Although uteroglobin is known to have an immunomodulatory property and prevents the deposition of immune-complexes on the glomeruli of mice, the therapeutic potential of uteroglobin is uncertain in glomerulonephritis. To test the hypothesis that uteroglobin can prevent glomerulonephritis, we have studied the effects of recombinant uteroglobin on the development of experimental crescentic glomerulonephritis that is induced by anti-glomerular basement membrane (anti-GBM) antibodies.

Serum amyloid A (SAA) production is increased by inflamed arthritic synovial tissue, where it acts as a cytokine/chemoattractant for inflammatory and immune cells and as an inducer of matrix degrading enzymes. SAA has been shown to bind lipoxin A4 receptor, a member of the formyl-peptide related 2 G-protein coupled receptor family (ALX) and elicit proinflammatory activities in human primary fibroblast-like synoviocytes (FLS). We report on the identification of uteroglobin, a small globular protein with potent anti-inflammatory activities, as a possible ligand of ALX. Uteroglobin-specific association with ALX was demonstrated by an enzyme immunoassay experiment employing a cell line engineered to express the human ALX receptor. Uteroglobin's interaction with ALX resulted in the inhibition of SAA responses, such as attenuation of phospholipase A2 activation and cellular chemotaxis. In FLS, uteroglobin showed an antagonism against SAA-induced interleukin-8 release and decreased cell migration. These novel roles described for uteroglobin via ALX may help elucidate genetic and clinical observations indicating that a polymorphism in the uteroglobin promoter is linked to disease outcome, specifically prediction of bone erosion in patients with rheumatoid arthritis or severity of IgA glomerulonephritis and sarcoidosis.

Uteroglobin (UG) is an antiinflammatory protein secreted by the epithelial lining of all organs communicating with the external environment. We reported previously that UG-knockout mice manifest exaggerated inflammatory response to allergen, characterized by increased eotaxin and Th2 cytokine gene expression, and eosinophil infiltration in the lungs. In this study, we uncovered that the airway epithelia of these mice also express high levels of cyclooxygenase (COX)-2, a key enzyme for the production of proinflammatory lipid mediators, and the bronchoalveolar lavage fluid (BALF) contain elevated levels of prostaglandin D2. These effects are abrogated by recombinant UG treatment. Although it has been reported that prostaglandin D2 mediates allergic inflammation via its receptor, DP, neither the molecular mechanism(s) of DP signaling nor the mechanism by which UG suppresses DP-mediated inflammatory response are clearly understood. Here we report that DP signaling is mediated via p38 mitogen-activated protein kinase, p44/42 mitogen-activated protein kinase, and protein kinase C pathways in a cell type-specific manner leading to nuclear factor-kappaB activation stimulating COX-2 gene expression. Further, we found that recombinant UG blocks DP-mediated nuclear factor-kappaB activation and suppresses COX-2 gene expression. We propose that UG is an essential component of a novel innate homeostatic mechanism in the mammalian airways to repress allergen-induced inflammatory responses.

To get further insights on the estrogen regulation of the uteroglobin (UG) gene, the 5'-flanking region of the UG gene from the brown hare (Lepus capensis) (Lc) was cloned and compared with those from two phylogenetically related species: the rabbit (Orictolagus cuniculus) (Oc) and the volcano rabbit (Romerolagus diazi) (Rd). The Lc-UG gene is very similar to those from rabbits (94%) and volcano rabbits (95%), and shares a number of genetic elements, including an estrogen response element (ERE). The estrogen-regulated transcription of a series of progressive 5'-deletion mutants of the Lc-UG gene, identified a functional ERE in the promoter region exhibiting the same orientation and relative position than that previously described in rabbits. The Lc-ERE is identical to the Oc-ERE, but different from both the Rd-ERE and the consensus ERE (c-ERE) by one nucleotide. We also detected important species-specific differences in the estrogen-regulated transcription of the UG gene. A luciferase reporter driven by 333 base pairs (bp) of the Lc-UG promoter elicited a higher response to estradiol than its related counterparts when expressed in estrogen-sensitive MCF-7 cells. Several ERE-like motifs which failed to act as functional EREs were also identified; one of them exhibited two mismatches in its palindromic sequence, a characteristic exhibited in many other natural occurring EREs, including the Rd-ERE.

