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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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Balduit, Vidergar, Zacchi, Mangogna, Agostinis, Grandolfo, Bottin, Salton, Confalonieri, Rocca, Zanconati, Confalonieri, Kishore, Ghebrehiwet and Bulla.)

Although only 0.8-1% of SARS-CoV-2 infections are in the 0-9 age-group, pneumonia is still the leading cause of infant mortality globally. Antibodies specifically directed against SARS-CoV-2 spike protein (S) are produced during severe COVID-19 manifestations. Following vaccination, specific antibodies are also detected in the milk of breastfeeding mothers. Since antibody binding to viral antigens can trigger activation of the complement classical - pathway, we investigated antibody-dependent complement activation by anti-S immunoglobulins (Igs) present in breast milk following SARS-CoV-2 vaccination. This was in view of the fact that complement could play a fundamentally protective role against SARS-CoV-2 infection in newborns. Thus, 22 vaccinated, lactating healthcare and school workers were enrolled, and a sample of serum and milk was collected from each woman. We first tested for the presence of anti-S IgG and IgA in serum and milk of breastfeeding women by ELISA. We then measured the concentration of the first subcomponents of the three complement pathways (i.e., C1q, MBL, and C3) and the ability of anti-S Igs detected in milk to activate the complement in vitro. The current study demonstrated that vaccinated mothers have anti-S IgG in serum as well as in breast milk, which is capable of activating complement and may confer a protective benefit to breastfed newborns.

Nonhealing wounds place a significant burden on both quality of life of affected patients and health systems. Skin substitutes are applied to promote the closure of nonhealing wounds, although their efficacy is limited by inadequate vascularization. The stromal vascular fraction (SVF) from the adipose tissue is a promising therapy to overcome this limitation. Despite a few successful clinical trials, its incorporation in the clinical routine has been hampered by their inconsistent results. All these studies concluded by warranting pre-clinical work aimed at both characterizing the cell types composing the SVF and shedding light on their mechanism of action. Here, we established a model of nonhealing wound, in which we applied the SVF in combination with a clinical-grade skin substitute. We purified the SVF cells from transgenic animals to trace their fate after transplantation and observed that it gave rise to a mature vascular network composed of arteries, capillaries, veins, as well as lymphatics, structurally and functionally connected with the host circulation. Then we moved to a human-in-mouse model and confirmed that SVF-derived endothelial cells formed hybrid human-mouse vessels, that were stabilized by perivascular cells. Mechanistically, SVF-derived endothelial cells engrafted and expanded, directly contributing to the formation of new vessels, while a population of fibro-adipogenic progenitors stimulated the expansion of the host vasculature in a paracrine manner. These data have important clinical implications, as they provide a steppingstone toward the reproducible and effective adoption of the SVF as a standard care for nonhealing wounds., (© 2023. The Author(s).)

C1q, the recognition molecule of the classical pathway of the complement system, plays a central role in pregnancy. Lack of C1q is characterized by poor trophoblast invasion and pregnancy failure. C1q can be the target of an antibody response: anti-C1q autoantibodies (anti-C1q) are present in several infectious and autoimmune diseases. The presence of these autoantibodies has been detected also in 2-8% of the general population. Recent evidence indicates that women who undergo assisted reproductive technology (ART) have an increased risk of developing pre-eclampsia (PE), particularly oocyte donation (OD) pregnancies. The aim of this study was to characterize the levels of C1q and anti-C1q in PE gestations, in healthy spontaneous, homologous and heterologous ART pregnancies. Serum of the following four groups of women, who were followed throughout two or three trimesters, were collected: PE, patients diagnosed with PE; OD, oocyte donation recipients; HOM, homologous ART women; Sp, spontaneous physiological pregnancy. Our results indicate that PE patients have lower levels of anti-C1q. In ART pregnant women, the trend of C1q and anti-C1q levels were similar to PE patients, even though these women did not develop PE-like symptoms during pregnancy. This finding suggests an immunological dysfunction at the foetal-maternal interface in ART pregnancies, a hypothesis confirmed by the observation of C1q deposition in placentae derived from OD, comparable to PE. Since significantly lower levels of anti-C1q were detected in PE compared to healthy control sera, we hypothesize the possible binding on placental syncytiotrophoblast microvesicles (STBM), which are increased in the circulation of PE mothers. Furthermore, the characterization of the binding-epitope of anti-C1q revealed that "physiological" autoantibodies were mainly directed against C1q globular domain. We concluded that anti-C1q could have a physiological role in pregnancy: during the healthy spontaneous pregnancy the raised levels of these autoantibodies can be important for the clearance of STBM. In PE and in pathological pregnancies (but also in OD pregnancies), the increase in syncytiotrophoblast apoptosis and consequent increase of the circulating STMB levels lead to a consumption of C1q and anti-C1q., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Agostinis, Zito, Toffoli, Peterlunger, Simoni, Balduit, Curtolo, Mangogna, Belmonte, Vacca, Romano, Stampalija, Salviato, Defendi, Di Simone, Kishore, Ricci and Bulla.)

SARS-CoV-2 is a devastating virus that induces a range of immunopathological mechanisms including cytokine storm, apoptosis, inflammation and complement and coagulation pathway hyperactivation. However, how the infection impacts pregnant mothers is still being worked out due to evidence of vertical transmission of the SARS-CoV-2, and higher incidence of pre-eclampsia, preterm birth, caesarian section, and fetal mortality. In this study, we assessed the levels of the three main receptors of SARS-CoV-2 (ACE2, TMPRSS2 and CD147) in placentae derived from SARS-CoV-2 positive and negative mothers. Moreover, we measured the effects of Spike protein on placental cell lines, in addition to their susceptibility to infection. SARS-CoV-2 negative placentae showed elevated levels of CD147 and considerably low amount of TMPRSS2, making them non-permissive to infection. SARS-CoV-2 presence upregulated TMPRSS2 expression in syncytiotrophoblast and cytotrophoblast cells, thereby rendering them amenable to infection. The non-permissiveness of placental cells can be due to their less fusogenicity due to infection. We also found that Spike protein was capable of inducing pro-inflammatory cytokine production, syncytiotrophoblast apoptosis and increased vascular permeability. These events can elicit pre-eclampsia-like syndrome that marks a high percentage of pregnancies when mothers are infected with SARS-CoV-2. Our study raises important points relevant to SARS-CoV-2 mediated adverse pregnancy outcomes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Agostinis, Toffoli, Spazzapan, Balduit, Zito, Mangogna, Zupin, Salviato, Maiocchi, Romano, Crovella, Fontana, Braga, Confalonieri, Ricci, Kishore and Bulla.)

