MFAP4-Deficiency Aggravates Age-Induced Changes in Resistance Artery Structure, While Ameliorating Hypertension.

BACKGROUND: Abnormalities of resistance arteries may play essential roles in the pathophysiology of aging and hypertension. Deficiency of the vascular extracellular matrix protein MFAP4 (microfibrillar-associated protein 4) has previously been observed as protective against aberrant arterial remodeling. We hypothesized that MFAP4-deficiency would reduce age- and hypertension-dependent arterial changes in extracellular matrix composition and stiffening. METHODS: Mesenteric arteries were isolated from old (20-23 months) littermate Mfap4+/+ and Mfap4-/- mice, and 2-photon excitation microscopy imaging was used to quantify elastin and collagen volumes and dimensions in the vascular wall. Ten-week-old littermate Mfap4+/+ and Mfap4-/- mice were subjected to 20 days of continuous Ang II (angiotensin II) infusion and hypertension was monitored using invasive blood pressure measurements. Arterial stiffness, responses to vascular constrictors, and myogenic tone were monitored using wire- or pressure-myography. Collagen contents were assessed by Western blotting. RESULTS: MFAP4-deficiency significantly increased collagen volume and elastin fragmentation in aged mesenteric arteries without affecting arterial stiffness. MFAP4-deficient mice exhibited reduced diastolic pressure in Ang II-induced hypertension. There was no significant effect of MFAP4-deficiency on mesenteric artery structural remodeling or myogenic tone, although collagen content in mesenteric arteries was tendentially increased in hypertensive Mfap4+/+ mice relative to Mfap4-/- mice. Increased efficacy of vasoconstrictors (phenylephrine, thromboxane) and reduced stiffness were observed in Ang II-treated Mfap4-/- mouse mesenteric arteries in ex vivo myography recordings. CONCLUSIONS: MFAP4-deficiency reduces the elastin/collagen ratio in the aging resistance artery without affecting arterial stiffness. In contrast, MFAP4-deficiency reduces the stiffness of resistance arteries and ameliorates Ang II-induced hypertension.

Crystal structures of human immune protein FIBCD1 suggest an extended binding site compatible with recognition of pathogen-associated carbohydrate motifs.

Fibrinogen C domain-containing protein 1 (FIBCD1) is an immune protein proposed to be involved in host recognition of chitin on the surface of pathogens. As FIBCD1 readily binds acetylated molecules, we have determined the high-resolution crystal structures of a recombinant fragment of the FIBCD1 C-terminal domain complexed with small N-acetyl-containing ligands to determine the mode of recognition. All ligands bind at the conserved N-acetyl-binding site (S1) with galactose and glucose-derived ligands rotated 180° relative to each other. One subunit of a native structure derived from protein expressed in mammalian cells binds glycosylation from a neighboring subunit, in an extended binding site. Across the various structures, the primary S1 binding pocket is occupied by N-acetyl-containing ligands or acetate, with N-acetyl, acetate, or sulfate ion in an adjacent pocket S1(2). Inhibition binding studies of N-acetylglucosamine oligomers, (GlcNAc)n, n = 1, 2, 3, 5, 11, via ELISA along with microscale thermophoresis affinity assays indicate a strong preference of FIBCD1 for longer N-acetylchitooligosaccharides. Binding studies of mutant H396A, located beyond the S1(2) site, showed no significant difference from wildtype, but K381L, within the S1(2) pocket, blocked binding to the model ligand acetylated bovine serum albumin, suggesting that S1(2) may have functional importance in ligand binding. The binding studies, alongside structural definition of diverse N-acetyl monosaccharide binding in the primary S1 pocket and of additional, adjacent binding pockets, able to accommodate both carbohydrate and sulfate functional groups, suggest a versatility in FIBCD1 to recognize chitin oligomers and other pathogen-associated carbohydrate motifs across an extended surface.

Expression of FIBCD1 by intestinal epithelial cells alleviates inflammation-driven tumorigenesis in a mouse model of colorectal cancer.