Uteroglobin-Related Protein 1 (UGRP1) is a secretoglobulin protein which has been suggested to play a role in lung inflammation and allergic diseases. UGRP1 has also been shown to be an important pneumoprotein, with diagnostic potential as a biomarker of lung damage. Previous genetic studies evaluating the association between variations on UGRP1 and allergic phenotypes have yielded mixed results. The aim of this present study was to identify genetic polymorphisms in UGRP1 and investigate if they were associated with asthma and allergic rhinitis in the Singapore Chinese population.

Escherichia coli is a robust, economic and rapid expression system for the production of recombinant therapeutic proteins. However, the expression in bacterial systems of complex molecules such as antibodies and fusion proteins is still affected by several drawbacks. We have previously described a procedure based on uteroglobin (UG) for the engineering of very soluble and stable polyvalent and polyspecific fusion proteins in mammalian cells (Ventura et al. 2009. J. Biol. Chem. 284∶26646-26654.) Here, we applied the UG platform to achieve the expression in E. coli of a bivalent human recombinant antibody (L19) toward the oncofetal fibronectin (B-FN), a pan-tumor target. Purified bacterial L19-UG was highly soluble, stable, and, in all molecules, the L19 moiety maintained its immunoreactivity. About 50-70% of the molecules were covalent homodimer, however after refolding with the redox couple reduced-glutathione/oxidized-glutathione (GSH/GSSG), 100% of molecules were covalent dimers. Mass spectrometry studies showed that the proteins produced by E. coli and mammalian cells have an identical molecular mass and that both proteins are not glycosylated. L19-UG from bacteria can be freeze-dried without any loss of protein and immunoreactivity. In vivo, in tumor-bearing mice, radio-iodinated L19-UG selectively accumulated in neoplastic tissues showing the same performance of L19-UG from mammalian cells. The UG-platform may represent a general procedure for production of various biological therapeutics in E. coli.

The pathophysiological changes occurring in the trabecular meshwork in primary open angle glaucoma are poorly understood, but are thought to include increased extracellular matrix deposition, trabecular meshwork cell apoptosis, inflammation, trabecular meshwork calcification and altered protein composition of the aqueous humor. Although many proteins are present in aqueous humor, relatively few have been studied extensively, and their potential roles in primary open angle glaucoma are unknown.

Sepsis is a serious clinical condition which can cause life-threatening organ dysfunction, and has limited therapeutic options. The paradigm of limiting excessive inflammation and promoting anti-inflammatory responses is a simplified concept. Yet, the absence of intrinsic anti-inflammatory signaling at the early stage of an infection can lead to an exaggerated activation of immune cells, including monocytes and macrophages. There is emerging evidence that endogenous molecules control those mechanisms. Here we aimed to identify and describe the dynamic changes in monocyte and macrophage subsets and lung damage in CL57BL/6N mice undergoing blunt chest trauma with subsequent cecal ligation and puncture. We showed that early an increase in systemic and activated Ly6C+CD11b+CD45+Ly6G- monocytes was paralleled by their increased emigration into lungs. The ratio of pro-inflammatory Ly6ChighCD11b+CD45+Ly6G- to patrolling Ly6ClowCD11b+CD45+Ly6G- monocytes significantly increased in blood, lungs and bronchoalveolar lavage fluid (BALF) suggesting an early transition to inflammatory phenotypes during early sepsis development. Similar to monocytes, the level of pro-inflammatory Ly6ChighCD45+F4/80+ macrophages increased in lungs and BALF, while tissue repairing Ly6ClowCD45+F4/80+ macrophages declined in BALF. Levels of inflammatory mediators TNF-α and MCP-1 in blood and RAGE in lungs and BALF were elevated, and besides their boosting of inflammation via the recruitment of cells, they may promote monocyte and macrophage polarization, respectively, toward the pro-inflammatory phenotype. Neutralization of uteroglobin increased pro-inflammatory cytokine levels, activation of inflammatory phenotypes and their recruitment to lungs; concurrent with increased pulmonary damage in septic mice. In in vitro experiments, the influence of uteroglobin on monocyte functions including migratory behavior, TGF-β1 expression, cytotoxicity and viability were proven. These results highlight an important role of endogenous uteroglobin as intrinsic anti-inflammatory signal upon sepsis-induced early lung injury, which modules the early monocyte/macrophages driven inflammation.

Ligand-targeted approaches have proven successful in improving the therapeutic index of a number of drugs. We hypothesized that the specific targeting of TNF-alpha antagonists to inflamed tissues could increase drug efficacy and reduce side effects.