Pre-eclampsia is a pregnancy complication characterized by defective vascular remodeling in maternal decidua responsible for reduced blood flow leading to functional and structural alterations in the placenta. We have investigated the contribution of the complement system to decidual vascular changes and showed that trophoblasts surrounding unremodeled vessels prevalent in preeclamptic decidua fail to express C1q that are clearly detected in cells around remodeled vessels predominant in control placenta. The critical role of C1q is supported by the finding that decidual trophoblasts of female C1qa -/- pregnant mice mated to C1qa +/+ male mice surrounding remodeled vessels express C1q of paternal origin. Unlike C1qa -/- pregnant mice, heterozygous C1qa +/- and wild type pregnant mice share a high percentage of remodeled vessels. C1q was also found in decidual vessels and stroma of normal placentae and the staining was stronger in preeclamptic placentae. Failure to detect placental deposition of C1r and C1s associated with C1q rules out complement activation through the classical pathway. Conversely, the intense staining of decidual endothelial cells and villous trophoblast for ficolin-3, MASP-1 and MASP-2 supports the activation of the lectin pathway that proceeds with the cleavage of C4 and C3 and the assembly of the terminal complex. These data extend to humans our previous findings of complement activation through the lectin pathway in an animal model of pre-eclampsia and provide evidence for an important contribution of C1q in decidual vascular remodeling., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Belmonte, Mangogna, Gulino, Cancila, Morello, Agostinis, Bulla, Ricci, Fraggetta, Botto, Garred and Tedesco.)

We investigated the role of rhIL-35, at low concentrations compatible with those produced by human trophoblast cells (less than 1 ng/mL), on human T helper (Th) cell functions and the presence of decidual IL-35-producing Th cells in human pregnancy. We found that human trophoblast cells produced IL-35 but not IL-4 or IL-10. RhIL-35, at concentrations produced by human trophoblasts, polarized T cells towards IL-35+, IL-10+, IL-4+ Th2-type cells and to Foxp3+ EBI3+ p35+ T reg cells producing IL-35 but not IL-10 and IL-4. Moreover, rhIL-35 at low concentrations did not suppress the proliferation of Th cells but stimulated IL-4 and IL-10 production by established Th clones. In particular, Th1-type clones acquired the capacity to produce IL-4. In addition, purified human trophoblast cell supernatants containing IL-35 upregulated IL-4 and IL-10 production by Th clones. Finally, IL-35+, IL-10+, IL-4+ Th2-type cells, which were found to be induced by low concentrations of IL-35 compatible with those produced by human trophoblasts, are exclusively present in the decidua of a successful pregnancy and at the embryo implantation site, suggesting their stringent dependence on trophoblast cells. Thus, the proximity of Th cells to IL-35-producing trophoblasts could be the determining factor for the differentiation of IL-35+, IL-10+, IL-4+ Th2-type cells that are crucial for human pregnancy success.

COVID-19 is characterized by virus-induced injury leading to multi-organ failure, together with inflammatory reaction, endothelial cell (EC) injury, and prothrombotic coagulopathy with thrombotic events. Complement system (C) via its cross-talk with the contact and coagulation systems contributes significantly to the severity and pathological consequences due to SARS-CoV-2 infection. These immunopathological mechanisms overlap in COVID-19 and pre-eclampsia (PE). Thus, mothers contracting SARS-CoV-2 infection during pregnancy are more vulnerable to developing PE. SARS-CoV-2 infection of ECs, via its receptor ACE2 and co-receptor TMPRSS2, can provoke endothelial dysfunction and disruption of vascular integrity, causing hyperinflammation and hypercoagulability. This is aggravated by bradykinin increase due to inhibition of ACE2 activity by the virus. C is important for the progression of normal pregnancy, and its dysregulation can impact in the form of PE-like syndrome as a consequence of SARS-CoV-2 infection. Thus, there is also an overlap between treatment regimens of COVID-19 and PE. C inhibitors, especially those targeting C3 or MASP-2, are exciting options for treating COVID-19 and consequent PE. In this review, we examine the role of C, contact and coagulation systems as well as endothelial hyperactivation with respect to SARS-CoV-2 infection during pregnancy and likely development of PE., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Agostinis, Mangogna, Balduit, Aghamajidi, Ricci, Kishore and Bulla.)

The loss of skin integrity has always represented a major challenge for clinicians dealing with dermal defects, such as ulcers (diabetic, vascular and chronic), postoncologic resections (i.e., radical vulvectomy) or dermatologic disorders. The introduction in recent decades of acellular dermal matrices (ADMs) supporting the repair and restoration of skin functionality represented a significant step toward achieving clean wound repair before performing skin grafts. Hard-to-heal ulcers generally depend on local ischemia and nonadequate vascularization. In this context, one possible innovative approach could be the prevascularization of matrices with vessel-forming cells (inosculation). This paper presents a comparative analysis of the most widely used dermal templates, i.e., Integra ® Bilayer Matrix Wound Dressing, PELNAC ® , PriMatrix ® Dermal Repair Scaffold, Endoform ® Natural Dermal Template, and Myriad Matrix ® , testing their ability to be colonized by human adult dermal microvascular endothelial cells (ADMECs) and to induce and support angiogenesis in vitro and in vivo. By in vitro studies, we demonstrated that Integra ® and PELNAC ® possess superior pro-adhesive and pro-angiogenetic properties. Animal models allowed us to demonstrate the ability of preseeded ADMECs on Integra ® to promote the engraftment, integration and vascularization of ADMs at the site of application.

The complement system is a major component of humoral innate immunity, acting as a first line of defense against microbes via opsonization and lysis of pathogens. However, novel roles of the complement system in inflammatory and immunological processes, including in cancer, are emerging. Endometriosis (EM), a benign disease characterized by ectopic endometrial implants, shows certain unique features of cancer, such as the capacity to invade surrounding tissues, and in severe cases, metastatic properties. A defective immune surveillance against autologous tissue deposited in the peritoneal cavity allows immune escape for endometriotic lesions. There is evidence that the glandular epithelial cells found in endometriotic implants produce and secrete the complement component C3. Here, we show, using immunofluorescence and RT-qPCR, the presence of locally synthesized C3 in the ectopic endometriotic tissue, but not in the eutopic tissue. We generated a murine model of EM via injection of minced uterine tissue from a donor mouse into the peritoneum of recipient mice. The wild type mice showed greater amount of cyst formation in the peritoneum compared to C3 knock-out mice. Peritoneal washings from the wild type mice with EM showed more degranulated mast cells compared to C3 knock-out mice, consistent with higher C3a levels in the peritoneal fluid of EM patients. We provide evidence that C3a participates in an auto-amplifying loop leading to mast cell infiltration and activation, which is pathogenic in EM. Thus, C3 can be considered a marker of EM and its local synthesis can promote the engraftment of the endometriotic cysts., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Agostinis, Zorzet, Balduit, Zito, Mangogna, Macor, Romano, Toffoli, Belmonte, Morello, Martorana, Borelli, Ricci, Kishore and Bulla.)

Increased hyaluronic acid (HA) production is often associated with cancer progression. In malignant pleural mesothelioma (MPM), HA is found at elevated levels in pleural effusions and sera of patients, and it has been widely debated whether MPM cells are able to produce HA by themselves or through the release of growth factors stimulating other cells. Another key component of the MPM microenvironment is C1q, which can act as a pro-tumorigenic factor favoring cell adhesion, migration and proliferation. The aim of the current study was to prove that MPM primary cells are able to synthesize HA and to inquire the stimulus given by C1q-HA matrix to HA synthesis. We confirmed the presence of a HA coat and cable-like structures around MPM primary cells, as well as an intracellular pool, mainly localized in the cytoplasmic and perinuclear region. After evaluating HA synthase (HAS) enzymes' basal expression in MPM primary cells, we found that C1q bound to HA was able to impinge upon HA homeostasis by upregulating HAS3 both at the mRNA and the protein levels. High expression of HAS3 has been correlated with a shorter life expectancy in MPM by bioinformatical analysis. These data confirmed that C1q bound to HA may exert pro-tumorigenic activity and identified HAS3 as a potential target in MPM.

