMD) and C3 glomerulonephritis (C3G) are challenged by the absence of effective therapies to reverse and eliminate their disease burden. Capitalizing on complement dysregulation as both a significant risk factor for AMD and the known pathophysiology of C3G, we investigated the potential for adeno-associated virus (AAV) delivery of complement factor H (CFH) to rescue C3G in a Cfh-/- mouse model of C3G. While past efforts to treat C3G using exogenous human CFH resulted in limited success before immune rejection led to a foreign protein response, our findings demonstrate the capacity for long-term AAV-mediated delivery of truncated CFH (tCFH) to restore inhibition of the alternative pathway of complement and ultimately reverse C3G without immune rejection. Comparing results from the administration of several tCFH vectors also revealed significant differences in their relative efficiency and efficacy. These discoveries pave the way for subsequent development of AAV-mediated tCFH replacement therapy for patients with C3G, while simultaneously demonstrating proof of concept for a parallel AAV-mediated tCFH gene augmentation therapy for patients with AMD.

Complement Factor B (CFB) inhibits the malignant progression of lung adenocarcinoma by downregulating the Ras/MAPK signaling pathway.

Lung adenocarcinoma (LUAC) as the most common lung cancer, and its incidence is increasing. Complement factor B (CFB) is an important factor in the alternative complement pathway. CFB has been reported to be involved in the progression of many cancers, including in pancreatic cancer, cutaneous squamous cell carcinoma, and nasopharyngeal carcinoma, but the function and molecular mechanism of CFB in LUAC remains unclear. The present study aimed to explore the role of CFB in LUAC malignant progression. In our previous study, we found that CFB was downregulated expression in LUAC clinical samples. Here, we firstly detected the cell function in vitro. Cell proliferation and migration were increased, while cell apoptosis and cell cycle arrest were suppressed after CFB knockdown. Overexpression of CFB repressed the malignant progression of LUAC in vitro. Besides, in vivo experiments revealed that upregulation of CFB inhibited tumor growth and Ki67 expression. Additionally, our data indicated that CFB negatively regulated Ras/mitogen-activated protein kinase (MAPK) signaling pathway. Furthermore, upregulation of CFB inhibited the progression of LUAC was reversed by Ras/MAPK pathway activators (ML-098 or C16-PAF). Our study uncovered that CFB acts as a tumor suppressor repressed tumorigenesis of LUAC through inhibiting the Ras/MAPK pathway, suggesting that CFB may be a potential biomarker and therapeutic target for LUAC.

Complement Component C5a and Fungal Pathogen Induce Diverse Responses through Crosstalk between Transient Receptor Potential Channel (TRPs) Subtypes in Human Conjunctival Epithelial Cells.

The conjunctiva has immune-responsive properties to protect the eye from infections. Its innate immune system reacts against external pathogens, such as fungi. The complement factor C5a is an important contributor to the initial immune response. It is known that activation of transient-receptor-potential-vanilloid 1 (TRPV1) and TRP-melastatin 8 (TRPM8) channels is involved in different immune reactions and inflammation in the human body. The aim of this study was to determine if C5a and mucor racemosus e voluminae cellulae (MR) modulate Ca2+-signaling through changes in TRPs activity in human conjunctival epithelial cells (HCjECs). Furthermore, crosstalk was examined between C5a and MR in mediating calcium regulation. Intracellular Ca2+-concentration ([Ca2+]i) was measured by fluorescence calcium imaging, and whole-cell currents were recorded using the planar-patch-clamp technique. MR was used as a purified extract. Application of C5a (0.05-50 ng/mL) increased both [Ca2+]i and whole-cell currents, which were suppressed by either the TRPV1-blocker AMG 9810 or the TRPM8-blocker AMTB (both 20 µM). The N-terminal peptide C5L2p (20-50 ng/mL) blocked rises in [Ca2+]i induced by C5a. Moreover, the MR-induced rise in Ca2+-influx was suppressed by AMG 9810 and AMTB, as well as 0.05 ng/mL C5a. In conclusion, crosstalk between C5a and MR controls human conjunctival cell function through modulating interactions between TRPV1 and TRPM8 channel activity.

Enhancement of complement-dependent cytotoxicity by linking factor-H derived short consensus repeats 19-20 to CD20 antibodies.

