The novel vaccines targeting interleukin-1 receptor type I.

OBJECTIVE: There is mounting evidence indicating that atherosclerosis represents a persistent inflammatory process, characterized by the presence of inflammation at various stages of the disease. Interleukin-1 (IL-1) precisely triggers inflammatory signaling pathways by binding to interleukin-1 receptor type I (IL-1R1). Inhibition of this signaling pathway contributes to the prevention of atherosclerosis and myocardial infarction. The objective of this research is to develop therapeutic vaccines targeting IL-1R1 as a preventive measure against atherosclerosis and myocardial infarction. METHODS: ILRQß-007 and ILRQß-008 vaccines were screened, prepared and then used to immunize high-fat-diet fed ApoE-/- mice and C57BL/6J mice following myocardial infarction. Progression of atherosclerosis in ApoE-/- mice was assessed primarily by oil-red staining of the entire aorta and aortic root, as well as by detecting the extent of macrophage infiltration. The post-infarction cardiac function in C57BL/6J mice were evaluated using cardiac ultrasound and histological staining. RESULTS: ILRQß-007 and ILRQß-008 vaccines stimulated animals to produce high titers of antibodies that effectively inhibited the binding of interleukin-1ß and interleukin-1α to IL-1R1. Both vaccines effectively reduced atherosclerotic plaque area, promoted plaque stabilization, decreased macrophage infiltration in plaques and influenced macrophage polarization, as well as decreasing levels of inflammatory factors in the aorta, serum, and ependymal fat in ApoE-/- mice. Furthermore, these vaccines dramatically improved cardiac function and macrophage infiltration in C57BL/6J mice following myocardial infarction. Notably, no significant immune-mediated damage was observed in immunized animals. CONCLUSION: The vaccines targeting the IL-1R1 would be a novel and promising treatment for the atherosclerosis and myocardial infarction.

Epithelial IFNγ signalling and compartmentalized antigen presentation orchestrate gut immunity.

All nucleated cells express major histocompatibility complex I and interferon-γ (IFNγ) receptor1, but an epithelial cell-specific function of IFNγ signalling or antigen presentation by means of major histocompatibility complex I has not been explored. We show here that on sensing IFNγ, colonic epithelial cells productively present pathogen and self-derived antigens to cognate intra-epithelial T cells, which are critically located at the epithelial barrier. Antigen presentation by the epithelial cells confers extracellular ATPase expression in cognate intra-epithelial T cells, which limits the accumulation of extracellular adenosine triphosphate and consequent activation of the NLRP3 inflammasome in tissue macrophages. By contrast, antigen presentation by the tissue macrophages alongside inflammasome-associated interleukin-1α and interleukin-1ß production promotes a pathogenic transformation of CD4+ T cells into granulocyte-macrophage colony-stimulating-factor (GM-CSF)-producing T cells in vivo, which promotes colitis and colorectal cancer. Taken together, our study unravels critical checkpoints requiring IFNγ sensing and antigen presentation by epithelial cells that control the development of pathogenic CD4+ T cell responses in vivo.

Immune suppressive function of IL-1α release in the tumor microenvironment regulated by calpain 1.

Interleukin-1α (IL-1α) plays an important role in inflammation and hematopoiesis. Many tumors have increased IL-1α expression. However, the immune regulatory role of secreted IL-1α in tumor development and whether it can be targeted for cancer therapy are still unclear. Here, we found that tumoral-secreted IL-1α significantly promoted hepatocellular carcinoma (HCC) development in vivo. Tumoral-released IL-1α were found to inhibit T and NK cell activation, and the killing capacity of CD8+ T cells. Moreover, MDSCs were dramatically increased by tumoral-released IL-1α in both spleens and tumors. Indeed, higher tumoral IL-1α expression is associated with increased tumoral infiltration of MDSCs in HCC patients. Further studies showed that tumoral-released IL-1α promoted MDSC recruitment to the tumor microenvironment through a CXCR2-dependent mechanism. Depletion of MDSCs could diminish the tumor-promoting effect of tumoral-released IL-1α. On the contrary, systemic administration of recombinant IL-1α protein significantly inhibited tumor development by activating T cells. In fact, IL-1α protein could promote T cell activation and enhance the cytotoxicity of CD8+ T cells in vitro. Thus, our study demonstrated that tumoral-released IL-1α promoted tumor development through recruiting MDSCs to inhibit T cell activation, while systemic IL-1α directly promoted anti-tumor T cell responses. We further identified calpain 1 as the major intracellular protease mediating tumoral IL-1α secretion. Calpain 1 KO tumors had diminished IL-1α release and reduced tumor development. Thus, our findings provide new insights into the functions of secreted IL-1α in tumor immunity and its implications for immunotherapy.