Background: Colorectal cancer (CRC) ranks as the third most prevalent cancer globally, highlighting the pressing need to address its development. Inflammation plays a crucial role in augmenting the risk of CRC and actively contributes to all stages of tumorigenesis. Consequently, targeting early inflammatory responses in the intestinal tract to restore homeostasis holds significant potential for preventing and treating CRC. Fibrinogen C domain-containing 1 (FIBCD1), a chitin-binding transmembrane protein predominantly found on human intestinal epithelial cells (IECs), has garnered attention in previous research for its ability to effectively suppress inflammatory responses and promote tissue homeostasis at mucosal barriers. Methods: In this study, we investigated the role of FIBCD1 in CRC development using transgenic mice that mimic human expression of FIBCD1 at the intestinal mucosal barrier. To model aspects of CRC, we employed the azoxymethane/dextran sodium sulfate (AOM/DSS) mouse model. Additionally, we examined the expression pattern of FIBCD1 in surgical specimens obtained from human CRC patients by immunohistochemical methods. By accessing public data repositories, we further evaluated FIBCD1 expression in colon adenocarcinoma and explored survival outcomes associated with FIBCD1 expression. Results: Here, we demonstrate that FIBCD1 substantially impacts CRC development by significantly reducing intestinal inflammation and suppressing colorectal tumorigenesis in mice. Furthermore, we identify a soluble variant of FIBCD1 that is significantly increased in feces during acute inflammation. Finally, we demonstrate increased expression of FIBCD1 by immunohistochemistry in human CRC specimens at more developed tumor stages. These results are further supported by bioinformatic analyses of publicly available repositories, indicating increased FIBCD1 expression in tumor tissues, where higher expression is associated with unfavorable prognosis. Conclusion: Collectively, these findings suggest that FIBCD1 influences early inflammatory responses in the AOM/DSS model, leading to a reduction in tumor size and burden. The increased expression of FIBCD1 in human CRC samples raises intriguing questions regarding its role in CRC, positioning it as a compelling candidate and novel molecular target for future research.

Microfibrillar-associated protein 4 as a potential marker of acute relapse in inflammatory demyelinating diseases of the central nervous system: Pathological and clinical aspects.

BACKGROUND: Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix protein not previously described in the human central nervous system (CNS). OBJECTIVES: We determined MFAP4 CNS expression and measured cerebrospinal fluid (CSF) and serum levels. METHODS: Tissue was sampled at autopsy from patients with acute multiple sclerosis (MS) (n = 3), progressive MS (n = 3), neuromyelitis optica spectrum disorder (NMOSD) (n = 2), and controls (n = 9), including 6 healthy controls (HC). MFAP4 levels were measured in 152 patients: 49 MS, 62 NMOSD, 22 myelin oligodendrocyte glycoprotein-associated disease (MOGAD), and 19 isolated optic neuritis (ION). RESULTS: MFAP4 localized to meninges and vascular/perivascular spaces, intense in the optic nerve. At sites of active inflammation, MFAP4 reactivity was reduced in NMOSD and acute MS and less in progressive MS. CSF MFAP4 levels were reduced during relapse and at the onset of diseases (mean U/mL: MS 14.3, MOGAD 9.7, and ION 14.6 relative to HC 17.9. (p = 0.013, p = 0.000, and p = 0.019, respectively). Patients with acute ON (n = 68) had reduced CSF MFAP4 (mean U/mL: 14.5, p = 0.006). CSF MFAP4 levels correlated negatively with relapse severity (rho = -0.41, p = 0.017). CONCLUSION: MFAP4 immunoreactivity was reduced at sites of active inflammation. CSF levels of MFAP4 were reduced following relapse and may reflect disease activity.

Large-Scale Protein Analysis of Experimental Retinal Artery Occlusion.

Retinal artery occlusion (RAO) is a devastating condition with no effective treatment. The management of RAO could potentially be improved through an in-depth understanding of the molecular alterations in the condition. This study combined advanced proteomic techniques and an experimental model to uncover the retinal large-scale protein profile of RAO. In 13 pigs, RAO was induced with an argon laser and confirmed by fluorescein angiography. Left eyes serving as controls received a sham laser without inducing occlusion. Retinal samples were collected after one, three, or six days and analyzed with liquid chromatography-tandem mass spectrometry. In RAO, 36 proteins were differentially regulated on day one, 86 on day three, and 557 on day six. Upregulated proteins included clusterin, vitronectin, and vimentin, with several proteins increasing over time with a maximum on day six, including clusterin, vimentin, osteopontin, annexin-A, signal transducer, and the activator of transcription 3. On day six, RAO resulted in the upregulation of proteins involved in cellular response to stress, hemostasis, innate immune response, and cytokine signaling. Downregulated proteins were involved in transmission across chemical synapses and visual phototransduction. This study identified the upregulation of multiple inflammatory proteins in RAO and the downregulation of proteins involved in visual pathways.