The complement system is readily triggered by the presence of damage-associated molecular patterns on the surface of tumor cells. The complement alternative pathway provides rapid amplification of the molecular stress signal, leading to complement cascade activation to deal with pathogens or malignant cells. Properdin is the only known positive regulator of the alternative pathway. In addition, properdin promotes the phagocytic uptake of apoptotic T cells by macrophages and dendritic cells without activating the complement system, thus, establishing its ability to recognize "altered-self". Dysregulation of properdin has been implicated in substantial tissue damage in the host, and in some cases, chronic unresolved inflammation. A corollary of this may be the development of cancer. Hence, to establish a correlation between properdin presence/levels in normal and cancer tissues, we performed bioinformatics analysis, using Oncomine and UALCAN. Survival analyses were performed using UALCAN and PROGgeneV2 to assess if properdin can serve as a potential prognostic marker for human lung adenocarcinoma (LUAD), liver hepatocellular carcinoma (LIHC), cervical squamous cell carcinoma (CESC), and pancreatic adenocarcinoma (PAAD). We also analyzed levels of tumor-infiltrating immune cells using TIMER, a tool for characterizing immune cell composition in cancers. We found that in LUAD and LIHC, there was a lower expression of properdin in the tumors compared to normal tissues, while no significant difference was observed in CESC and PAAD. Survival analysis demonstrated a positive association between properdin mRNA expression and overall survival in all 4 types of cancers. TIMER analysis revealed that properdin expression correlated negatively with tumor purity and positively with levels of infiltrating B cells, cytotoxic CD8 + T cells, CD4 + helper T cells, macrophages, neutrophils and dendritic cells in LUAD, CESC and PAAD, and with levels of B cells, CD8 + T cells and dendritic cells in LIHC. Immunohistochemical analysis revealed that infiltrating immune cells were the most likely source of properdin in the tumor microenvironment. Thus, complement protein properdin shows promise as a prognostic marker in cancer and warrants further study., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Mangogna, Varghese, Agostinis, Alrokayan, Khan, Stover, Belmonte, Martorana, Ricci, Bulla and Kishore.)

Endometriosis (EM) is a chronic disease characterized by the presence and proliferation of functional endometrial glands and stroma outside the uterine cavity. Ovaries and pelvic peritoneum are the most common locations for endometrial ectopic tissue, followed by deep infiltrating EM sites. The cyclic and recurrent bleeding, the progressive fibrosis and the peritoneal adhesions of ectopic endometrial glands, may cause different symptoms depending on the origin involved. EM is a frequent clinical condition affecting around 10% of women of mainly reproductive age, as well as in post-menopausal women and adolescents, especially with uterine anomalies. The risk of developing EM depends on a complex interaction between genetic, immunological, hormonal, and environmental factors. It is largely considered to arise due to a dysfunction of immunological surveillance. In fact, women with EM exhibit altered functions of peritoneal macrophages, lymphocytes and natural killer cells, as well as levels of inflammatory mediators and growth factors in the peritoneal fluid. In EM patients, peritoneal macrophages are preponderant and highly active compared to healthy women. Peritoneal macrophages are able to regulate the events that determine the production of cytokines, prostaglandins, growth factors and complement components. Several studies have shown alteration in the regulation of the complement activation, leading to chronic inflammation characteristic of EM. Aberrant regulation/activation of the complement system has been observed in the peritoneal cavity of women affected by EM. Thus, complement inhibition may represent a new approach for the treatment of EM, given that a number of complement inhibitors are under pre-clinical and clinical development. Such an intervention may provide a broader therapeutic control of complement-mediated inflammatory damage in EM patients. This review will focus on our current understanding of the role of complement activation in EM and possible modalities available for complement-based therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Agostinis, Balduit, Mangogna, Zito, Romano, Ricci, Kishore and Bulla.)

Gastroesophageal reflux disease (GERD) is a common digestive disorder that causes esophagitis and injuries to the esophageal mucosa. GERD symptoms are recurrent during pregnancy and their treatment is focused on lifestyle changes and nonprescription medicines. The aim of this study was to characterize the mechanism of action of a new patented medical device, an oral formulation containing hyaluronic acid, rice extract, and amino acids dispersed in a bioadhesive polymer matrix, by assessing its protective effects in in vitro and ex vivo models of esophageal mucosa damage. Acidic bile salts and pepsin cocktail (BSC) added to CP-A and COLO-680 N esophagus cells were used as an in vitro GERD model to evaluate the binding capacities, anti-inflammatory effects and reparative properties of the investigational product (IP) in comparison to a viscous control. Our results showed that the IP prevents cell permeability and tight junction dysfunction induced by BSC. Furthermore, the IP was also able to down-regulate IL-6 and IL-8 mRNA expression induced by BSC stimulation and to promote tissue repair and wound healing. The results were confirmed by ex vivo experiments in excised rat esophagi through the quantification of Evans Blue permeability assay. These experiments provided evidence that the IP is able to bind to the human esophagus cells, preventing the damage caused by gastroesophageal reflux, showing potential anti-irritative, soothing, and reparative properties., (Copyright © 2020 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Most Cystic Fibrosis (CF) patients succumb to airway inflammation and pulmonary infections due to Pseudomonas aeruginosa . D -BMAP18, a membrane-permeabilizing antimicrobial peptide composed of D-amino acids, was evaluated as a possible antibacterial aimed to address this issue. The antipseudomonal activity of D -BMAP18 was tested in a pathophysiological context. The peptide displayed activity against CF isolates of Pseudomonas aeruginosa in the presence of CF sputum when combined with sodium chloride and DNase I. In combination with DNase I, D -BMAP18 discouraged the deposition of new biofilm and eradicated preformed biofilms of some P. aeruginosa strains. In addition, D -BMAP18 down regulated the production of TNF-α, IL1-β, and TGF-β in LPS-stimulated or IFN-γ macrophages derived from THP-1 cells indicating an anti-inflammatory activity. The biocompatibility of D -BMAP18 was assessed using four different cell lines, showing that residual cell-specific cytotoxicity at bactericidal concentrations could be abolished by the presence of CF sputum. Overall, this study suggests that D -BMAP18 may be an interesting molecule as a starting point to develop a novel therapeutic agent to simultaneously contrast lung infections and inflammation in CF patients.

β2-Glycoprotein I (β2-GPI) is an abundant plasma glycoprotein with unknown physiological function and is currently recognized as the main target of antiphospholipid Abs responsible for complement activation and vascular thrombosis in patients with antiphospholipid syndrome (APS). In this study, we provide evidence that mannose-binding lectin (MBL) binds to β2-GPI in Ca ++ and a dose-dependent manner and that this interaction activates complement and promotes complement-dependent thrombin generation. Surprisingly, a significant binding was observed between MBL and isolated domains II and IV of β2-GPI, whereas the carbohydrate chains, domain I and domain V, were not involved in the interaction, documenting a noncanonical binding mode between MBL and β2-GPI. Importantly, this interaction may occur on endothelial cells because binding of MBL to β2-GPI was detected on the surface of HUVECs, and colocalization of MBL with β2-GPI was observed on the endothelium of a biopsy specimen of a femoral artery from an APS patient. Because β2-GPI-mediated MBL-dependent thrombin generation was increased after priming the endothelium with TNF-α, our data suggests that this mechanism could play an important yet unrecognized role under physiological conditions and may be upregulated in pathological situations. Moreover, the complement activation and the procoagulant effects of the β2-GPI/MBL complex may contribute to amplify similar activities of anti-β2-GPI Abs in APS and possibly act independently of Abs, raising the issue of developing appropriate therapies to avoid recurrences and disability in patients at risk for these clinical conditions., (Copyright © 2020 by The American Association of Immunologists, Inc.)