Antibody-mediated complement-dependent cytotoxicity (CDC) on malignant cells is regulated by several complement control proteins, including the inhibitory complement factor H (fH). fH consists of 20 short consensus repeat elements (SCRs) with specific functional domains. Previous research revealed that the fH-derived SCRs 19-20 (SCR1920) can displace full-length fH on the surface of chronic lymphocytic leukemia (CLL) cells, which sensitizes CLL cells for e.g. CD20-targeting therapeutic monoclonal antibody (mAb) induced CDC. Therefore, we constructed lentiviral vectors for the generation of cell lines that stably produce mAb-SCR-fusion variants starting from the clinically approved parental mAbs rituximab, obinutuzumab and ofatumumab, respectively. Flow-cytometry revealed that the modification of the mAbs by the SCRs does not impair the binding to CD20. Increased in vitro lysis potency compared to their parental mAbs was corroborated by showing specific and dose dependent target cell elimination by CDC when compared to their parental mAbs. Lysis of CLL cells was not affected by the depletion of NK cells, suggesting that antibody-dependent cellular cytotoxicity plays a minor role in this context. Overall, this study emphasizes the crucial role of CDC in the elimination of CLL cells by mAbs and introduces a novel approach for enhancing CDC by directly fusing fH SCR1920 with mAbs.

Complement factor I knockdown inhibits colon cancer development by affecting Wnt/ß-catenin/c-Myc signaling pathway and glycolysis.

BACKGROUND: Colon cancer (CC) occurrence and progression are considerably influenced by the tumor microenvironment. However, the exact underlying regulatory mechanisms remain unclear. AIM: To investigate immune infiltration-related differentially expressed genes (DEGs) in CC and specifically explored the role and potential molecular mechanisms of complement factor I (CFI). METHODS: Immune infiltration-associated DEGs were screened for CC using bioinformatics. Quantitative reverse transcription polymerase chain reaction was used to examine hub DEGs expression in the CC cell lines. Stable CFI-knockdown HT29 and HCT116 cell lines were constructed, and the diverse roles of CFI in vitro were assessed using CCK-8, 5-ethynyl-2'-deoxyuridine, wound healing, and transwell assays. Hematoxylin and eosin staining and immunohistochemistry staining were employed to evaluate the influence of CFI on the tumorigenesis of CC xenograft models constructed using BALB/c male nude mice. Key proteins associated with glycolysis and the Wnt pathway were measured using western blotting. RESULTS: Six key immune infiltration-related DEGs were screened, among which the expression of CFI, complement factor B, lymphoid enhancer binding factor 1, and SRY-related high-mobility-group box 4 was upregulated, whereas that of fatty acid-binding protein 1, and bone morphogenic protein-2 was downregulated. Furthermore, CFI could be used as a diagnostic biomarker for CC. Functionally, CFI silencing inhibited CC cell proliferation, migration, invasion, and tumor growth. Mechanistically, CFI knockdown downregulated the expression of key glycolysis-related proteins (glucose transporter type 1, hexokinase 2, lactate dehydrogenase A, and pyruvate kinase M2) and the Wnt pathway-related proteins (ß-catenin and c-Myc). Further investigation indicated that CFI knockdown inhibited glycolysis in CC by blocking the Wnt/ß-catenin/c-Myc pathway. CONCLUSION: The findings of the present study demonstrate that CFI plays a crucial role in CC development by influencing glycolysis and the Wnt/ß-catenin/c-Myc pathway, indicating that it could serve as a promising target for therapeutic intervention in CC.

Urinary complement factor D is increased in primary malignant hypertension: a single-center, cross-sectional study.

Kidney injury is one of the detrimental consequences of primary malignant hypertension (pMHTN). There is a paucity of non-invasive biomarkers to enhance diagnosis and elucidate the underlying mechanisms. This study aims to explore urine protein biomarkers for pMHTN associated renal damage. In the discovery phase, urine samples were collected from 8 pMHTN, 19 disease controls (DCs), and 5 healthy controls (HCs). In-gel digestion combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach was used for identification of proteins associated with pMHTN. In the validation phase, the differentially expressed proteins were validated by ELISA assay in cohort with 10 pMHTN patients, 37 DCs, and 30 HCs. Compared to DCs and HCs, a specific band between 15 and 25 kDa was found in 7 out of 8 patients with pMHTN. Further LC-MS/MS analysis revealed 5 differentially expressed proteins. ELISA validation demonstrated that urinary complement factor D (CFD) was significantly up regulated in pMHTN. By receiver operating characteristic curve analysis, urinary CFD/Cr showed moderate potential in discriminating pMHTN from DCs (the area under curve: 0.822, 95% CI 0.618-0.962). Urinary CFD may be a potential biomarker for pMHTN with its elevation indicative of the activation of the alternative complement pathway in pMHTN.