Serum MFAP4, a novel potential biomarker for liver cirrhosis screening, correlates with transient elastography in NAFLD patients.

Background and Aim: Non-alcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease in different countries. Liver fibrosis is considered as the most appropriate predictor of NAFLD-associated outcome. Microfibrillar-associated protein 4 (MFAP4) is a glycoprotein located in the extracellular matrix. Circulatory MFAP4 has been suggested as a noninvasive biomarker for the assessment of hepatitis C virus and alcoholic liver disease associated liver fibrosis. In this study, we aimed to investigate the association between serum MFAP4 and liver fibrosis severity in NAFLD patients. Methods: A case-control study was conducted in which NAFLD patients (n = 25) and healthy participants (n = 12) were recruited. Liver fibrosis/cirrhosis was assessed by transient elastography (TE) and biochemical parameters were collected. Serum MFAP4 was measured by sandwich ELISA based on two monoclonal anti-MFAP4 antibodies and calibrated with a standard of recombinant MFAP4. Results: Serum MFAP4 levels increased with fibrosis severity and were highly upregulated in patients with cirrhosis (F4 fibrosis stage). In addition, serum MFAP4 levels positively correlated with TE measurement and showed significant association with the severely advanced fibrotic stage in NAFLD patients, in multiple linear regression analysis following adjustment for age, gender, and body mass index. Conclusion: This study suggests the use of MFAP4 as a potential diagnostic noninvasive biomarker for cirrhosis screening in NAFLD patients.

Using thoracic ultrasound to detect interstitial lung disease in patients with rheumatoid arthritis: a protocol for the diagnostic test accuracy AURORA study.

INTRODUCTION: Pulmonary diseases are significant contributors to morbidity and mortality in patients with rheumatoid arthritis (RA). RA-associated interstitial lung disease (RA-ILD) may be prevalent in up to 30% and clinically evident in 10% of patients with RA. Feasible methods to detect concomitant ILD in RA are warranted. Our objective is to determine the diagnostic accuracy of thoracic ultrasound (TUS) for ILD in patients with RA with respiratory symptoms, by using chest high-resolution CT (HRCT) as the reference standard. Further, we aim to evaluate the diagnostic accuracy for the promising blood biomarkers surfactant protein-D and microfibrillar-associated protein 4 in the detection of ILD in this group of patients. METHODS AND ANALYSIS: By use of a standardised 14 zone protocol patients suspected of having RA-ILD will undergo TUS as index test performed by a junior resident in rheumatology (BKS), who is certified by the European Respiratory Society in performing TUS assessments. Participants form a consecutive series of up to 80 individuals in total. The anonymised TUS images will be stored and scored by the junior resident as well as two senior rheumatologists, who have received training in TUS, and a TUS-experienced pulmonologist. HRCT will be used as the gold standard for ILD diagnosis (reference standard). The two basic measures for quantifying the diagnostic test accuracy of the TUS test are the sensitivity and specificity in comparison to the HRCT. ETHICS AND DISSEMINATION: Data will be collected and stored in the Research Electronic Data Capture database. The study is approved by the Committees on Health Research Ethics and the Danish Data Protection Agency. The project is registered at clinicaltrials.gov (NCT05396469, pre-results) and data will be published in peer-reviewed journals.

Dexamethasone Intravitreal Implant Is Active at the Molecular Level Eight Weeks after Implantation in Experimental Central Retinal Vein Occlusion.