Purpose: To evaluate the vitreous concentration of different nonsteroidal anti-inflammatory drugs (NSAIDs) after topical administration and the related prostaglandin E2 (PGE2) levels in patients undergoing pars plana vitrectomy., Methods: A prospective, randomized, investigator-masked study was performed. One hundred four patients scheduled for a pars plana vitrectomy for an epiretinal membrane or a macular hole were randomized to receive topical diclofenac 0.1%, indomethacin 0.5%, nepafenac 0.3%, bromfenac 0.09%, or placebo 3 days before surgery. At the beginning of surgery, a sample of undiluted vitreous was collected in each patient to assess NSAIDs concentration and PGE2 levels., Results: The median vitreous concentrations were 203.35 (interquartile range 146.54-264.18) pg/mL for diclofenac, 243.45 (interquartile range 156.96-365.37) pg/mL for nepafenac, 438.21 pg/mL (interquartile range, 282.52-645.87) for its active metabolite amfenac, 350.14 (interquartile range, 290.88-481.95) pg/mL for indomethacin, and 274.59 (245.43-358.25) pg/mL for bromfenac. Vitreous PGE2 levels were significantly lower for all the NSAIDs groups compared with the control group (P < 0.001). A statistically significant higher vitreous PGE2 level was found in the diclofenac group compared with the other NSAIDs groups (P < 0.05)., Conclusion: Topical NSAIDs achieve sufficient vitreous concentration to decrease vitreous PGE2 levels compared with the control group. The different efficacy in reducing PGE2 concentration may affect the management of posterior segment inflammation.

Checkpoint inhibitors have substantially improved the prognosis for patients with advanced malignancy. Treatment with immunomodulants has the ability to reactivate the immune system against tumor cells, but can also trigger the development of immune-related adverse events that reflects a loss of tolerance of the immune system for self-antigens. Regarding the endocrine system, thyroid and pituitary are the most frequent glands involved; in particular hypophysitis is commonly observed with anti-CTLA4 with a variable impaired anterior pituitary dysfunction (mainly ACTH and TSH dysregulation) while a posterior pituitary dysfunction has been rarely described. A 68-year-old man with a diagnosis of metastatic mesothelioma started in September 2016 first-line treatment with tremelimumab and durvalumab. After 3 cycles he presented sudden onset of polydipsia and polyuria without other symptoms. Diagnostic work-up, including a water deprivation test, established a diagnosis of central diabetes insipidus. Patient started sublingual desmopressin 60 mcg three times a day, that was subsequently increased up to 480 mcg/die. At magnetic resonance imaging the posterior lobe of pituitary gland did not show high signal intensity on T1-weighted images. After regression of diabetes insipidus symptoms under desmopressin, patient restarted cancer treatment and received additional 10 doses without worsening of endocrinological toxicity or further treatment-related toxicities, maintaining the same desmopressin dosage. Posterior pituitary dysfunction has been rarely observed in patients treated with immunomodulants. To our knowledge, this is the first observation of permanent central diabetes insipidus in patients treated with combined immune checkpoint inhibitors (tremelimumab and durvalumab).

Human surfactant protein D (SP-D) belongs to the family of collectins that is composed of a characteristic amino-terminal collagenous region and a carboxy-terminal C-type lectin domain. Being present at the mucosal surfaces, SP-D acts as a potent innate immune molecule and offers protection against non-self and altered self, such as pathogens, allergens, and tumor. Here, we examined the effect of a recombinant fragment of human SP-D (rfhSP-D) on a range of breast cancer lines. Breast cancer has four molecular subtypes characterized by varied expressions of estrogen (ER), progesterone (PR), and epidermal growth factor (EGF) receptors (HER2). The cell viability of HER2-overexpressing (SKBR3) and triple-positive (BT474) breast cancer cell lines [but not of a triple-negative cell line (BT20)] was reduced following rfhSP-D treatment at 24 h. Upregulation of p21/p27 cell cycle inhibitors and p53 phosphorylation (Ser15) in rfhSP-D-treated BT474 and SKBR3 cell lines signified G2/M cell cycle arrest. Cleaved caspases 9 and 3 were detected in rfhSP-D-treated BT474 and SKBR3 cells, suggesting an involvement of the intrinsic apoptosis pathway. However, rfhSP-D-induced apoptosis was nullified in the presence of hyaluronic acid (HA) whose increased level in breast tumor microenvironment is associated with malignant tumor progression and invasion. rfhSP-D bound to solid-phase HA and promoted tumor cell proliferation. rfhSP-D-treated SKBR3 cells in the presence of HA showed decreased transcriptional levels of p53 when compared to cells treated with rfhSP-D only. Thus, HA appears to negate the anti-tumorigenic properties of rfhSP-D against HER2-overexpressing and triple-positive breast cancer cells., (Copyright © 2020 Murugaiah, Agostinis, Varghese, Belmonte, Vieni, Alaql, Alrokayan, Khan, Kaur, Roberts, Madan, Bulla and Kishore.)

Background: An aberrant and persistent inflammatory state at the fetal-maternal interface is considered as a key contributor in compromised pregnancies. Decidual endothelial cells (DECs) play a pivotal role in the control of the local decidual inflammation. The aim of the current study was to determine whether dietary supplement with zinc oxide (ZnO), due to its very low adverse effects, may be useful for modulating the inflammatory response in the first trimester of pregnancy., Methods: The anti-inflammatory properties of ZnO in pregnancy were evaluated by in vitro tests on endothelial cells isolated from normal deciduas and on a trophoblast cell line (HTR8/Svneo). The effects of this treatment were analyzed in terms of adhesion molecule expression and inflammatory cytokine secretion, by real time-quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA)., Results: Our data showed that ZnO was able to reduce the inflammatory response of DECs, in terms of vascular cell adhesion molecule-1 (VCAM-1), interleukin (IL)-8, IL-6, tumor necrosis factor-α (TNF-α) and monocyte chemoattractant protein-1 (MCP-1) expression induced by TNF-α stimulation. This compound exerted no effect on intracellular adhesion molecule-1 (ICAM-1) exocytosis induced by TNF-α on stimulated trophoblast cells, but significantly reduced their IL-6 expression., Conclusion: According to these results, it can be suggested that the ZnO supplement, through its modulation of the pro-inflammatory response of DECs, can be used in pregnancy for the prevention of local decidual inflammation.