Bioinformatics and in-silico findings reveal medical features and pharmacological targets of biochanin A against colorectal cancer and COVID-19.

Severe mortality due to the COVID-19 pandemic resulted from the lack of effective treatment. Although COVID-19 vaccines are available, their side effects have become a challenge for clinical use in patients with chronic diseases, especially cancer patients. In the current report, we applied network pharmacology and systematic bioinformatics to explore the use of biochanin A in patients with colorectal cancer (CRC) and COVID-19 infection. Using the network pharmacology approach, we identified two clusters of genes involved in immune response (IL1A, IL2, and IL6R) and cell proliferation (CCND1, PPARG, and EGFR) mediated by biochanin A in CRC/COVID-19 condition. The functional analysis of these two gene clusters further illustrated the effects of biochanin A on interleukin-6 production and cytokine-cytokine receptor interaction in CRC/COVID-19 pathology. In addition, pathway analysis demonstrated the control of PI3K-Akt and JAK-STAT signaling pathways by biochanin A in the treatment of CRC/COVID-19. The findings of this study provide a therapeutic option for combination therapy against COVID-19 infection in CRC patients.

VGLL3 activates inflammatory responses by inducing interleukin-1α secretion.

Vestigial-like family member 3 (VGLL3), a member of the vestigial-like family, is a cofactor of the TEA-domain-containing transcription factor (TEAD). Although elevation in VGLL3 expression is associated with inflammatory diseases, such as inflammatory sarcomas and autoimmune diseases, the molecular mechanisms underlying VGLL3-mediated inflammation remain largely unknown. In this study, we analyzed the relationship between elevated VGLL3 expression and the levels of NF-κB, a transcription factor that plays a pivotal role in inflammation. NF-κB was found to be activated in a cell line stably expressing VGLL3. Mechanistically, VGLL3 was shown to promote the expression and secretion of the potent NF-κB-activating cytokine interleukin (IL)-1α, probably through its association with TEADs. As VGLL3 is a target of transforming growth factor ß (TGF-ß) signaling, we analyzed IL-1α induction upon TGF-ß stimulation. TGF-ß stimulation was observed to induce IL-1α secretion and NF-κB activation, and VGLL3 was associated with this phenomenon. The TGF-ß transcription factors Smad3 and Smad4 were shown to be necessary for inducing VGLL3 and IL-1α expression. Lastly, we found that VGLL3-dependent IL-1α secretion is involved in constitutive NF-κB activation in highly malignant breast cancer cells. Collectively, the findings suggested that VGLL3 expression and TGF-ß stimulation activate the inflammatory response by inducing IL-1α secretion.

Tumor Necrosis Factor Alpha-Induced Interleukin-1 Alpha Synthesis and Cell Death Is Increased in Mouse Epithelial Cells Infected With Chlamydia muridarum.

Chlamydia trachomatis-genital infection in women can be modeled in mice using Chlamydia muridarum. Using this model, it has been shown that the cytokines tumor necrosis factor (TNF)α and interleukin (IL)-1α lead to irreversible tissue damage in the oviducts. In this study, we investigated the contribution of TNFα on IL-1α synthesis in infected epithelial cells. We show that C muridarum infection enhanced TNFα-induced IL-1α expression and release in a mouse epithelial cell line. In addition to IL-1α, several TNFα-induced inflammatory genes were also highly induced, and infection enhanced TNF-induced cell death. In the mouse model of genital infection, oviducts from mice lacking the TNFα receptor displayed minimal staining for IL-1α compared with wild-type oviducts. Our results suggest TNFα and IL-1α enhance each other's downstream effects resulting in a hyperinflammatory response to chlamydial infection. We propose that biologics targeting TNF-induced IL-1α synthesis could be used to mitigate tissue damage during chlamydial infection.