Central retinal vein occlusion (CRVO) is a visually disabling condition resulting from a thrombus in the major outflow vessel of the eye. The inflammatory response in CRVO is effectively treated with a dexamethasone (DEX) intravitreal implant. Uncovering the proteome changes following DEX implant intervention in CRVO may identify key proteins that mediate the beneficial effects of DEX. In six Göttingen minipigs, CRVO was induced in both eyes with an argon laser using a well-established experimental model. The right eyes were treated with a DEX intravitreal implant (Ozurdex, Allergan), while the left control eyes received a sham injection. Eight weeks after DEX intervention, retinal samples were collected and analyzed with tandem mass tag-based mass spectrometry. DEX implant intervention resulted in the upregulation of peptidyl-prolyl cis-trans isomerase FKBP5 (FKBP5) and ubiquilin-4. Immunohistochemistry showed expression of FKBP5 in the nuclei in all cellular layers of the retina. Cell adhesion molecule 3, tumor necrosis factor receptor superfamily member 16, and trans-1,2-dihydrobenzene-1,2-diol dehydrogenase were downregulated following DEX intervention. The upregulation of the corticosteroid-sensitive protein FKBP5 suggests that the implant remained active at the molecular level after eight weeks of treatment. Future studies may investigate if FKBP5 regulates the efficacy and duration of the DEX implant.

MFAP4-Mediated Effects in Elastic Fiber Homeostasis, Integrin Signaling and Cancer, and Its Role in Teleost Fish.

Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix (ECM) protein belonging to the fibrinogen-related domain superfamily. MFAP4 is highly expressed in elastin-rich tissues such as lung, blood vessels and skin. MFAP4 is involved in organization of the ECM, regulating proper elastic fiber assembly. On the other hand, during pathology MFAP4 actively contributes to disease development and progression due to its interactions with RGD-dependent integrin receptors. Both tissue expression and circulating MFAP4 levels are associated with various disorders, including liver fibrosis and cancer. In other experimental models, such as teleost fish, MFAP4 appears to participate in host defense as a macrophage-specific innate immune molecule. The aim of this review is to summarize the accumulating evidence that indicates the importance of MFAP4 in homeostasis as well as pathological conditions, discuss its known biological functions with special focus on elastic fiber assembly, integrin signaling and cancer, as well as describe the reported functions of non-mammalian MFAP4 in fish. Overall, our work provides a comprehensive overview on the role of MFAP4 in health and disease.

Microfibrillar-associated protein 4 in health and disease.

Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix protein belonging to the fibrinogen-related domain family. It has been localized to elastic fiber-rich regions in several tissues including the arteries, lungs, heart and skin. MFAP4 binds collagen, fibrillins and tropoelastin and contributes to the process of microfibrillar assembly and maturation of elastic fibers. MFAP4 can also bind RGD-dependent integrins, predominantly αVß3 and αVß5 through its N-terminal RGD sequence, modulating cellular behavior. Circulating MFAP4 was suggested as a robust biomarker for hepatitis C virus- and alcoholic liver disease-related liver fibrosis, cardiovascular disorders and chronic obstructive pulmonary disease. In mice, MFAP4 seems to have a widely redundant role under homeostatic conditions, as global MFAP4 deficiency results in a mild pulmonary phenotype, causing emphysema-like airspace enlargement that progresses with age. However, emerging in vivo and in vitro data suggest that MFAP4 is actively involved in the pathogenesis of remodeling-associated diseases, including fibrosis, cardiovascular disorders, aging, asthma and cancer through activation of integrin-mediated signaling as well as by modulating TGF-ß pathway, thus supporting maladaptive matrix remodeling. This review summarizes the current knowledge about MFAP4 structure and localization, its mechanisms of action in disease-induced tissue remodeling as well as its potential role as a clinical biomarker.

Proteome Analysis of Aflibercept Intervention in Experimental Central Retinal Vein Occlusion.