The development of personalized therapies for ovarian carcinoma patients is still hampered by several limitations, mainly the difficulty of predicting patients' responses to chemotherapy in tumor cells isolated from peritoneal fluids. The main reason for the low predictive power of in vitro assays is related to the modification of the cancer cells' phenotype induced by the culture conditions, which results in changes to the activation state and drug sensitivity of tumor cells compared to their in vivo properties. We have defined the optimal culture conditions to set up a prognostic test to predict high-grade serous ovarian carcinoma (HGSOC) patients' responses to platinum chemotherapy. We evaluated the effects of hyaluronic acid (HA) and fibronectin matrices and the contribution of freezing/thawing processes to the cell response to platinum-based treatment, collecting spheroids from the ascitic fluids of 13 patients with stage II or III HGSOC. Our findings indicated that an efficient model used to generate predictive data for in vivo sensitivity to platinum is culturing fresh spheroids on HA, avoiding the use of previously frozen primary tumor cells. The establishment of this easy, reproducible and standardized testing method can significantly contribute to an improvement in therapeutic effectiveness, thus bringing the prospect of personalized therapy closer for ovarian carcinoma patients.

Background and Objective: Echinacea angustifolia DC. and Zingiber officinale Roscoe are two natural products with documented immunomodulatory activity, both able to modulate the expression of important immune-related genes. Thus, their use in combination seems to be particularly promising. In this context, we have considered the oral supplementation of a highly standardized lipophilic extract combining both above-mentioned phytocomplexes, formulated in attractive softgel capsules, with two objectives: on the one hand to study oral pharmacokinetic of main active extracts' components and on the other hand to examine the immunomodulation and anti-inflammatory properties by gene expression profiling., Methods: Softgel capsules containing a combination of E. angustifolia DC. and Z. officinale Roscoe (5 mg and 25 mg, respectively) were given by oral administration to 10 healthy volunteers. The plasma concentrations of dodeca-2E,4E,8Z,10E/Z-tetraenoic isobutylamide (tetraene) for E. angustifolia DC., 6-gingerol and 6-shogaol (free and glucuronide) for Z. officinale Roscoe were determined by LC-MS analysis, and the pharmacokinetic analysis was performed. To understand the functional mechanisms responsible for the documented health benefits, we also examined the overall transcriptional remodeling induced in the peripheral blood mononuclear cells and performed an integrative functional analysis on the generated gene expression., Results: All bioactive components were absorbed very rapidly, and their t max were detected in plasma from 30 min to 1.40 h. The peak concentrations of tetraene, 6-gingerol, 6-shogaol and their glucuronide metabolites were 14.74, 5.66, 9.25, 29.2 and 22.24 ng/ml, respectively. Integrated analysis performed on the generated gene expression data highlighted immunomodulatory and anti-inflammatory effects similar to those exerted by hydrocortisone., Conclusion: These data demonstrated that the bioactive ingredients are highly and rapidly absorbed from softgel capsules containing the combination of the above-mentioned lipophilic extracts, providing evidence to support their immunomodulatory and anti-inflammatory properties. These data also help in defining the mechanistic pathways underlying the health benefits of these plant-derived bioactive compounds., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

The female reproductive tract harbors distinct microbial communities, as in the vagina, cervical canal, uterus, and fallopian tubes. The nature of the vaginal microbiota is well-known; in contrast, the upper reproductive tract remains largely unexplored. Alteration in the uterine microbiota, which is dependent on the nutrients and hormones available to the uterus, is likely to play an important role in uterine-related diseases such as hysteromyoma, adenomyosis, and endometriosis. Uterine mucosa is an important tissue barrier whose main function is to offer protection against pathogens and other toxic factors, while maintaining a symbiotic relationship with commensal microbes. These characteristics are shared by all the mucosal tissues; however, the uterine mucosa is unique since it changes cyclically during the menstrual cycle as well as pregnancy. The immune system, besides its role in the defense process, plays crucial roles in reproduction as it ensures local immune tolerance to fetal/paternal antigens, trophoblast invasion, and vascular remodeling. The human endometrium contains a conspicuous number of immune cells, mainly Natural Killers (NK) cells, which are phenotypically distinct from peripheral cytotoxic NK, cells and macrophages. The endometrium also contains few lymphoid aggregates comprising B cell and CD8 + T cells. The number and the phenotype of these cells change during the menstrual cycle. It has become evident in recent years that the immune cell phenotype and function can be influenced by microbiota. Immune cells can sense the presence of microbes through their pattern recognition receptors, setting up host-microbe interaction. The microbiota exerts an appropriately controlled defense mechanism by competing for nutrients and mucosal space with pathogens. It has recently been considered that uterus is a non-sterile compartment since it seems to possess its own microbiota. There has been an increasing interest in characterizing the nature of microbial colonization within the uterus and its apparent impact on fertility and pregnancy. This review will examine the potential relationship between the uterine microbiota and the immune cells present in the local environment., (Copyright © 2019 Agostinis, Mangogna, Bossi, Ricci, Kishore and Bulla.)

The contribution of the complement system in the pathophysiology of brain cancers has been recently considered in light of its well-known involvement in carcinogenesis. Complement system represents an important component of the inflammatory response, which acts as a functional bridge between the innate and adaptive immune response. C1q, the first recognition subcomponent of the complement classical pathway, has recently been shown to be involved in a range of pathophysiological functions that are not dependent on complement activation. C1q is expressed in the microenvironment of various types of human tumors, including melanoma, prostate, mesothelioma, and ovarian cancers, where it can exert a protective or a harmful effect on cancer progression. Despite local synthesis of C1q in the central nervous system, the involvement of C1q in glioma pathogenesis has been poorly investigated. We, therefore, performed a bioinformatics analysis, using Oncomine dataset and UALCAN database in order to assess whether the expression of the genes encoding for the three chains of C1q ( C1qA, C1qB , and C1qC ) could serve as a potential prognostic marker for gliomas. The obtained results were then validated using an independent glioma cohort from the Chinese Glioma Genome Atlas datasets. Our bioinformatics analysis, coupled with immunohistochemistry and fluorescence microscopy, appears to suggest a positive correlation between higher levels of C1q expression and unfavorable prognosis in a diverse grade of gliomas., (Copyright © 2019 Mangogna, Belmonte, Agostinis, Zacchi, Iacopino, Martorana, Rodolico, Bonazza, Zanconati, Kishore and Bulla.)

Background: Prognostic factors affecting outcome of traumatic brain injury (TBI), despite their importance, are still under discussion. The purpose of this study was to describe risk factors of in-hospital mortality and outcome at 1 year in a homogeneously treated population of patients with moderate/severe TBI., Methods: A total of 193 consecutive patients with moderate or severe TBI (Glasgow Coma Scale [GCS] score 13-3, including patients with initial GCS score of 13 at high risk for subsequent neurologic deterioration), admitted to the intensive care unit, were retrospectively analyzed. In-hospital mortality and unfavorable outcome at 1 year, based on a Glasgow Outcome Scale-Extended score ≤4, were considered as primary and secondary outcomes., Results: At 1 year, unfavorable outcome occurred in 47.2%, including an in-hospital mortality of 19.7%. Increasing age, GCS motor score <3, coagulation disorders, and intracranial hypertension were acute risk factors of in-hospital mortality. In the 155 remaining survivors, Oxford Handicap Scale (OHS), posttraumatic cerebral infarction, cerebrospinal fluid disturbances, and length of intensive care unit stay were associated with unfavorable outcome at 1 year, in univariate analysis. A cutoff OHS score ≥3 discriminated the probability of an unfavorable outcome (area under the curve, 0.87; P < 0.001; specificity, 74%; sensitivity, 84%). Combining the effect of acute and subacute variables in a multivariate analysis, increasing age and OHS score were independent predictors of outcome., Conclusions: The results of this retrospective study confirmed age as the main acute risk factor and identified OHS as new potential subacute predictor of unfavorable outcome in moderate and severe TBI., (Copyright © 2019 Elsevier Inc. All rights reserved.)

