Complement protein C1q stimulates hyaluronic acid degradation via gC1qR/HABP1/p32 in malignant pleural mesothelioma

Complement component C1q can act as a pro-tumorigenic factor in the tumor microenvironment (TME). The TME in malignant pleural mesothelioma (MPM) is rich in C1q and hyaluronic acid (HA), whose interaction enhances adhesion, migration and proliferation of malignant cells. HA-bound C1q is also capable of modulating HA synthesis. Thus, we investigated whether HA-C1q interaction would affect HA degradation, analyzing the main degradation enzymes, hyaluronidase (HYAL)1 and HYAL2, and a C1q receptor candidate. We first proceeded with the characterization of HYALs in MPM cells, especially HYAL2, since bioinformatics survival analysis revealed that higher HYAL2 mRNA levels have an unfavorable prognostic index in MPM patients. Interestingly, Real-Time quantitative PCR, flow cytometry and Western blot highlighted an upregulation of HYAL2 after seeding of primary MPM cells onto HA-bound C1q. In an attempt to unveil the receptors potentially involved in HA-C1q signaling, a striking co-localization between HYAL2 and globular C1q receptor/HABP1/p32 (gC1qR) was found by immunofluorescence, surface biotinylation and proximity ligation assays. RNA interference experiments revealed a potentially regulatory function exerted by gC1qR on HYAL2 expression, since C1QBP (gene for gC1qR) silencing unexpectedly caused HYAL2 downregulation. In addition, the functional blockage of gC1qR by a specific antibody hindered HA-C1q signaling and prevented HYAL2 upregulation. Thus, C1q-HA interplay is responsible for enhanced HYAL2 expression, suggesting an increased rate of HA catabolism and the release of pro-inflammatory and pro-tumorigenic HA fragments in the MPM TME. Our data support the notion of an overall tumor-promoting property of C1q. Moreover, the overlapping localization and physical interaction between HYAL2 and gC1qR suggests a potential regulatory effect of gC1qR within a putative HA-C1q macromolecular complex

Complement component C1q and hyaluronic acid act as novel "signaling complex" in malignant pleural mesothelioma microenvironment to promote cancer progression