Anti-C5 monoclonal antibody treatment showing pathological resolution of complement-mediated atypical hemolytic uremic syndrome: a case report.

BACKGROUND: No reports have shown histological changes before and after anti-C5 monoclonal antibody treatment in patients with atypical hemolytic uremic syndrome (aHUS). Here, we report a rare case of complement-mediated aHUS with a complement factor H (CFH) mutation and anti-CFH antibodies who underwent multiple kidney biopsies. CASE PRESENTATION: A 53-year-old woman developed aHUS with CFH gene mutation [c.3572C > T (p. Ser1191 Leu)] and anti-CFH antibodies. Her father had succumbed to acute kidney injury (AKI) in his 30 s. She exhibited AKI, thrombocytopenia, and hemolytic anemia with schistocytes. After improving the platelet count with one session of plasma exchange, a kidney biopsy was performed one month after the onset of symptoms. Blood vessel thrombosis, obvious endothelial swelling, endocapillary hypercellularity, and subendothelial exudative lesions in the glomeruli and arterioles were detected. Anti-C5 monoclonal antibody treatment with eculizumab immediately improved disease activity. A second biopsy 3 months later revealed marked improvement of endothelial injuries with residual membrane double contours and exudative lesions. A third biopsy at 17 months after gradual improvement of kidney function showed a further decrease of double contours along with alterations of the exudative lesions to fibrous intimal thickening. CONCLUSIONS: This is the first report showing the pathophysiology of aHUS in the kidneys and the efficacy of anti-C5 monoclonal antibody treatment by presenting serial kidney pathological features before and after anti-C5 monoclonal antibody treatment. Since her CFH mutation was considered the most important pathological condition, treatment centered on eculizumab was administered, resulting in a good long-term prognosis. In addition, kidney pathological resolution in aHUS occurred over 1 year after anti-C5 monoclonal antibody treatment.

Bilateral Retinal Venous Occlusion in Atypical Hemolytic-Uremic Syndrome Due to Complement Factor H Mutation.

PURPOSE: Atypical hemolytic uremic syndrome (aHUS) is a rare progressive thrombotic microangiopathy caused by overactivation in the alternative complement pathway. A wide spectrum of environmental triggers, such as viruses, vaccination, drugs, pregnancy, neoplasms, transplant, and autoimmune diseases can cause aHUS in genetically susceptible individuals. In this report, the diagnosis and treatment process of aHUS and bilateral retinal venous occlusion (RVO) will be presented. METHODS: Single-case, retrospective management of ophthalmological and systemic manifestations. RESULTS: A 28-year-old G2P2 female with acute blurred vision and history of acute renal failure. She was diagnosed with preeclampsia in her gestation history. After the laboratory work-up, the diagnosis of aHUS was confirmed. She was treated with eculizumab following 14 days of plasmapheresis. However, her visual acuity was 20/20 on the right and 20/60 on the left at the time of admission. Retinal examination revealed flame-shaped hemorrhages, exudation, and macular edema. The patient was diagnosed with branch RVO in the right eye. Subsequently, central RVO was occurred in the left eye. Intravitreal dexamethasone implant was administered for both eyes since there was no reasonable regression in retinal findings with bevacizumab treatment. She went into remission and her BCVA reached 20/25 during the 12-month follow-up period under the eculizumab therapy. CONCLUSION: Diagnosis of aHUS is challenging especially during pregnancy and the postpartum period. Although ocular involvement is quite rare, we described bilateral RVO in aHUS case with homozygous nonsense mutation (c.2134 G > T p.G712). Dexamethasone implant should be considered for the treatment of RVO in aHUS cases.

Hot Spot of Complement Factor I Rare Variant p.Ile357Met in Patients With Hemolytic Uremic Syndrome.