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.

Aortic coarctations in adults are mainly represented by recurrent critical narrowing at the site of previous surgical correction, or less frequently by native forms of complex obstructive malformations of the distal arch and isthmus. We present our experience with an unusual form of native adult aortic coarctation presenting as a complete interruption of the aortic arch., Competing Interests: The authors declare no conflict of interest related to the article., (Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.)

C1q is the first subcomponent of the classical pathway of the complement system and belongs to the C1q/Tumor Necrosis Factor superfamily. C1q can perform a diverse range of immune and non-immune functions in a complement-dependent as well as -independent manner. Being a pattern recognition molecule of the innate immunity, C1q can recognize a number of self, non-self and altered-self ligands and bring about effector mechanisms designed to clear pathogens via opsonisation and inflammatory response. C1q is locally synthesized by macrophages and dendritic cells, and thus, can get involved in a range of biological processes, such as angiogenesis and tissue remodeling, immune modulation, and immunologic tolerance. The notion of C1q involvement in the pathogenesis of cancer is still evolving. C1q appears to have a dual role in cancer: tumor promoting as well as tumor-protective, depending on the context of the disease. In the current study, we performed a bioinformatics analysis to investigate whether C1q can serve as a potential prognostic marker for human carcinoma. We used the Oncomine database and the survival analysis platforms Kaplan-Meier plotter. Our results showed that high levels of C1q have a favorable prognostic index in basal-like breast cancer for disease-free survival, and in HER2-positive breast cancer for overall survival, while it showed a pro-tumorigenic role of C1q in lung adenocarcinoma, and in clear cell renal cell carcinoma. This in silico study, if validated via a retrospective study, can be a step forward in establishing C1q as a new tool as a prognostic biomarker for various carcinoma.

Background: In pregnancy, excessive inflammation and break down of immunologic tolerance can contribute to miscarriage. Endothelial cells (ECs) are able to orchestrate the inflammatory processes by secreting pro-inflammatory mediators and bactericidal factors by modulating leakiness and leukocyte trafficking, via the expression of adhesion molecules and chemokines. The aim of this study was to analyse the differences in the phenotype between microvascular ECs isolated from decidua (DECs) and ECs isolated from human skin (ADMECs)., Methods: DECs and ADMECs were characterized for their basal expression of angiogenic factors and adhesion molecules. A range of immunological responses was evaluated, such as vessel leakage, reactive oxygen species (ROS) production in response to TNF-α stimulation, adhesion molecules expression and leukocyte migration in response to TNF-α and IFN-γ stimulation., Results: DECs produced higher levels of HGF, VEGF-A and IGFBP3 compared to ADMECs. DECs expressed adhesion molecules, ICAM-2 and ICAM-3, and a mild response to TNF-α was observed. Finally, DECs produced high levels of CXCL9/MIG and CXCL10/IP-10 in response to IFN-γ and selectively recruited Treg lymphocytes., Conclusion: DEC phenotype differs considerably from that of ADMECs, suggesting that DECs may play an active role in the control of immune response and angiogenesis at the foetal-maternal interface.

Surfactant protein D (SP-D) is a pattern recognition molecule belonging to the Collectin (collagen-containing C-type lectin) family that has pulmonary as well as extra-pulmonary existence. In the lungs, it is a well-established opsonin that can agglutinate a range of microbes, and enhance their clearance via phagocytosis and super-oxidative burst. It can interfere with allergen-IgE interaction and suppress basophil and mast cell activation. However, it is now becoming evident that SP-D is likely to be an innate immune surveillance molecule against tumor development. SP-D has been shown to induce apoptosis in sensitized eosinophils derived from allergic patients and a leukemic cell line via p53 pathway. Recently, SP-D has been shown to suppress lung cancer progression via interference with the epidermal growth factor signaling. In addition, a truncated form of recombinant human SP-D has been reported to induce apoptosis in pancreatic adenocarcinoma via Fas-mediated pathway in a p53-independent manner. To further establish a correlation between SP-D presence/levels and normal and cancer tissues, we performed a bioinformatics analysis, using Oncomine dataset and the survival analysis platforms Kaplan-Meier plotter, to assess if SP-D can serve as a potential prognostic marker for human lung cancer, in addition to human gastric, breast, and ovarian cancers. We also analyzed immunohistochemically the presence of SP-D in normal and tumor human tissues. We conclude that (1) in the lung, gastric, and breast cancers, there is a lower expression of SP-D than normal tissues; (2) in ovarian cancer, there is a higher expression of SP-D than normal tissue; and (3) in lung cancer, the presence of SP-D could be associated with a favorable prognosis. On the contrary, at non-pulmonary sites such as gastric, breast, and ovarian cancers, the presence of SP-D could be associated with unfavorable prognosis. Correlation between the levels of SP-D and overall survival requires further investigation. Our analysis involves a large number of dataset; therefore, any trend observed is reliable. Despite apparent complexity within the results, it is evident that cancer tissues that produce less levels of SP-D compared to their normal tissue counterparts are probably less susceptible to SP-D-mediated immune surveillance mechanisms via infiltrating immune cells.

The tumor suppressor DAB2IP contributes to modulate the network of information established between cancer cells and tumor microenvironment. Epigenetic and post-transcriptional inactivation of this protein is commonly observed in multiple human malignancies, and can potentially favor progression of tumors driven by a variety of genetic mutations. Performing a high-throughput screening of a large collection of human microRNA mimics, we identified miR-149-3p as a negative post-transcriptional modulator of DAB2IP. By efficiently downregulating DAB2IP, this miRNA enhances cancer cell motility and invasiveness, facilitating activation of NF-kB signaling and promoting expression of pro-inflammatory and pro-angiogenic factors. In addition, we found that miR-149-3p secreted by prostate cancer cells induces DAB2IP downregulation in recipient vascular endothelial cells, stimulating their proliferation and motility, thus potentially remodeling the tumor microenvironment. Finally, we found that inhibition of endogenous miR-149-3p restores DAB2IP activity and efficiently reduces tumor growth and dissemination of malignant cells. These observations suggest that miR-149-3p can promote cancer progression via coordinated inhibition of DAB2IP in tumor cells and in stromal cells.

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-α 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-α completely abrogated the effect induced by pathologic sera., Conclusion: Trophoblast cells are the main producer of PCT in PE placentae. TNF-α, 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., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

C1q is the first recognition subcomponent of the complement classical pathway, which acts toward the clearance of pathogens and apoptotic cells. C1q is also known to modulate a range of functions of immune and non-immune cells, and has been shown to be involved in placental development and sensorial synaptic pruning. We have recently shown that C1q can promote tumor by encouraging their adhesion, migration, and proliferation in addition to angiogenesis and metastasis. In this study, we have examined the role of human C1q in the microenvironment of malignant pleural mesothelioma (MPM), a rare form of cancer commonly associated with exposure to asbestos. We found that C1q was highly expressed in all MPM histotypes, particularly in epithelioid rather than in sarcomatoid histotype. C1q avidly bound high and low molecular weight hyaluronic acid (HA) via its globular domain. C1q bound to HA was able to induce adhesion and proliferation of mesothelioma cells (MES) via enhancement of ERK1/2, SAPK/JNK, and p38 phosphorylation; however, it did not activate the complement cascade. Consistent with the modular organization of the globular domain, we demonstrated that C1q may bind to HA through ghA module, whereas it may interact with human MES through the ghC. In conclusion, C1q highly expressed in MPM binds to HA and enhances the tumor growth promoting cell adhesion and proliferation. These data can help develop novel diagnostic markers and molecular targets for MPM.