Amyloid beta acts synergistically as a pro-inflammatory cytokine.

The amyloid beta (Aß) peptide is believed to play a central role in Alzheimer's disease (AD), the most common age-related neurodegenerative disorder. However, the natural, evolutionarily selected functions of Aß are incompletely understood. Here, we report that nanomolar concentrations of Aß act synergistically with known cytokines to promote pro-inflammatory activation in primary human astrocytes (a cell type increasingly implicated in brain aging and AD). Using transcriptomics (RNA-seq), we show that Aß can directly substitute for the complement component C1q in a cytokine cocktail previously shown to induce astrocyte immune activation. Furthermore, we show that astrocytes synergistically activated by Aß have a transcriptional signature similar to neurotoxic "A1" astrocytes known to accumulate with age and in AD. Interestingly, we find that this biological action of Aß at low concentrations is distinct from the transcriptome changes induced by the high/supraphysiological doses of Aß often used in in vitro studies. Collectively, our results suggest an important, cytokine-like function for Aß and a novel mechanism by which it may directly contribute to the neuroinflammation associated with brain aging and AD.

Evidence for the involvement of interleukin-1α during development of experimental cytomegalovirus retinitis in immunosuppressed mice.

Interleukin-1α (IL-1α) is an alarmin involved in the recruitment of macrophages and neutrophils during tissue inflammation. IL-1α can undergo cleavage by proteases, such as calpain-1, that enhances IL-1α binding to its receptor, although proteolytic cleavage is not necessary for biological activity. Macrophages and neutrophils are involved in the retinal inflammation associated with development of AIDS-related human cytomegalovirus (HCMV) retinitis. We therefore performed studies to test the hypothesis that IL-1α gene expression is stimulated intraocularly during retinitis development using two mouse models of murine cytomegalovirus (MCMV) retinitis that differ in method of immunosuppression, one by retrovirus-induced immunosuppression (MAIDS) and the other by corticosteroid-induced immunosuppression. MCMV-infected eyes of groups of retinitis-susceptible mice with MAIDS of 10 weeks duration (MAIDS-10 mice) and retinitis-susceptible corticosteroid-treated mice showed significant stimulation of IL-1α mRNA. Western blot analysis confirmed IL-1α protein production within the MCMV-infected eyes of MAIDS-10 mice. Whereas significant intraocular calpain-1 mRNA and protein production were also observed within MCMV-infected eyes of MAIDS-10 mice, the MCMV-infected eyes of retinitis-susceptible corticosteroid-treated mice showed a pattern of mRNA synthesis equivalent to that found within the MCMV-infected eyes of healthy mice that fail to develop retinitis. Our findings suggest a role for the alarmin IL-1α in the pathogenesis of MCMV retinitis in immunosuppressed mice. These findings may extend to the pathogenesis of HCMV retinitis in patients with AIDS or other forms of immunosuppression.

L-carnosine mitigates interleukin-1α-induced dry eye disease in rabbits via its antioxidant, anti-inflammatory, antiapoptotic, and antifibrotic effects.

OBJECTIVE: To elucidate the implications of L-carnosine on interleukin-1α (IL-1α)-induced inflammation of lacrimal glands (LGs). MATERIALS AND METHODS: Forty rabbits were divided equally into four groups: control group (G1), IL-1α (G2), L-carnosine (G3), and L-carnosine plus IL-1α (G4). Several clinical, histopathological, immunohistochemical, morphometric, and biochemical investigations were performed, followed by statistical analysis to diagnose the presence of dry eye disease (DED). RESULTS: The LGs of G2 rabbits showed degeneration of the acinar cells, increased deposition of collagen fibers, and marked immunoexpression of FasL; elevated levels of interferon-γ, tumor necrosis factor-α, transforming growth factor-ß1, and malondialdehyde; and decreased levels of glutathione peroxidase, superoxide dismutase, catalase, and reactive oxygen species compared with those of G1 rabbits. In contrast, administration of L-carnosine to G4 rabbits revealed marked improvement of all previously harmful changes in G2 rabbits, indicating the cytoprotective effects of L-carnosine against IL-1α-induced inflammation of LGs. CONCLUSIONS: IL-1α induced inflammation of LGs and eye dryness via oxidative stress, proinflammatory, apoptotic, and profibrotic effects, whereas L-carnosine mitigated DED through antioxidant, anti-inflammatory, antiapoptotic, and antifibrotic effects on LGs. Therefore, this work demonstrates for the first time that L-carnosine may be used as adjuvant therapy for the preservation of visual integrity in patients with DED.HighlightsIL-1α induced dry eye disease through its oxidative stress, proinflammatory, apoptotic and profibrotic effects on the lacrimal glands of rabbit.L-carnosine has antioxidant, anti-inflammatory, antiapoptotic and antifibrotic effects.L-carnosine mitigated IL-1α induced dry eye disease via elevating the levels of FasL, IFN-γ, TNF-α, TGFß1 and MDA as well as reducing the levels of antioxidants (GPx, SOD, and catalase) and ROS in the lacrimal glands of rabbit.L-carnosine could be used as a novel adjuvant therapy for the treatment of dry eye disease.