Aflibercept is a frequently used inhibitor of vascular endothelial growth factor (VEGF) in the treatment of macular edema following central retinal vein occlusion (CRVO). Retinal proteome changes following aflibercept intervention in CRVO remain largely unstudied. Studying proteomic changes of aflibercept intervention may generate a better understanding of mechanisms of action and uncover aspects related to the safety profile. In 10 Danish Landrace pigs, CRVO was induced in both eyes with an argon laser. Right eyes were treated with intravitreal aflibercept while left control eyes received isotonic saline water. Retinal samples were collected 15 days after induced CRVO. Proteomic analysis by tandem mass tag-based mass spectrometry identified a total of 21 proteins that were changed in content following aflibercept intervention. In retinas treated with aflibercept, high levels of aflibercept components were reached, including the VEGF receptor-1 and VEGF receptor-2 domains. Fold changes in the additional proteins ranged between 0.70 and 1.19. Aflibercept intervention resulted in a downregulation of pigment epithelium-derived factor (PEDF) (fold change = 0.84) and endoplasmin (fold change = 0.91). The changes were slight and could thereby not be confirmed with less precise immunohistochemistry and Western blotting. Our data suggest that aflibercept had a narrow mechanism of action in the CRVO model. This may be an important observation in cases when macular edema secondary to CRVO is resistant to aflibercept intervention.

A serological biomarker of type I collagen degradation is related to a more severe, high neutrophilic, obese asthma subtype.

BACKGROUND: Asthma is a heterogeneous disease; therefore, biomarkers that can assist in the identification of subtypes and direct therapy are highly desirable. Asthma is a chronic inflammatory disease that leads to changes in the extracellular matrix (ECM) by matrix metalloproteinases (MMPs) degradation causing fragments of type I collagen that is released into circulation. OBJECTIVE: Here, we asked if MMP-generated type I collagen (C1M) was associated with subtypes of asthma. METHODS: C1M was serologically assessed at baseline in the adult participants of the All Age Asthma study (ALLIANCE) (n = 233), and in The Prospective Epidemiological Risk Factor study (PERF) (n = 283). In addition, C1M was assessed in mice sensitized to ovalbumin (OVA) and challenged with OVA aerosol. C1M was evaluated in mice with and without acute neutrophilic inflammation provoked by poly(cytidylic-inosinic) acid and mice treated with CP17, a peptide inhibiting neutrophil accumulation. RESULTS: Serum C1M was significantly increased in asthmatics compared to healthy controls (p = 0.0005). We found the increased C1M levels in asthmatics were related to blood neutrophil and body mass index (BMI) in the ALLIANCE cohort, which was validated in the PERF cohort. When patients were stratified into obese (BMI > 30) asthmatics with high neutrophil levels and uncontrolled asthma, this group had a significant increase in C1M compared to normal-weight (BMI < 25) asthmatics with low neutrophil levels and controlled asthma (p = 0.0277). C1M was significantly elevated in OVA mice with acute neutrophilic inflammation compared to controls (P = 0.0002) and decreased in mice treated with an inhibitor of neutrophil infiltration (p = 0.047). CONCLUSION & CLINICAL RELEVANCE: C1M holds the potential to identify a subtype of asthma that relates to severity, obesity, and high neutrophils. These data suggest that C1M is linked to a subtype of overall inflammation, not only derived from the lung. The link between C1M and neutrophils were further validated in in vivo model. TRIAL REGISTRATION: (ALLIANCE, NCT02419274 ).

Surfactant protein D inhibits growth, alters cell surface polysaccharide exposure and immune activation potential of Aspergillus fumigatus.

Humoral immunity plays a defensive role against invading microbes. However, it has been largely overlooked with respect to Aspergillus fumigatus, an airborne fungal pathogen. Previously, we have demonstrated that surfactant protein D (SP-D), a major humoral component in human lung-alveoli, recognizes A. fumigatus conidial surface exposed melanin pigment. Through binding to melanin, SP-D opsonizes conidia, facilitates conidial phagocytosis, and induces the expression of protective pro-inflammatory cytokines in the phagocytic cells. In addition to melanin, SP-D also interacts with galactomannan (GM) and galactosaminogalactan (GAG), the cell wall polysaccharides exposed on germinating conidial surfaces. Therefore, we aimed at unravelling the biological significance of SP-D during the germination process. Here, we demonstrate that SP-D exerts direct fungistatic activity by restricting A. fumigatus hyphal growth. Conidial germination in the presence of SP-D significantly increased the exposure of cell wall polysaccharides chitin, α-1,3-glucan and GAG, and decreased ß-1,3-glucan exposure on hyphae, but that of GM was unaltered. Hyphae grown in presence of SP-D showed positive immunolabelling for SP-D. Additionally, SP-D treated hyphae induced lower levels of pro-inflammatory cytokine, but increased IL-10 (anti-inflammatory cytokine) and IL-8 (a chemokine) secretion by human peripheral blood mononuclear cells (PBMCs), compared to control hyphae. Moreover, germ tube surface modifications due to SP-D treatment resulted in an increased hyphal susceptibility to voriconazole, an antifungal drug. It appears that SP-D exerts its anti-A. fumigatus functions via a range of mechanisms including hyphal growth-restriction, hyphal surface modification, masking of hyphal surface polysaccharides and thus altering hyphal immunostimulatory properties.