Atypical hemolytic uremic syndrome (aHUS) is a rare kidney disease due to a dysregulation of the complement alternative pathway. Complement factor I (CFI) negatively regulates the alternative pathway and CFI gene rare variants have been associated to aHUS with a low disease penetrance. We report 10 unrelated cases of HUS associated to a rare CFI variant, p.Ile357Met (c.1071T>G). All patients with isolated p.Ile357Met CFI missense variant were retrospectively identified among patients included between January 2007 and January 2022 in the French HUS Registry. We identified 10 unrelated patients (70% women; median age at HUS diagnosis, 36.5 years) who carry the same rare variant p.Ile357Met in the CFI gene. Seven patients (cases 1-7) presented with aHUS in the native kidney associated with malignant hypertension in 5 patients. None received a C5 inhibitor. Two of these cases occurred in the peripartum period with complete recovery of kidney function, while 5 of these patients reached kidney failure requiring replacement therapy (KFRT). Four patients with KFRT subsequently underwent kidney transplantation. Three later developed C3 glomerulopathy in their kidney graft, but none had aHUS recurrence. Three other patients (cases 8-10) experienced de novo thrombotic microangiopathy after kidney transplantation, precipitated by various triggers. The rare CFI variant p.Ile357Met appears to be a facilitating genetic factor for HUS and for some forms of secondary HUS.

Inhibition of the Alternative Complement Pathway May Cause Secretion of Factor B, Enabling an Early Detection of Pancreatic Cancer.

This study aims to elucidate the cellular mechanisms behind the secretion of complement factor B (CFB), known for its dual roles as an early biomarker for pancreatic ductal adenocarcinoma (PDAC) and as the initial substrate for the alternative complement pathway (ACP). Using parallel reaction monitoring analysis, we confirmed a consistent ∼2-fold increase in CFB expression in PDAC patients compared with that in both healthy donors (HD) and chronic pancreatitis (CP) patients. Elevated ACP activity was observed in CP and other benign conditions compared with that in HD and PDAC patients, suggesting a functional link between ACP and PDAC. Protein-protein interaction analyses involving key complement proteins and their regulatory factors were conducted using blood samples from PDAC patients and cultured cell lines. Our findings revealed a complex control system governing the ACP and its regulatory factors, including Kirsten rat sarcoma viral oncogene homolog (KRAS) mutation, adrenomedullin (AM), and complement factor H (CFH). Particularly, AM emerged as a crucial player in CFB secretion, activating CFH and promoting its predominant binding to C3b over CFB. Mechanistically, our data suggest that the KRAS mutation stimulates AM expression, enhancing CFH activity in the fluid phase through binding. This heightened AM-CFH interaction conferred greater affinity for C3b over CFB, potentially suppressing the ACP cascade. This sequence of events likely culminated in the preferential release of ductal CFB into plasma during the early stages of PDAC. (Data set ID PXD047043.).

Complement factor H: a novel innate immune checkpoint in cancer immunotherapy.

The elimination of cancer cells critically depends on the immune system. However, cancers have evolved a variety of defense mechanisms to evade immune monitoring, leading to tumor progression. Complement factor H (CFH), predominately known for its function in inhibiting the alternative pathway of the complement system, has recently been identified as an important innate immunological checkpoint in cancer. CFH-mediated immunosuppression enhances tumor cells' ability to avoid immune recognition and produce an immunosuppressive tumor microenvironment. This review explores the molecular underpinnings, interactions with immune cells, clinical consequences, and therapeutic possibilities of CFH as an innate immune checkpoint in cancer control. The difficulties and opportunities of using CFH as a target in cancer immunotherapy are also explored.

Gliosis-dependent expression of complement factor H truncated variants attenuates retinal neurodegeneration following ischemic injury.