Il mesotelioma pleurico maligno (MPM) \ue8 una forma rara di cancro che si sviluppa dalle cellule del mesotelio pleurico ed \ue8 pi\uf9 comunemente associato all'esposizione all'amianto. La crescita del tumore non dipende solo da anomalie genetiche che si accumulano nelle cellule tumorali, ma anche dal microambiente tumorale la cui composizione puo' promuovere la loro sopravvivenza, crescita e migrazione. Uno dei componenti pi\uf9 abbondanti della matrice extracellulare del mesotelioma pleurico maligno \ue8 rappresentato dall'acido ialuronico (HA). Inoltre, il fattore C1q, il primo componente della via clasica di attivazione del complemento, \ue8 risultato essere altamente espresso nel microambiente tumorale di tutte le varianti istologiche di mesotelioma. Con questo studio abbiamo dimostrato che il fattore complementare C1q pu\uf2 legare l' HA con un'affinit\ue0 simile a quella delle Immunoglobuline. Questa osservazione ha sollevato la possibilit\ue0 che il C1q legato ad HA possa funzionare come un nuovo "complesso di segnalazione" in grado di sostenere lo sviluppo e la progressione del tumore. In effetti, l'HA legato al C1q \ue8 risultato capace di migliorare l'adesione, la migrazione e la proliferazione delle cellule di mesotelioma, attraverso l'attivazione di diverse vie di segnalazione correlate al cancro, come MAP3K e l'mTOR. Sempre il complesso HA-C1q \ue8 risutato capace di alterare il metabolismo dell'HA promuovendo l'espressione di ialuronidasi 2 (HYAL2), un enzima coinvolto nella degradazione dell'HA, che potrebbe condurre alla produzione locale di HA a basso peso molecolare (LMW-HA). Queste specie di HA sono note per svolgere un ruolo cruciale nella tumorigenesi influenzando la proliferazione e la motilit\ue0 cellulare, esibendo effetti pro-angiogenici e promuovendo la produzione di specie reattive dell'ossigeno (ROS). Sulla base di questi risultati possiamo ipotezzare che C1q-HA, aumentando la degradazione di HA attraverso HYAL2, alimenter\ue0 la deposizione di LMW-HA nel microambiente tumorale che, a sua volta, favorir\ue0 la produzione di ROS e l'ulteriore degradazione di HA. In ultima analisi \ue8 possibile ipotizzare che tutti questi eventi agiscano di concerto per alterare le propriet\ue0 di segnalazione del microambiente tumorale, rendendolo ancora pi\uf9 permissivo allo sviluppo e alla progressione del cancro.Malignant pleural mesothelioma (MPM) is a rare form of cancer that develops from cells of the pleural mesothelium and is most commonly associated with exposure to asbestos. Tumor growth is not only dependent on genetic abnormalities accumulating in cancer cells, but also on the local microenvironment which can provide a permissive niche for their survival, growth and migration. One of the most abundanly expressed component of the MPM extracellular matrix is represented by the hyaluronic acid (HA). Moreover C1q, the first component of the classical complement pathway, has emerged to be also highly expressed in the tumour microenvironment of all MPM hystotypes variants. Interstingly, we demonstrated that C1q can bind HA with an affinity similar to IgG. This observation raised the possibility that C1q bound to HA would function as a novel \u201csignaling complex\u201d able to sustain MPM development and progression. Indeed C1q-bound HA emerged to enhance mesothelioma cells adhesion, migration and proliferation, through the activation of several cancer-related signaling pathways, such as MAP3Ks and the mTOR. Morevoer C1q-bound HA can affect HA homeostasis by enhancing the expression of hyaluronidase 2 (HYAL2), an enzyme involved in HA degradation, possibly leading to local production of Low Molecular Weight-HA (LMW-HA). These HA species are known to play a crucial role in tumorigenesis by affecting cell proliferation and motility, by exhibiting pro-angiogenic effects and by promoting reactive oxygen species (ROS) production. Based on these findings we can evisage that C1q-HA, by enhancing HA degradation via HYAL2, will feed LMW-HA deposition in the tumor microenvironment which, in turn, will favour ROS production and further HA degradation. These events are expected to act in concert to alter the signaling properties of the tumor microenvironment, render it even more permissive to cancer development and progression

COMPLEMENT PROTEIN C1Q PRODUCTION IN MALIGNANT PLEURAL MESOTHELIOMA

PURPOSE The complement component C1q has been shown to be abundantly expressed in the microenvironment of several solid tumors where it shows pro-tumor activities (1). We demonstrated that C1q is abundantly present in malignant pleural mesothelioma (MPM), and promote adhesion, migration and invasion of MPM tumor cells. The aim of our study was to investigate the cells and the mechanisms responsible for its local production. METHODS MPM sections were analyzed by immunohistochemistry for the presence of C1q in the microenvironment. MPM human primary mesothelioma cells were isolated from pleural biopsy, characterized by immunofluorescence, cytofluorimetric analysis and their production of cytokines was evaluated by qPCR and ELISA. Human macrophages were incubated with MPM conditioned medium and their phenotype and their production of C1q, was evaluated by qPCR and ELISA. RESULTS C1q was express in tumor-associated stroma of different histotypes of mesothelioma. C1q pattern distribution seems connected to tumor-infiltrating myeloid elements. MPM cells were unable to produce C1q. M\u3a6 treated with MPM conditioned medium have shown an M2-like phenotypic profile (CD206 and IL-10 upregulation) and a significant upregulation in C1q production. No variation was detected for C1s gene. DISCUSSION C1q has been shown able to induce M2-like polarization of M\u3c6 (2). MPM cells increase the C1q production by M\u3c6. Higher C1q presence in the microenvironment could lead to a stronger M2-like polarization of M\u3c6 producing a self-sustained cycle that could promote tumor malignant progression.CONCLUSIONS C1q fulfill a key role as an immunosuppressive and cancer-promoting factor in mesothelioma microenvironment. BIBLIOGRAPHY 1. Bulla R et al. Nat Commun. 2016 Feb 1;7:10346. 2. Son M et al. Blood. 2016 Nov 3;128(18):2218-2228. (595