Association of serum surfactant protein D and SFTPD gene variants with asthma in Danish children, adolescents, and young adults.

BACKGROUND: Surfactant Protein D (SP-D) is a pattern recognition molecule belonging to the family of collectins expressed in multiple human organ systems, including the lungs. Previous studies have shown that SP-D levels in bronchoalveolar lavage samples decrease and serum levels increase in patients suffering from asthma, possibly due to a combination of induced SP-D synthesis and decreased air-blood barrier integrity. The aims of this study were to investigate whether serum levels of SP-D and common variants in the SP-D gene were associated with asthma in adolescents and young adults. METHODS: Prospective observational study including 449 adolescents and young adults (age 11-27 years) previously diagnosed with asthma during a 2-year period from 2003 to 2005 (0-16 years). At follow-up from 2016 to 2017, 314 healthy controls with no history of asthma were recruited. Serum SP-D was analyzed on samples obtained at baseline as well as samples obtained at follow-up. SP-D genotyping was performed for rs721917, rs2243639, and rs3088308. RESULTS: No differences were found in mean levels of sSP-D and SFTPD genotype among subjects with current asthma, no current asthma, and controls. Serum SP-D and SFTPD genotype were not associated with any clinical parameters of asthma. Furthermore, baseline sSP-D was not associated with asthma at follow-up. CONCLUSION: Serum surfactant protein D and common SP-D gene variants were not associated with asthma in Danish adolescents and young adults with mild to moderate asthma. Serum surfactant protein D did not demonstrate any value as a clinical biomarker of asthma.

MFAP4 Deficiency Attenuates Angiotensin II-Induced Abdominal Aortic Aneurysm Formation Through Regulation of Macrophage Infiltration and Activity.

Objective: Abdominal aortic aneurysm (AAA) is a common age-related vascular disease characterized by progressive weakening and dilatation of the aortic wall. Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix (ECM) protein involved in the induction of vascular remodeling. This study aimed to investigate if MFAP4 facilitates the development of AAA and characterize the underlying MFAP4-mediated mechanisms. Approach and Results: Double apolipoprotein E- and Mfap4-deficient (ApoE -/- Mfap4 -/-) and control apolipoprotein E-deficient (ApoE -/-) mice were infused subcutaneously with angiotensin II (Ang II) for 28 days. Mfap4 expression was localized within the adventitial and medial layers and was upregulated after Ang II treatment. While Ang II-induced blood pressure increase was independent of Mfap4 genotype, ApoE -/- Mfap4 -/- mice exhibited significantly lower AAA incidence and reduced maximal aortic diameter compared to ApoE -/- littermates. The ApoE -/- Mfap4 -/- AAAs were further characterized by reduced macrophage infiltration, matrix metalloproteinase (MMP)-2 and MMP-9 activity, proliferative activity, collagen content, and elastic membrane disruption. MFAP4 deficiency also attenuated activation of integrin- and TGF-ß-related signaling within the adventitial layer of AAA tissues. Finally, MFAP4 stimulation promoted human monocyte migration and significantly upregulated MMP-9 activity in macrophage-like THP-1 cells. Conclusion: This study demonstrates that MFAP4 induces macrophage-rich inflammation, MMP activity, and maladaptive remodeling of the ECM within the vessel wall, leading to an acceleration of AAA development and progression. Collectively, our findings suggest that MFAP4 is an essential aggravator of AAA pathology that acts through regulation of monocyte influx and MMP production.

Corrigendum to 'Level of MFAP4 in ascites independently predicts 1-year transplant-free survival in patients with cirrhosis' [JHEP Reports 3 (2021) 100287].