Inherited, age-related, and acute retinal diseases are often exacerbated by an aberrant or excessive activity of the complement system. Consequently, cells not directly affected by an acute event or genetic variants may degenerate, resulting in enhanced visual impairment. The therapeutic potential of supplementation of complement factor H (FH), a key regulator of the complement cascade, is therefore particularly promising in the context of retinal diseases caused by complement activation. In this study, we engineered adeno-associated viruses (AAVs) containing sequences of two truncated human FH variants. The expression of these variants was regulated by the glial fibrillary acidic protein (GFAP) promoter, which is selectively active in gliotic Müller cells. Both FH variants consisted of FH domains 19-20, which were connected to domains 1-4 and 1-7, respectively, by a polyglycine linker. These AAVs were intravitreally injected following ischemic injury of C57BL/6J mouse retinas. We observed transgene expression in gliotic Müller cells and to some extent in astrocytes. The expression correlated directly with damage severity. Interventions resulted in decreased complement activation, accelerated normalization of microglia activity and morphological improvements. Reduced levels of C3 transcripts and C3d protein in conjunction with higher transcript levels of inhibitory regulators like Cfi and Cfh, hinted at attenuated complement activity. This study demonstrates the great potential of complement regulatory gene addition therapy. With further in vivo testing it could be applied to treat a wide range of retinal diseases where no causative therapies are available.

The Role and Potential Mechanism of Complement Factor D in Fibromyalgia Development.

Purpose: Fibromyalgia (FM) is a syndrome characterized by chronic musculoskeletal pain. Its clinical symptoms include both somatic and psychiatric symptoms, making the treatment of FM extremely challenging. The cause of FM is still unknown, and some patients do not improve their symptoms even after long-term active treatment. Thus, the development of new targeted therapies is important to reduce pain and improve quality of life for FM patients. Methods: In this study, we screened genes and secreted factors that play key roles in FM through bioinformatics and big data analysis. Furthermore, we performed CCK-8, qRT-PCR, glucose, ATP and lactate content testing, dual luciferase reporter gene assay to investigate the potential mechanism of complement factor D in fibromyalgia development. Results: In bioinformatics and big data analysis, we identified CFD was negatively correlated with the pro-inflammatory factor IL-6 and positively correlated with the anti-inflammatory factor IL-4, which suggested that CFD may be an anti-inflammatory factor. Through cellular assays, we verified that CFD reversed the decrease in IL-4 expression and the increase in IL-6 expression in BV2 cells caused by ATP. Conclusion: In summary, based on bioinformatic methods and big data mining we obtained a new target CFD for FM, and further experiments verified that CFD has significant inhibition of ATP-induced neuropathic pain. These findings provide a new theoretical basis for the clinical translation of CFD.

Complement factor H attenuates TNF-α-induced inflammation by upregulating EIF3C in rheumatoid arthritis.

OBJECTIVE: To explore the role and underlying mechanism of Complement Factor H (CFH) in the peripheral and joint inflammation of RA patients. METHODS: The levels of CFH in the serum and synovial fluid were determined by ELISA. The pyroptosis of monocytes was determined by western blotting and flow cytometry. The inflammation cytokine release was tested by ELISA. The cell migration and invasion ability of fibroblast-like synoviocytes (FLS) were tested by Wound healing Assay and transwell assay, respectively. The potential target of CFH was identified by RNA sequencing. RESULTS: CFH levels were significantly elevated in the serum and synovial fluid from RA and associated with high sensitivity C-reactive protein (hs-CRP), erythrocyte sedimentation rate (ESR), and disease activity score 28 (DAS28). TNF-α could inhibit CFH expression, and CFH combined with TNF-α significantly decreased cell death, cleaved-caspase 3, gasdermin E N-terminal (GSDME-N), and inflammatory cytokines release (IL-1ß and IL-6) of RA-derived monocytes. Stimulated with TNF-α increased CFH levels in RA FLS and CFH inhibits the migration, invasion, and TNF-α-induced production of inflammatory mediators, including proinflammatory cytokines (IL-6, IL-8) as well as matrix metalloproteinases (MMPs, MMP1 and MMP3) of RA FLSs. The RNA-seq results showed that CFH treatment induced upregulation of eukaryotic translation initiation factor 3 (EIF3C) in both RA monocytes and FLS. The migration of RA FLSs was promoted and the expressions of IL-6, IL-8, and MMP-3 were enhanced upon EIF3C knockdown under the stimulation of CFH combined with TNF-α. CONCLUSION: In conclusion, we have unfolded the anti-inflammatory roles of CFH in the peripheral and joints of RA, which might provide a potential therapeutic target for RA patients.

New Immunohistochemical Markers for Pleural Mesothelioma Subtyping.