Evaluation of the Interplay Between the Complement Protein C1q and Hyaluronic Acid in Promoting Cell Adhesion

It has been increasingly demonstrated that the tumor microenvironment plays an active role in neoplasia growth and metastasis. Through different pathways, tumor cells can efficiently recruit stromal, immune and endothelial cells by secreting stimulatory factors, chemokines and cytokines. In turn, these cells can alter the signaling properties of the microenvironment by releasing growth-promoting signals, metabolites and extracellular matrix components to sustain high proliferation and metastatic competence. In this context, we identify that the complement component C1q, highly expressed locally by a range of human malignant tumors, upon interacting with the extracellular matrix hyaluronic acid, strongly affects the behavior of primary cells isolated from human tumor specimens. Here, we describe a method to test how C1q bound to hyaluronic acid (HA) impacts tumor cell adhesion, underlying the fact that the biological properties of key components of the extracellular matrix (in this case HA) can be shaped by bioactive signals toward tumor progression. Video Lin

Pre-eclampsia is associated with defective production of C1q by invasive trophoblast

Purpose: We have previously demonstrated that C1q, the first component of the classic complement cascade, is involved in the placentation process acting as a molecular bridge between endovascular trophoblast and decidual endothelial cell1 and promoting trophoblast interstitial invasion2. We observed also a deficient trophoblast invasion in implantation sites of C1q-/- mice if compared to WT animals; so we hypothesized that C1q could had a role in the onset of pre-eclampsia, a multisystem syndrome characterized by a defect of placentation. Methods: Placental mRNA derived from 7 pre-eclamptic (PE) patients and 6 healthy matched controls were analyzed by qPCR for C1q expression. PE sections were stained for C1q and cytokeratin 7 to identify trophoblast. The expression of MMP12 by on freshly isolated trophoblast cells adhering to C1q, FN or poly-L-Lysine was detected by qPCR and immunofluorescence. Results: C1q expression was found to be significantly lower in PE placentae compared to healthy women. Histological evidences on PE decidual sections showed that trophoblast cells surrounding non-remodelled spiral artery do not express C1q in comparison to non pathological placentae. In vitro studies on trophoblast cells demonstrated that the expression of MMP-12, a marker of vascular remodelling3, is upregulated in response to C1q interaction. Discussion: The defective staining of C1q by PE perivascular trophoblast seems to be directly related to absence of vascular remodelling. The upregulation of MMP-12 expression by trophoblast cells in response to C1q indicated a functional role of C1q in trophoblast vascular remodelling. Conclusions: Collectively, these data support the pivotal role played by C1q in placental development. The importance of this component at the placental level is evidenced by its involvement in pregnancy disorders such as pre-eclampsia, characterized by poor trophoblast invasion

Pre-eclampsia affects procalcitonin production in placental tissue

PROBLEM: Procalcitonin (PCT) is the prohormone of calcitonin which is usually released from neuroendocrine cells of the thyroid gland (parafollicular) and the lungs (K cells). PCT is synthesized by almost all cell types and tissues, including monocytes and parenchymal tissue, upon LPS stimulation. To date, there is no evidence for PCT expression in the placenta both in physiological and pathological conditions. METHOD: Circulating and placental PCT levels were analysed in pre-eclamptic (PE) and control patients. Placental cells and macrophages (PBDM), stimulated with PE sera, were analysed for PCT expression. The effect of anti-TNF-\u3b1 antibody was analysed. RESULTS: Higher PCT levels were detected in PE sera and in PE placentae compared to healthy women. PE trophoblasts showed increased PCT expression compared to those isolated from healthy placentae. PE sera induced an upregulation of PCT production in macrophages and placental cells. The treatment of PBDM with PE sera in the presence of anti-TNF-\u3b1 completely abrogated the effect induced by pathologic sera. CONCLUSION: Trophoblast cells are the main producer of PCT in PE placentae. TNF-\u3b1, in association with other circulating factors present in PE sera, upregulates PCT production in macrophages and normal placental cells, thus contributing to the observed increased in circulating PCT in PE sera