Antiflammin-1 (AF-1), a derivative of uteroglobin (UG), is a synthetic nonapeptide with diverse biological functions. In the present study, we investigated whether AF-1 has a protective effect against bleomycin-induced pulmonary fibrosis.

Cigarette smoking is the main risk factor for COPD (Chronic Obstructive Pulmonary Disease), yet only a subset of smokers develops COPD. Family members of patients with severe early-onset COPD have an increased risk to develop COPD and are therefore defined as "susceptible individuals". Here we perform unbiased analyses of proteomic profiles to assess how "susceptible individuals" differ from age-matched "non-susceptible individuals" in response to cigarette smoking. Epithelial lining fluid (ELF) was collected at baseline and 24 hours after smoking 3 cigarettes in young individuals susceptible or non-susceptible to develop COPD and older subjects with established COPD. Controls at baseline were older healthy smoking and non-smoking individuals. Five samples per group were pooled and analysed by stable isotope labelling (iTRAQ) in duplicate. Six proteins were selected and validated by ELISA or immunohistochemistry. After smoking, 23 proteins increased or decreased in young susceptible individuals, 7 in young non-susceptible individuals, and 13 in COPD in the first experiment; 23 proteins increased or decreased in young susceptible individuals, 32 in young non-susceptible individuals, and 11 in COPD in the second experiment. SerpinB3 and Uteroglobin decreased after acute smoke exposure in young non-susceptible individuals exclusively, whereas Peroxiredoxin I, S100A9, S100A8, ALDH3A1 (Aldehyde dehydrogenase 3A1) decreased both in young susceptible and non-susceptible individuals, changes being significantly different between groups for Uteroglobin with iTRAQ and for Serpin B3 with iTRAQ and ELISA measures. Peroxiredoxin I, SerpinB3 and ALDH3A1 increased in COPD patients after smoking. We conclude that smoking induces a differential protein response in ELF of susceptible and non-susceptible young individuals, which differs from patients with established COPD. This is the first study applying unbiased proteomic profiling to unravel the underlying mechanisms that induce COPD. Our data suggest that SerpinB3 and Uteroglobin could be interesting proteins in understanding the processes leading to COPD.

The classical manifestation of COVID-19 is pulmonary infection. After host cell entry via human angiotensin-converting enzyme II (hACE2), the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus can infect pulmonary epithelial cells, especially the AT2 (alveolar type II) cells that are crucial for maintaining normal lung function. However, previous hACE2 transgenic models have failed to specifically and efficiently target the cell types that express hACE2 in humans, especially AT2 cells. In this study, we report an inducible, transgenic hACE2 mouse line and showcase three examples for specifically expressing hACE2 in three different lung epithelial cells, including AT2 cells, club cells, and ciliated cells. Moreover, all these mice models develop severe pneumonia after SARS-CoV-2 infection. This study demonstrates that the hACE2 model can be used to precisely study any cell type of interest with regard to COVID-19-related pathologies.

Blunt thoracic trauma (TxT) deteriorates clinical post-injury outcomes. Ongoing inflammatory changes promote the development of post-traumatic complications, frequently causing Acute Lung Injury (ALI). Club Cell Protein (CC)16, a pulmonary anti-inflammatory protein, correlates with lung damage following TxT. Whether CC16-neutralization influences the inflammatory course during ALI is elusive. Ninety-six male CL57BL/6N mice underwent a double hit model of TxT and cecal ligation puncture (CLP, 24 h post-TxT). Shams underwent surgical procedures. CC16 was neutralized by the intratracheal application of an anti-CC16-antibody, either after TxT (early) or following CLP (late). Euthanasia was performed at 6 or 24 h post-CLP. Systemic and pulmonary levels of IL-6, IL-1β, and CXCL5 were determined, the neutrophils were quantified in the bronchoalveolar lavage fluid, and histomorphological lung damage was assessed. ALI induced a significant systemic IL-6 increase among all groups, while the local inflammatory response was most prominent after 24 h in the double-hit groups as compared to the shams. Significantly increased neutrophilic infiltration upon double hit was paralleled with the enhanced lung damage in all groups as compared to the sham, after 6 and 24 h. Neutralization of CC16 did not change the systemic inflammation. However, early CC16-neutralization increased the neutrophilic infiltration and lung injury at 6 h post-CLP, while 24 h later, the lung injury was reduced. Late CC16-neutralization increased neutrophilic infiltration, 24 h post-CLP, and was concurrent with an enhanced lung injury. The data confirmed the anti-inflammatory potential of endogenous CC16 in the murine double-hit model of ALI.