Virus-mediated inactivation of anti-apoptotic Bcl-2 family members promotes Gasdermin-E-dependent pyroptosis in barrier epithelial cells.

Two sets of innate immune proteins detect pathogens. Pattern recognition receptors (PRRs) bind microbial products, whereas guard proteins detect virulence factor activities by the surveillance of homeostatic processes within cells. While PRRs are well known for their roles in many types of infections, the role of guard proteins in most infectious contexts remains less understood. Here, we demonstrated that inhibition of protein synthesis during viral infection is sensed as a virulence strategy and initiates pyroptosis in human keratinocytes. We identified the BCL-2 family members MCL-1 and BCL-xL as sensors of translation shutdown. Virus- or chemical-induced translation inhibition resulted in MCL-1 depletion and inactivation of BCL-xL, leading to mitochondrial damage, caspase-3-dependent cleavage of gasdermin E, and release of interleukin-1α (IL-1α). Blocking this pathway enhanced virus replication in an organoid model of human skin. Thus, MCL-1 and BCL-xL can act as guard proteins within barrier epithelia and contribute to antiviral defense.

Pharmacological inhibition of MDA-9/Syntenin blocks breast cancer metastasis through suppression of IL-1ß.

Melanoma differentiation associated gene-9 (MDA-9), Syntenin-1, or syndecan binding protein is a differentially regulated prometastatic gene with elevated expression in advanced stages of melanoma. MDA-9/Syntenin expression positively associates with advanced disease stage in multiple histologically distinct cancers and negatively correlates with patient survival and response to chemotherapy. MDA-9/Syntenin is a highly conserved PDZ-domain scaffold protein, robustly expressed in a spectrum of diverse cancer cell lines and clinical samples. PDZ domains interact with a number of proteins, many of which are critical regulators of signaling cascades in cancer. Knockdown of MDA-9/Syntenin decreases cancer cell metastasis, sensitizing these cells to radiation. Genetic silencing of MDA-9/Syntenin or treatment with a pharmacological inhibitor of the PDZ1 domain, PDZ1i, also activates the immune system to kill cancer cells. Additionally, suppression of MDA-9/Syntenin deregulates myeloid-derived suppressor cell differentiation via the STAT3/interleukin (IL)-1ß pathway, which concomitantly promotes activation of cytotoxic T lymphocytes. Biologically, PDZ1i treatment decreases metastatic nodule formation in the lungs, resulting in significantly fewer invasive cancer cells. In summary, our observations indicate that MDA-9/Syntenin provides a direct therapeutic target for mitigating aggressive breast cancer and a small-molecule inhibitor, PDZ1i, provides a promising reagent for inhibiting advanced breast cancer pathogenesis.

IL-1α Mediates Innate and Acquired Resistance to Immunotherapy in Melanoma.

Inflammation has long been associated with cancer initiation and progression; however, how inflammation causes immune suppression in the tumor microenvironment and resistance to immunotherapy is not well understood. In this study, we show that both innate proinflammatory cytokine IL-1α and immunotherapy-induced IL-1α make melanoma resistant to immunotherapy. In a mouse melanoma model, we found that tumor size was inversely correlated with response to immunotherapy. Large tumors had higher levels of IL-1α, Th2 cytokines, polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs), and regulatory T cells but lower levels of IL-12, Th1 cytokines, and activated T cells. We found that therapy with adenovirus-encoded CD40L (rAd.CD40L) increased tumor levels of IL-1α and PMN-MDSCs. Blocking the IL-1 signaling pathway significantly decreased rAd.CD40L-induced PMN-MDSCs and their associated PD-L1 expression in the tumor microenvironment and enhanced tumor-specific immunity. Similarly, blocking the IL-1 signaling pathway improved the antimelanoma activity of anti-PD-L1 Ab therapy. Our study suggests that blocking the IL-1α signaling pathway may increase the efficacy of immunotherapies against melanoma.