Complement component C1q is one of the recognition molecules of the complement system which can serve several functions unrelated to complement activation. This molecule is produced at foeto-maternal interface by macrophages as wells as by decidual endothelial cells and invading trophoblast. Foetal trophoblast cells migrating through the decidua in the early stages of pregnancy synthesize and express C1q on their surface, which is actively involved in promoting trophoblast endovascular and interstitial invasion of the decidua. These functions are mediated by two cell surface receptors, gC1qR and α4β1 integrin, which promote trophoblast adhesion and migration through the activation of ERK1/2 MAPKs. C1q -/- mice manifest increased frequency of foetal resorption, reduced foetal weight, and smaller litter size when compared to their wild-type counterparts, suggesting that defective local production of C1q may be involved in pregnancy disorders, such as pre-eclampsia. C1q acts also as a strong angiogenic factor and promotes neovascularization. These studies suggest novel and unexpected roles of this complement component in physiological and pathological pregnancies., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Problem: We have previously found that C1q is constitutively expressed by invading trophoblast and endothelial cells of decidua and contributes to vascular and tissue remodeling. Based on these findings, we sought to determine whether there were changes in the circulating level of C1q that may be used as a diagnostic and predictive marker of preeclampsia., Method of Study: We measured the levels of C1q, C4, and complement activation products in serum or plasma of normal pregnant women and preeclamptic patients from different cohorts., Results: We observed a marked decrease in the concentration of C1q associated with a reduced level of C4 in preeclamptic patients as compared to matched healthy pregnant woman but no significant difference in the circulating level of the activating products C5a and the soluble terminal complement complex sC5b-9. Analysis of serum samples collected at early phase of pregnancy from women who later developed preeclampsia failed to show a decrease in C1q level., Conclusion: The results of the present investigation demonstrate that low levels of C1q and C4 are associated with preeclampsia but cannot be used as predictive markers., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Overweight and obesity have been associated with a pro-inflammatory state. We aimed to assess the ability of different measures of overall and abdominal adiposity for identifying adverse levels of inflammatory and metabolic markers in adolescents., Methods: This is a cross-sectional analysis with 529 Portuguese adolescents (267 girls), mean age 14.3 ± 1.7 years. Weight, height, sitting height, waist circumference (WC), and body fat percentage (BF%) were measured; and BMI, waist-to-height ratio (WHtR), and waist-to-sitting-height ratio (WsHtR) were calculated. We measured C-reactive protein (CRP), fibrinogen, erythrocyte sedimentation rate (ESR), complements C3 and C4, leptin, and adiponectin levels. Receiver operating characteristic (ROC) curves were used to estimate the ability of the adiposity measures to discriminate between low/high values of biomarkers., Results: The highest areas under the ROC curves were presented by BF% for fibrinogen and complement C3 in both sexes and for ESR, complement C4, and adiponectin only in girls; by BMI for CRP in girls and for leptin in both sexes; by WHtR for leptin in both sexes and for CRP, fibrinogen, and adiponectin only in girls; by waist circumference for CRP, fibrinogen, and complement C3 only in boys and for complement C4 in girls; and by WsHtR for complement C3 in girls; p < 0.05 for all., Conclusions: The measures that more often presented discriminatory power were, for overall adiposity, BF% in both sexes, and for abdominal adiposity, WHtR in girls and WC in boys. However, small differences in discriminatory capabilities don't allow us to clearly defend the adoption of a single measure above all others.

Complement C1q is the activator of the classical pathway. However, it is now recognized that C1q can exert functions unrelated to complement activation. Here we show that C1q, but not C4, is expressed in the stroma and vascular endothelium of several human malignant tumours. Compared with wild-type (WT) or C3- or C5-deficient mice, C1q-deficient (C1qa(-/-)) mice bearing a syngeneic B16 melanoma exhibit a slower tumour growth and prolonged survival. This effect is not attributable to differences in the tumour-infiltrating immune cells. Tumours developing in WT mice display early deposition of C1q, higher vascular density and an increase in the number of lung metastases compared with C1qa(-/-) mice. Bone marrow (BM) chimeras between C1qa(-/-) and WT mice identify non-BM-derived cells as the main local source of C1q that can promote cancer cell adhesion, migration and proliferation. Together these findings support a role for locally synthesized C1q in promoting tumour growth.

An attractive herbal product, softgel capsules containing 10 mg of Echinacea angustifolia lipophilic extract, was given in a single oral administration to 10 human volunteers to perform a pharmacokinetic and immunological study. The plasma concentration of the major constituent was monitored, quantifying at predetermined time points the dodeca-2E,4E,8Z,10E/Z-tetraenoic isobutylamides (tetraene). The plasmatic levels of IL-2, IL-6, IL-8, IL-10 and TNF-a in samples collected before and 24 h after drug administration were analyzed by cytokine assay. The total RNA was extracted from limpho-monocyte isolated from the same blood samples and the same cytokines in terms of gene expression were evaluated. With the help of proper statistical tests the differences between the values obtained at 0 and 24 h were evaluated. Results of pharmacokinetic studies attest an approximately 3.5-fold improvement of tetraene oral bioavailability compared with previously published studies. Dodeca-2E,4E-dienoic acid isobutylamide exerts immunomodulatory effects down-regulating the gene expression and reducing the protein plasmatic levels of pro-inflammatory cytokines such as IL-6, TNF-α and IL-8, and up-regulating the expression of anti-inflammatory molecules as IL-10. Student's two-sided paired t-test and non-parametric Wilcoxon-Mann-Whitney signed rank test agree in the conclusions about the differences between the ln values at 24 h and corresponding ln values at 0 h., (Copyright © 2015 Elsevier B.V. All rights reserved.)

It is known that excessive inflammation at fetal-maternal interface is a key contributor in a compromised pregnancy. Female genital tract is constantly in contact with microorganisms and several strategies must be adopted to avoid pregnancy failure. Decidual endothelial cells (DECs) lining decidual microvascular vessels are the first cells that interact with pro-inflammatory stimuli released into the environment by microorganisms derived from gestational tissues or systemic circulation. Here, we show that DECs are hypo-responsive to LPS stimulation in terms of IL-6, CXCL8 and CCL2 production. Our results demonstrate that DECs express low levels of TLR4 and are characterized by a strong constitutive activation of the non-canonical NF-κB pathway and a low responsiveness of the canonical pathway to LPS. In conclusion, DECs show a unique hypo-responsive phenotype to the pro-inflammatory stimulus LPS in order to control the inflammatory response at feto-maternal interface.

Streptococcus pyogenes is a rare but aggressive cause of meningitis, which often evolves in a poor outcome with fatal consequences. Although lumbar puncture and CT scan of the brain are the gold standard of diagnosis of cerebral infections, they can have some limitations. We report and describe the clinical history and neuroimaging of a 36-year-old woman admitted to the emergency department of our hospital three days after the onset of earache and otorrhoea. When the patient developed an emergent refractory status epilepticus, the CT scan of the brain showed an unusual pneumocephalus. However, the MRI study of the brain revealed a pachymeningitis with partial thrombosis of the right transverse sinus and subdural empyema due to a S. pyogenes otitis media. Prompt diagnosis and the specific findings of the MRI allowed rapid correct treatment and thus led to a good outcome for the patient.