Madhuca longifolia, a tropical tree used as medicine and food, is known to have a beneficial effect against stomach gastric toxicity. Madhuca longifolia is used in treating cough, skin disease and nerve disorders. Diclofenac, a non-steroidal anti-inflammatory drug (NSAID), with overdosage and prolonged use, is known to cause gastric toxicity. Silymarin (SLY), a polyphenolic antioxidant flavonoid, is a derivative of Silybum marianum extracted from milk thistle seeds and fruits, has been widely used in the treatment of gastric ulcer. SLY was used as the standard drug to compare the effects with the Madhuca longifolia aqueous leaf extract treatment. The aim of the current study is to understand the effect of Madhuca longifolia aq. leaf extract on rat stomach and intestine against diclofenac-administered toxicity. Rats (n = 30) were divided into Group I normal control, Group II treated with diclofenac, Group III treated with diclofenac and Madhuca longifolia leaf extract, Group IV treated with diclofenac and silymarin, and Group V was treated with Madhuca longifolia leaf extract alone. After the study duration, rats were euthanized and tissue samples were analyzed for antioxidant, cytokine, protein expression levels and histopathological changes. Diclofenac treated rats had significant (p < 0.05) changes in levels of antioxidants, cytokines, protein expression and pathological changes as compared to rats treated with Madhuca longifolia. This study demonstrated that Madhuca longifolia leaf extract had gastroprotective activity in rats treated with diclofenac.

The presented comprehensive review of current knowledge about genetic factors predisposing to Graves' disease (GD) put emphasis on functional significance of observed associations. In particular, we discuss recent efforts aimed at refining diseases associations found within the HLA complex and implicating HLA class I as well as HLA-DPB1 loci. We summarize data regarding non-HLA genes such as PTPN22, CTLA4, CD40, TSHR and TG which have been extensively studied in respect to their role in GD. We review recent findings implicating variants of FCRL3 (gene for FC receptor-like-3 protein), SCGB3A2 (gene for secretory uteroglobin-related protein 1- UGRP1) as well as other unverified possible candidate genes for GD selected through their documented association with type 1 diabetes mellitus: Tenr-IL2-IL21, CAPSL (encoding calcyphosine-like protein), IFIH1(gene for interferon-induced helicase C domain 1), AFF3, CD226 and PTPN2. We also review reports on association of skewed X chromosome inactivation and fetal microchimerism with GD. Finally we discuss issues of genotype-phenotype correlations in GD.

We present the first study of the 3D kinetics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and the early host response in a large lung volume using a combination of tissue imaging and transcriptomics. This approach allowed us to make a number of important findings: Spatially restricted antiviral response is shown, including the formation of monocytic macrophage clusters and upregulation of the major histocompatibility complex II in infected epithelial cells. The monocyte-derived macrophages are linked to SARS-CoV-2 clearance, and the appearance of these cells is associated with post-infection endothelial damage; thus, we shed light on the role of these cells in infected tissue. An early onset of tissue repair occurring simultaneously with inflammatory and necrotizing processes provides the basis for longer-term alterations in the lungs.

The COVID-19 pandemic continues to be a health crisis with major unmet medical needs. The early responses from airway epithelial cells, the first target of the virus regulating the progression toward severe disease, are not fully understood. Primary human air-liquid interface cultures representing the broncho-alveolar epithelia were used to study the kinetics and dynamics of SARS-CoV-2 variants infection. The infection measured by nucleoprotein expression, was a late event appearing between day 4-6 post infection for Wuhan-like virus. Other variants demonstrated increasingly accelerated timelines of infection. All variants triggered similar transcriptional signatures, an "early" inflammatory/immune signature preceding a "late" type I/III IFN, but differences in the quality and kinetics were found, consistent with the timing of nucleoprotein expression. Response to virus was spatially organized: CSF3 expression in basal cells and CCL20 in apical cells. Thus, SARS-CoV-2 virus triggers specific responses modulated over time to engage different arms of immune response.