Overexpression of Interleukin-1α Suppresses Liver Metastasis of Lymphoma: Implications for Antitumor Effects of CD8+ T-cells.

Interleukin (IL)-1 plays a key role in carcinogenesis, tumor progression, and metastasis. Although IL-1 may enhance the expansion of CD8+ T-cells, the pathological contribution of IL-1-activated CD8+ T-cells to tumor metastasis remains unclear. This study used a liver metastasis model of the EL4 T-cell lymphoma cells transplanted into human IL (hIL)-1α conditional transgenic (hIL-1α cTg) mice. Overproduction of hIL-1α suppressed both macroscopic and histological liver metastasis of EL4 T-cell lymphoma. The hIL-1α-induced inflammatory state increased the number of CD8+ T-cells both within and around metastatic tumors. Moreover, larger numbers of CD8+ T-cells showed greater infiltration of liver blood vessels in hIL-1α cTg mice than in control wild-type mice. Terminal deoxynucleotidyl transferase dUTP nick-end labeling staining of liver tissue from hIL-1α cTg mice indicated increased apoptosis of cells in the tumor. Localization of apoptosis cells resembled that of CD8+ T-cells. In addition, cytotoxicity assay showed that CD8+ T-cell counts from tumor-bearing hIL-1α cTg mice correlated with cytotoxicity against EL4. In summary, IL-1α suppresses lymphoma metastasis, and IL-1α-activated CD8+ T-cells may play important roles in inhibiting both tumor metastasis and metastatic tumor growth.

IL-1α antibody inhibits dose-dependent exacerbation of eosinophilic inflammation by crude house-dust-mite antigen in the conjunctiva of an atopic keratoconjunctivitis mouse model.

PURPOSE: To investigate whether crude house-dust-mite antigen exacerbates eosinophilic inflammation in the conjunctival tissues of an atopic keratoconjunctivitis mouse model in a dose-dependent manner. MATERIALS AND METHODS: An atopic keratoconjunctivitis mouse model was established by percutaneous sensitization and crude house-dust-mite antigen application in NC/Nga mice. To assess the dose-dependent response, conjunctival specimens from groups that were administered high- (High-HDM) or low-dose house-dust-mite antigen (Low-HDM) following percutaneous sensitization and the control without house-dust-mite antigen administration (control group) were evaluated. Histological examination and immunofluorescence staining were performed to determine eosinophil density and the number of IL-13-positive cells. Polymerase chain reaction array was used to obtain adaptive and innate immunity-related factor profile, and quantitative polymerase chain reaction was used to determine Il13, Il17a, Ccl11, and Ccl24 expression. Atopic keratoconjunctivitis model mice injected with anti-IL-1α antibody (IL-1α group) or vehicle (vehicle group) to the upper and lower eyelids before atopic keratoconjunctivitis development were evaluated. RESULTS: Eosinophil density in the conjunctiva increased with house-dust-mite antigen application in a dose-dependent manner. CD4, CXCL10, CCR6, C3, and IL-13 mRNA levels increased more than 5-fold in the conjunctiva of the High-HDM group animals compared to those in control animals. mRNA expression of Il13 and Ccl11 in the conjunctiva of the High-HDM group animals significantly increased compared with that in the Low-HDM and control group animals. Conversely, the eosinophil density and Il13 mRNA expression significantly decreased in the IL-1α group compared with those in the vehicle group. CONCLUSIONS: The house-dust-mite antigen increased eosinophilic infiltration and Il13 mRNA expression in the conjunctiva of an atopic keratoconjunctivitis mouse model in a dose-dependent manner. These inflammatory alterations were partially alleviated by eyelid injection of anti-IL-1α antibody. These findings indicate that IL-1α-induced IL-13 production constitutes a major exacerbating factor for house-dust-mite antigen-induced atopic keratoconjunctivitis.