Pleural mesothelioma (PM) comprises three main subtypes: epithelioid, biphasic and sarcomatoid, which have different impacts on prognosis and treatment definition. However, PM subtyping can be complex given the inter- and intra-tumour morphological heterogeneity. We aim to use immunohistochemistry (IHC) to evaluate five markers (Mesothelin, Claudin-15, Complement Factor B, Plasminogen Activator Inhibitor 1 and p21-activated Kinase 4), whose encoding genes have been previously reported as deregulated among PM subtypes. Immunohistochemical expressions were determined in a case series of 73 PMs, and cut-offs for the epithelioid and non-epithelioid subtypes were selected. Further validation was performed on an independent cohort (30 PMs). For biphasic PM, the percentage of the epithelioid component was assessed, and IHC evaluation was also performed on the individual components separately. Mesothelin and Claudin-15 showed good sensitivity (79% and 84%) and specificity (84% and 73%) for the epithelioid subtype. CFB and PAK4 had inferior performance, with higher sensitivity (89% and 84%) but lower specificity (64% and 36%). In the biphasic group, all markers showed different expression when comparing epithelioid with sarcomatoid areas. Mesothelin, Claudin-15 and CFB can be useful in subtype discrimination. PAI1 and PAK4 can improve component distinction in biphasic PM.

Complement Factor I Gene Variant in an Atypical Hemolytic Uremic Syndrome Triggered by Hypereosinophilia Syndrome.

Atypical hemolytic uremic syndrome (aHUS) is a condition characterized by acute kidney injury (AKI), thrombocytopenia, and microangiopathic hemolytic anemia secondary to complement pathway dysregulation. Several triggers have been identified as causing aHUS in genetically susceptible patients; however, hypereosinophilia syndrome (HES)-triggered aHUS has not been reported. In this article, we present a case of aHUS presented with generalized urticarial rashes and angioedema. The initial investigations revealed hypereosinophilia (maximal absolute eosinophil count of 6,840 cells/µL) with normal bone-marrow analyses; hence, idiopathic HES was diagnosed. During hospitalization, the patient developed convulsion, stuporous, and full-blown thrombotic microangiopathy (TMA), with AKI requiring temporary hemodialysis. A kidney biopsy confirmed the existence of renal TMA. Next-generation sequencing of the coding regions of aHUS-related genes was performed, revealing an underlying complement factor I (CFI) deficiency, a heterozygous variant p.P64L of CFI gene. The patient was successfully treated with high-dose steroids and extended duration of plasmapheresis.

Characterization of the pleural microenvironment niche and cancer transition using single-cell RNA sequencing in EGFR-mutated lung cancer.

Background: Lung cancer is associated with a high mortality rate and often complicated with malignant pleural effusion (MPE), which has a very poor clinical outcome with a short life expectancy. However, our understanding of cell-specific mechanisms underlying the pathobiology of pleural metastasis remains incomplete. Methods: We analyzed single-cell transcriptomes of cells in pleural effusion collected from patients with lung cancer and congestive heart failure (as a control), respectively. Soluble and complement factors were measured using a multiplex cytokine bead assay. The role of ferroptosis was evaluated by GPX4 small interfering RNA (siRNA) transfection and overexpression. Results: We found that the mesothelial-mesenchymal transition (MesoMT) of the pleural mesothelial cells contributed to pleural metastasis, which was validated by lung cancer/mesothelial cell co-culture experiments. The ferroptosis resistance that protected cancer from death which was secondary to extracellular matrix detachment was critical for pleural metastasis. We found a universal presence of immune-suppressive lipid-associated tumor-associated macrophages (LA-TAMs) with complement cascade alteration in the MPE of the lung cancer patients. Specifically, upregulated complement factors were also found in the MPE, and C5 was associated with poor overall survival in the lung cancer patients with epidermal growth factor receptor mutation. Plasmacytoid dendritic cells (pDCs) exhibited a dysfunctional phenotype and pro-tumorigenic feature in the primary cancer. High expression of the gene set extracted from pDCs was associated with a poor prognosis in the lung cancer patients. Receptor-ligand interaction analysis revealed that the pleural metastatic niche was aggravated by cross-talk between mesothelial cells-cancer cells/immune cells via TNC and ICAM1. Conclusions: Taken together, our results highlight cell-specific mechanisms involved in the pathobiological development of pleural metastasis in lung cancer. These results provide a large-scale and high-dimensional characterization of the pleural microenvironment and offer a useful resource for the future development of therapeutic drugs in lung cancer.