Neonatal haemochromatosis is a rare alloimmune gestational disease with a high mortality. The hallmark of neonatal haemochromatosis is severe neonatal liver failure associated with extrahepatic siderosis. Thus far, no pituitary dysfunction has been reported to result from the tissue damage associated with extrahepatic siderosis. The present report describes a neonate with neonatal haemochromatosis and secondary hypothyroidism associated with pituitary iron deposition. Both the conditions were successfully treated by ABO-incompatible liver transplantation. Pituitary gland dysfunction is another possible extrahepatic manifestation of neonatal haemochromatosis, and it is reversible after liver transplantation., (© 2014 Steunstichting ESOT.)

A single-chain fragment variable (scFv) recognizing β2-glycoprotein 1 (β2GPI) from humans and other species was isolated from a human phage display library and engineered to contain an IgG1 hinge-CH2-CH3 domain. The scFv-Fc directed against β2GPI domain I-induced thrombosis and fetal loss, thus mimicking the effect of antibodies from patients with antiphospholipid syndrome (APS). Complement is involved in the biological effect of anti-β2GPI scFv-Fc, as demonstrated by its ability to promote in vitro and in vivo complement deposition and the failure to induce vascular thrombosis in C6-deficient rats and fetal loss in C5-depleted mice. A critical role for complement was also supported by the inability of the CH2-deleted scFv-Fc to cause vessel occlusion and pregnancy failure. This antibody prevented the pathological effects of anti-β2GPI antibodies from APS patients and displaced β2GPI-bound patient antibodies. The CH2-deleted antibody represents an innovative approach potentially useful to treat APS patients refractory to standard therapy., (© 2014 by The American Society of Hematology.)

We have previously shown that C1q is expressed on endothelial cells (ECs) of newly formed decidual tissue. Here we demonstrate that C1q is deposited in wound-healing skin in the absence of C4 and C3 and that C1q mRNA is locally expressed as revealed by real-time PCR and in situ hybridization. C1q was found to induce permeability of the EC monolayer, to stimulate EC proliferation and migration, and to promote tube formation and sprouting of new vessels in a rat aortic ring assay. Using a murine model of wound healing we observed that vessel formation was defective in C1qa(-/-) mice and was restored to normal after local application of C1q. The mean vessel density of wound-healing tissue and the healed wound area were significantly increased in C1q-treated rats. On the basis of these results we suggest that C1q may represent a valuable therapeutic agent that can be used to treat chronic ulcers or other pathological conditions in which angiogenesis is impaired, such as myocardial ischemia.

Diabetic nephropathy is the leading cause of chronic renal disease and one of the major causes of cardiovascular mortality. Evidence suggests that its progression is due to the chronic hyperglycemia consequent to the production and accumulation of advanced glycation endproducts (AGEs). Lysozyme was shown to posses AGE-sequestering properties and the capacity to reduce the severity of the early stage manifestations of the diabetic nephropathy. This study was aimed to contribute to the understanding the molecular mechanisms of lysozyme effectiveness in the diabetic nephropathy, using an in-vitro cellular model, represented by the HK-2 cells, human proximal tubular epithelial cells. Lysozyme significantly reduced the AGE-induced IL-6 mRNA and an ELISA assay showed also a decreased release of the functional protein with a dose-dependent trend. In addition, lysozyme prevented macrophage recruitment, suggesting its capacity to elicit an anti-inflammatory action. We may conclude that the protective action of lysozyme on the nephrotoxic effects of AGE may depend, at least in part, on its ability to prevent the production and release of inflammatory mediators, such as IL-6 and to reduce macrophage recruitment in the inflammatory sites.

Procalcitonin (PCT) is one of the best diagnostic and prognostic markers in clinical practice, widely used to evaluate the evolution of bacterial infections. Although it is mainly produced by thyroid, during sepsis almost all the peripheral tissues are involved in PCT production. Parenchymal cells have been suggested as the main source of PCT expression; however the contribution of macrophages is not clear yet. In response to environmental cues, tissue macrophages acquire distinct functional phenotypes, ranging from proinflammatory (M1) to anti-inflammatory (M2) phenotype. Macrophages at the fetal-maternal interface show immunosuppressive M2-like activities required for the maintenance of immunological homeostasis during pregnancy. This study aims to clarify the ability to synthesise PCT of fully differentiated (M0), polarized (M1/M2) macrophages and those cultured either in the presence of first trimester gravid serum (GS) or pregnancy hormones. We found out that M1 macrophages upregulate PCT expression following LPS stimulation compared to M0 and M2. The GS downregulates PCT expression in macrophages, skewing them towards an M2-like phenotype. This effect seems only partially mediated by the hormonal milieu. Our findings strengthen the key role of macrophages in counteracting inflammatory stimuli during pregnancy, suggesting PCT as a possible new marker of M1-like macrophages.

Purpose: Because the nongenotoxic inhibitor of the p53/MDM2 interactions Nutlin-3 has shown promising in vitro therapeutic activity against a variety of p53(wild-type) cancer cells, in this study we evaluated an innovative strategy able to specifically target Nutlin-3 toward CD20(+) malignant cells., Experimental Design: The cytotoxic effects of Nutlin-3 encapsulated into poly(lactide-co-glycolide) nanoparticles (NP-Nut) and into rituximab (anti-CD20 antibody)-engineered NP (NP-Rt-Nut) as well as of NPs engineered with rituximab alone (NP-Rt) were initially analyzed in vitro in JVM-2 B-leukemic cells, by assessing both the functional activation of the p53 pathway (by Nutlin-3) and/or the activation of the complement cascade (by rituximab). Moreover, the potential therapeutic efficacy of NP-Nut, NP-Rt, and NP-Rt-Nut were comparatively assessed in vivo in CD20(+) JVM-2 leukemic xenograft SCID mice., Results: Functional in vitro assays showed that NP-Nut and NP-Rt-Nut exhibited a comparable ability to activate the p53 pathway in the p53(wild-type) JVM-2 leukemic cells. On the other hand, NP-Rt and NP-Rt-Nut, but not NP nor NP-Nut, were able to promote activation of the complement cascade. Of note, the in vivo intratumoral injection in JVM-2 B-leukemic/xenograft mice showed that NP-Rt-Nut displayed the maximal therapeutic activity promoting a survival rate significantly higher not only with respect to control animals, treated either with vehicle or with empty NP, but also with respect to animals treated with NP-Nut or NP-Rt., Conclusions: Our data show for the first time the potential antileukemic activity of rituximab-engineered Nutlin-3-loaded NPs in xenograft SCID mice.

The use of inhaled nitric oxide in patients with traumatic brain injury, intracranial hypertension, and acute respiratory distress syndrome (ARDS) has been reported in an intensive care unit setting only in a few case reports. The use of the prone position for patients with traumatic brain injury and lung impairment has been reported only in selected cases. Here we report our experience with the use of both inhaled nitric oxide and the prone position together in the operating room in a patient with head injury and ARDS who underwent column stabilization.