Blocking PDGF-CC signaling ameliorates multiple sclerosis-like neuroinflammation by inhibiting disruption of the blood-brain barrier.

Disruption of blood-brain barrier (BBB) integrity is a feature of various neurological disorders. Here we found that the BBB is differently affected during the preclinical, progression and remission phase of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). We have identified an upregulation of pro-inflammatory and pro-angiogenic factors in the BBB transcriptome and down-regulation of endothelial tight junction members coinciding with elevated BBB leakage specifically during the progression phase. These changes were antagonized by blocking PDGFRα signaling with the small tyrosine kinase inhibitor imatinib. Moreover, targeting the PDGFRα ligand PDGF-CC using a neutralizing antibody, facilitated recovery of BBB integrity and improvement of EAE symptoms. Intracerebroventricular injection of PDGF-CC induced upregulation, whereas blocking PDGF-CC during EAE led to downregulation of Tnfa and Il1a at the BBB. Our findings suggest that blocking PDGF-CC counteracts fundamental aspects of endothelial cell activation and disruption of the BBB by decreasing Tnfa and Il1a expression. We also demonstrate that both PDGF-CC and its receptor PDGFRα were upregulated in MS lesions indicating that blocking PDGF-CC may be considered a novel treatment for MS.

Targeting IL-1ß in the Treatment of Atherosclerosis.

The role of inflammation in atherosclerosis has been recognized several decades ago and existing treatments provide benefits in part through non-specific anti-inflammatory actions. Compared with other cytokines, interleukin-1ß (IL-1ß) is associated with acute and chronic inflammation. Anti-inflammatory therapy with canakinumab targeting the IL-1ß innate immunity pathway could significantly reduce the rate of recurrent cardiovascular events than placebo. The results of CANTOS suggested an important role of IL-1ß in atherosclerosis. However, there are numerous mechanisms that are to be clarified. We herein discussed the important immunomodulatory effect IL-1ß exerts on atherosclerosis and the potential mechanisms underlying it. We also reviewed bench-to-bedside clinical translation of IL-1ß neutralizing strategies associated with the use of IL-1ß blockade in patients with atherosclerosis.

The Association between TNF-alpha, IL-1 alpha and IL-10 with Alzheimer's Disease.

BACKGROUND: Sporadic Alzheimer's Disease (AD) is assumed to be associated with different biological/genetic vulnerability, as well as with neuroinflammation, mediated by cytokines. The present study evaluated the role of cytokines in AD. OBJECTIVE: The aim was to determine the possible association of TNF-α (rs1800629), IL1-α (rs1800587) and IL-10 (rs1800896) polymorphisms with AD, and to assess serum TNF-α, IL-1α and IL-10 concentrations in patients with AD and in subjects with mild cognitive impairment (MCI). METHODS: The study included 645 Caucasian participants: 395 subjects with AD and 250 subjects with MCI. Genotyping was performed using real-time PCR in all 645 subjects, while serum concentrations of TNF-α, IL-1α and IL-10 and were determined using ELISA in 174 subjects. RESULTS: The frequency of the TNF-α rs1800629, IL1-α rs1800587 or IL-10 rs1800896 genotypes did not differ significantly between patients with AD and MCI. Serum concentration of IL-1α and IL-10 were significantly decreased, while the concentration of TNF-α was significantly higher in patients with AD than in MCI subjects. TNF-α, IL1-α or IL-10 concentrations were similar in subjects with AD or MCI subdivided into carriers of the corresponding TNF-α rs1800629, IL1-α rs1800587 or IL-10 rs1800896 genotypes. CONCLUSION: Similar distribution of the IL1-α rs1800587, TNF-α rs1800629 or IL-10 rs1800896 genotypes in subjects with AD and MCI failed to confirm that these specific risk genotypes are associated with vulnerability to develop AD. Alteration in IL-1α, IL-10 and TNF-α concentrations in patients with AD partially confirmed the association with the neuroinflammatory response in AD.

A Clearance Period after Soluble Lead Nanoparticle Inhalation Did Not Ameliorate the Negative Effects on Target Tissues Due to Decreased Immune Response.