[This corrects the article DOI: 10.1016/j.jhepr.2021.100287.].

No effect of deleted in malignant brain tumors 1 deficiency on chemotherapy induced murine intestinal mucositis.

Mucositis is a serious adverse effect of chemotherapeutic treatment. During intestinal mucositis, the mucosal barrier is compromised, increasing the risk of severe infections. Mucositis necessitates dose reduction or pauses in treatment, which affect the outcome of the treatment. Deleted in malignant brain tumors 1 (DMBT1) is a secreted scavenger protein with effects on innate immunity and epithelial regeneration. We have previously shown that jejunal DMBT1 expression is increased in piglets during chemotherapeutic treatment. We hypothesized that DMBT1 ameliorates doxorubicin-induced mucositis. Individually-caged Dmbt1+/+ (WT) and Dmbt1-/- (KO) female mouse littermates received intraperitoneal injections of either doxorubicin or saline. They were euthanized after three (D3) or seven days (D7). Weight loss was monitored every day, and serum citrulline levels were measured at termination. Intestinal tissue was analyzed for the expression of DMBT1 and proinflammatory cytokines (IL-1ß, IL-6, and TNF). Specimens from the small intestines and colon were scored for inflammation and epithelial and mucosal architecture changes. We detected no effect of DMBT1 on weight loss, serum citrulline levels, expression of proinflammatory cytokines, or histologic damage. We detected a significant increase in crypt depth in WT mice compared to that in KO mice on D3. In conclusion, DMBT1 does not affect doxorubicin-induced mucositis in mice.

Level of MFAP4 in ascites independently predicts 1-year transplant-free survival in patients with cirrhosis.

BACKGROUND & AIMS: Prognostic models of cirrhosis underestimate disease severity for patients with cirrhosis and ascites. Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix protein linked to hepatic neoangiogenesis and fibrogenesis. We investigated ascites MFAP4 as a predictor of transplant-free survival in patients with cirrhosis and ascites. METHODS: A dual-centre observational study of patients with cirrhosis and ascites recruited consecutively in relation to a paracentesis was carried out. Patients were followed up for 1 year, until death or liver transplantation (LTx). Ascites MFAP4 was tested with the model for end-stage liver disease (MELD-Na), CLIF Consortium Acute Decompensation (CLIF-C AD), and Child-Pugh score in Cox regression models. RESULTS: Ninety-three patients requiring paracentesis were included. Median ascites MFAP4 was 29.7 U/L [22.3-41.3], and MELD-Na was 19 [16-23]. A low MELD-Na score (<20) was observed in 49 patients (53%). During follow-up, 20 patients died (22%), and 6 received LTx (6%). High ascites MFAP4 (>29.7 U/L) was associated with 1-year transplant-free survival (p = 0.002). In Cox regression, ascites MFAP4 and MELD-Na independently predicted 1-year transplant-free survival (hazard ratio [HR] = 0.97, p = 0.03, and HR = 1.08, p = 0.01, respectively). Ascites MFAP4 and CLIF-C AD also predicted survival independently (HR = 0.96, p = 0.02, and HR = 1.05, p = 0.03, respectively), whereas only ascites MFAP4 did, controlling for the Child-Pugh score (HR = 0.97, p = 0.03, and HR = 1.18, p = 0.16, respectively). For patients with MELD-Na <20, ascites MFAP4 but not ascites protein predicted 1-year transplant-free survival (HR 0.91, p = 0.02, and HR = 0.94, p = 0.17, respectively). CONCLUSIONS: Ascites MFAP4 predicts 1-year transplant-free survival in patients with cirrhosis and ascites. In patients with low MELD-Na scores, ascites MFAP4, but not total ascites protein, significantly predicted 1-year transplant-free survival. LAY SUMMARY: Patients with cirrhosis who have fluid in the abdomen, ascites, are at an increased risk of death and in need for liver transplantation. Our study identified patients with ascites and a poor prognosis by measuring microfibrillar associated protein 4 (MFAP4), a protein present in the abdominal fluid. Patients with low levels of the MFAP4 protein are at particularly increased risk of death or liver transplantation, suggesting that clinical care should be intensified in this group of patients.