Complement factor H inhibits endothelial cell migration through suppression of STAT3 signaling.

Complement factor H (CFH), a major soluble inhibitor of complement, is a plasma protein that directly interacts with the endothelium of blood vessels. Mutations in the CFH gene lead to diseases associated with excessive angiogenesis; however, the underlying mechanisms are unknown. The present study aimed to determine the effects of CFH on endothelial cells and to explore the underlying mechanisms. The adenoviral plasmid expressing CFH was transduced into HepG2 cells, and the culture medium supernatant was collected and co-cultured with human umbilical vein endothelial cells (HUVECs). Cell proliferation was measured by CCK8 and MTT assays, and cell migration was measured by wound healing and Transwell assays. Reverse transcription-quantitative PCR was performed to detect gene transcription. Western blotting was used to determine protein levels. The results revealed that CFH can inhibit migration, but not viability, of HUVECs. In addition, CFH did not significantly alter MAPK or TGF-ß signaling, whereas it decreased STAT3 phosphorylation in HUVECs. Furthermore, CFH failed to reduce migration of HUVECs, with inhibition of STAT3 signaling by STATTIC or activation of STAT3 signaling by overexpression of STAT3 (Y705D) compromising CFH-inhibited HUVEC migration. CFH also decreased the expression levels of vascular endothelial growth factor receptor 2, a downstream effector of STAT3 mediating endothelial cell migration. In conclusion, the present study suggested that CFH may be a potential therapeutic target for angiogenesis-related diseases.

Low levels of complement factor C3 at diagnosis can predict outcome in antineutrophil antibody associated vasculitis.

INTRODUCTION: Experimental data support the involvement of complement in the pathogenesis of antineutrophil antibody associated vasculitis, and clinical studies describe a more severe disease phenotype in patients with antineutrophil antibody associated vasculitis and complement activation. In the present study, we looked for an association between circulating serum complement factor 3 levels at diagnosis and outcomes. METHODS: One hundred sixty-four patients with antineutrophil antibody associated vasculitis who underwent kidney biopsy at our center during the last 15 years were retrospectively reviewed. Patients were categorized according to their serum complement factor 3 level at diagnosis. Patient and renal survival were compared between those above and below the median serum complement factor 3 at diagnosis. RESULTS: During the first year, 6 patients died and 53 reached end-stage renal disease. Death or end-stage renal disease at one-year were significantly more common in the low serum complement factor 3 group (44 vs. 29%, p = 0.037). In the multivariable analysis, serum complement factor 3 was the strongest negative outcome predictor (HR, 95%CI 0.118, (0.021-0.670)). The lower the serum complement factor 3 level at baseline, the higher the risk of dialysis and death. The risk was particularly high for both endpoints if the serum complement factor 3 concentration was below 0.9 g/l at baseline. CONCLUSION: Complement activation at diagnosis may identify a distinct subgroup of patients with antineutrophil antibody associated vasculitis and higher risk for poor outcomes. However, it remains to be proven whether inhibition of serum complement factor 3 is beneficial and safe in the clinical setting.

Cell-autonomous regulation of complement C3 by factor H limits macrophage efferocytosis and exacerbates atherosclerosis.

Complement factor H (CFH) negatively regulates consumption of complement component 3 (C3), thereby restricting complement activation. Genetic variants in CFH predispose to chronic inflammatory disease. Here, we examined the impact of CFH on atherosclerosis development. In a mouse model of atherosclerosis, CFH deficiency limited plaque necrosis in a C3-dependent manner. Deletion of CFH in monocyte-derived inflammatory macrophages propagated uncontrolled cell-autonomous C3 consumption without downstream C5 activation and heightened efferocytotic capacity. Among leukocytes, Cfh expression was restricted to monocytes and macrophages, increased during inflammation, and coincided with the accumulation of intracellular C3. Macrophage-derived CFH was sufficient to dampen resolution of inflammation, and hematopoietic deletion of CFH in atherosclerosis-prone mice promoted lesional efferocytosis and reduced plaque size. Furthermore, we identified monocyte-derived inflammatory macrophages expressing C3 and CFH in human atherosclerotic plaques. Our findings reveal a regulatory axis wherein CFH controls intracellular C3 levels of macrophages in a cell-autonomous manner, evidencing the importance of on-site complement regulation in the pathogenesis of inflammatory diseases.