The inhalation of metal (including lead) nanoparticles poses a real health issue to people and animals living in polluted and/or industrial areas. In this study, we exposed mice to lead(II) nitrate nanoparticles [Pb(NO3)2 NPs], which represent a highly soluble form of lead, by inhalation. We aimed to uncover the effects of their exposure on individual target organs and to reveal potential variability in the lead clearance. We examined (i) lead biodistribution in target organs using laser ablation and inductively coupled plasma mass spectrometry (LA-ICP-MS) and atomic absorption spectrometry (AAS), (ii) lead effect on histopathological changes and immune cells response in secondary target organs and (iii) the clearance ability of target organs. In the lungs and liver, Pb(NO3)2 NP inhalation induced serious structural changes and their damage was present even after a 5-week clearance period despite the lead having been almost completely eliminated from the tissues. The numbers of macrophages significantly decreased after 11-week Pb(NO3)2 NP inhalation; conversely, abundance of alpha-smooth muscle actin (α-SMA)-positive cells, which are responsible for augmented collagen production, increased in both tissues. Moreover, the expression of nuclear factor κB (NF-κB) and selected cytokines, such as tumor necrosis factor alpha (TNFα), transforming growth factor beta 1 (TGFß1), interleukin 6(IL-6), IL-1α and IL-1ß , displayed a tissue-specific response to lead exposure. In summary, diminished inflammatory response in tissues after Pb(NO3)2 NPs inhalation was associated with prolonged negative effect of lead on tissues, as demonstrated by sustained pathological changes in target organs, even after long clearance period.

A novel serine protease inhibitor as potential treatment for dry eye syndrome and ocular inflammation.

Dry eye syndrome (DES), a multifactorial disorder which leads to ocular discomfort, visual disturbance and tear film instability, has a rising prevalence and limited treatment options. In this study, a newly developed trypsin-like serine protease inhibitor (UAMC-00050) in a tear drop formulation was evaluated to treat ocular inflammation. A surgical animal model of dry eye was employed to investigate the potential of UAMC-00050 on dry eye pathology. Animals treated with UAMC-00050 displayed a significant reduction in ocular surface damage after evaluation with sodium fluorescein, compared to untreated, vehicle treated and cyclosporine-treated animals. The concentrations of IL-1α and TNF-α were also significantly reduced in tear fluid from UAMC-00050-treated rats. Additionally, inflammatory cell infiltration in the palpebral conjunctiva (CD3 and CD45), was substantially reduced. An accumulation of pro-MMP-9 and a decrease in active MMP-9 were found in tear fluid from animals treated with UAMC-00050, suggesting that trypsin-like serine proteases play a role in activating MMP-9 in ocular inflammation in this animal model. Comparative qRT-PCR analyses on ocular tissue indicated the upregulation of tryptase, urokinase plasminogen activator receptor (uPAR) and protease-activated receptor 2 (PAR2). The developed UAMC-00050 formulation was stable up to 6 months at room temperature in the absence of light, non-irritating and sterile with compatible pH and osmolarity. These results provide a proof-of-concept for the in vivo modifying potential of UAMC-00050 on dry eye pathology and suggest a central role of trypsin-like serine proteases and PAR2 in dry eye derived ocular inflammation.

Cytokine-induced Killer T Cells Enhance the Cytotoxicity Against Carboplatin-resistant Ovarian Cancer Cells.

BACKGROUND/AIM: Ovarian cancer (OC) is typically diagnosed at an advanced stage with limitations for cure. Cytokine-induced killer (CIK) T cell therapy exerts significant cytotoxic effects against cancer cells and reduces the adverse effects of chemotherapy. Herein, we performed a flow cytometry-based method to evaluate the cytotoxicity of peripheral blood mononuclear cells-derived CIK cells against OC cells. MATERIALS AND METHODS: The CIK cells were induced and expanded using an interferon-γ/IL-2-based xeno-free medium system. The cytotoxicity of CIK cells or carboplatin against OC cells was examined. RESULTS: The CIK cells showed an NK-like phenotypic characteristic and dose-dependently increased cytotoxicity against OC cells. We found that the number of advanced OC cells, which were more resistant to carboplatin, was dramatically decreased by an additional one-shot CIK treatment. CONCLUSION: CIK cells have a potent cytotoxic ability that would be explored as an alternative strategy for cancer treatment in the near future.