The aging lungs are vulnerable to chronic pulmonary diseases; however, the underlying mechanisms are not well understood. In this study, we compared the aging lungs of 20-24-month-old mice with the young of 10-16-week-old mice, and found that aging airway epithelial cells significantly upregulated the expression of uteroglobin-related protein 1 (UGRP1), which was responsible for the higher levels of CCL6 in the aging lungs. Alveolar macrophages (AMs) changed intrinsically with aging, exhibiting a decrease in cell number and altered gene expression. Using terminal differentiation trajectories, a population of MARCO+ AMs with the ability to produce CCL6 was identified in the aging lungs. Upregulated UGRP1was demonstrated to modulate CCL6 production of AMs in the UGRP1-MARCO pair in vivo and in vitro. Furthermore, MARCO+ AMs aggravated bleomycin-induced pulmonary fibrosis in a CCL6-dependent manner in the aged mice, and blocking MARCO or neutralizing CCL6 significantly inhibited pulmonary fibrosis, similar to the depletion of AMs. The age-related upregulation of UGRP1 and MARCO+ AMs, involved in the progression of lung fibrosis, was also observed in human lung tissues. Thus, UGRP1 modulated MARCO+ AMs regarding the age-related lung fibrosis in a CCL6-dependent manner, which is key to establishing optimal targeting for the aging population.

Inflammation serves a key role in chronic obstructive pulmonary disease (COPD). However, changes in the immune profiles of patients with COPD remain unclear. The present prospective observational study aimed to determine the expression profiles of immune cells and inflammatory factors of patients with COPD and healthy controls, and to analyze the relationship between immune profiles and smoking history. A total of 140 subjects were enrolled in the present study between September 2018 and April 2019 at West China Hospital of Sichuan University (Chengdu, China). These included 87 patients with stable COPD and 24 patients with acute exacerbated COPD, as well as 29 healthy controls. Baseline characteristics were recorded during the screening period, and levels of immune cells were examined using flow cytometry. In addition, levels of inflammatory factors were measured using ELISAs. The results revealed increased expression of the CD64+/CD14+ mononuclear phagocyte system (MPS) and CD16+CD66+ neutrophils, and decreased expression of CD3+CD4+ T cells and CD3+ CD8+ T cells (all P<0.05) in the peripheral blood of patients with COPD and smokers relative to non‑smoking controls. In addition, significant differences were observed in protein levels of IL‑6, IL‑1β, TNF‑α, TGF‑α, IFN‑γ, IL‑8, IL‑17A and CRP among the three groups (all P<0.05). Furthermore, the IL‑17A, TNF and NF‑κB signaling pathways were found to serve a key role in the inflammatory network of COPD. Pearson's correlation analysis revealed a positive correlation between CD3+T lymphocyte counts and pulmonary function, and a negative correlation with smoking and exacerbations. Furthermore, neutrophils and MPS were negatively associated with pulmonary function, and IL‑8 was positively associated with cough. There was also a negative association between CRP and IL‑17A with pulmonary function but a positive correlation with symptoms and exacerbation. Club cell secretory protein was also negatively associated with emphysema parameters. In conclusion, the present findings revealed significant differences in profiles of immune factors among patients with COPD, smokers and non‑smoking controls and their association with clinical characteristics. The clinical trial registration number of the present study is: ChiCTR1800015700 (registered with http://www.chictr.org.cn/index.aspx, 2018/04/16).

Existing understanding of molecular composition of sputum and its role in tuberculosis patients is variously limited to its diagnostic potential. We sought to identify infection induced sputum proteome alteration in active/non tuberculosis patients (A/NTB) and their role in altered lung patho-physiology. Out of the study population (n = 118), sputum proteins isolated from discovery set samples (n = 20) was used for an 8-plex isobaric tag for relative and absolute concentration analysis. A minimum set of protein with at least log2(ATB/NTB) >±1.0 in ATB was selected as biosignature and validated in 32 samples. Predictive accuracy was calculated from area under the receiver operating characteristic curve (AUC of ROC) using a confirmatory set (n = 50) by Western blot analysis. Mass spectrometry analysis identified a set of 192 sputum proteins, out of which a signature of β-integrin, vitamin D binding protein:DBP, uteroglobin, profilin and cathelicidin antimicrobial peptide was sufficient to differentiate ATB from NTB. AUC of ROC of the biosignature was calculated to 0.75. A shift in DBP-antimicrobial peptide (AMP) axis in the lungs of tuberculosis patients is observed. The identified sputum protein signature is a promising panel to differentiate ATB from NTB groups and suggest a deregulated DBP-AMP axis in lungs of tuberculosis patients.