Spheroid formation induces chemokine production in trophoblast-derived Swan71 cells.

PROBLEM: In the cell column of anchoring villi, the cytotrophoblast differentiates into extravillous trophoblast (EVT) and invades the endometrium in contact with maternal immune cells. Recently, chemokines were proposed to regulate the decidual immune response. To investigate the roles of chemokines around the anchoring villi, we examined the expression profiles of chemokines in the first-trimester trophoblast-derived Swan71 cells using a three-dimensional culture model. METHOD OF STUDY: The gene expressions in the spheroid-formed Swan71 cells were examined by microarray and qPCR analyses. The protein expressions were examined by immunochemical staining. The chemoattractant effects of spheroid-formed Swan71 cells were examined by migration assay using monocyte-derived THP-1 cells. RESULTS: The expressions of an EVT marker, laeverin, and matrix metalloproteases, MMP2 and MMP9, were increased in the spheroid-cultured Swan71 cells. Microarray and qPCR analysis revealed that mRNA expressions of various chemokines, CCL2, CCL7, CCL20, CXCL1, CXCL2, CXCL5, CXCL6, CXCL8, and CXCL10, in the spheroid-cultured Swan71 cells were up-regulated as compared with those in the monolayer-cultured Swan71 cells. These expressions were significantly suppressed by hypoxia. Migration assay showed that culture media derived from the spheroid-formed Swan71 cells promoted THP-1 cell migration. CONCLUSION: This study indicated that chemokine expressions in Swan71 cells increase under a spheroid-forming culture and the culture media have chemoattractant effects. Since three-dimensional cell assembling in the spheroid resembles the structure of the cell column, this study also suggests that chemokines play important roles in the interaction between EVT and immune cells in their early differentiation stage.

Chemerin promotes proliferation and migration of ovarian cancer cells by upregulating expression of PD-L1.

Ovarian cancer is the third-most-common malignant reproductive tumor in women. According to the American Cancer Society, it has the highest mortality rate of gynecological tumors. The five-year survival rate was only 29% during the period from 1975 to 2008 (Reid et al., 2017). In recent decades, the five-year survival rate of ovarian cancer has remained around 30% despite continuous improvements in surgery, chemotherapy, radiotherapy, and other therapeutic methods. However, because of the particularity of the volume and location of ovarian tissue, the early symptoms of ovarian cancer are hidden, and there is a lack of highly sensitive and specific screening methods. Most patients have advanced metastasis, including abdominal metastasis, when they are diagnosed (Reid et al., 2017). Therefore, exploring the mechanism of ovarian cancer metastasis and finding early preventive measures are key to improving the survival rate and reducing mortality caused by ovarian cancer.

Nanocurcumin Potently Inhibits SARS-CoV-2 Spike Protein-Induced Cytokine Storm by Deactivation of MAPK/NF-κB Signaling in Epithelial Cells.

Interleukin-mediated deep cytokine storm, an aggressive inflammatory response to SARS-CoV-2 virus infection in COVID-19 patients, is correlated directly with lung injury, multi-organ failure, and poor prognosis of severe COVID-19 patients. Curcumin (CUR), a phenolic antioxidant compound obtained from turmeric (Curcuma longa L.), is well-known for its strong anti-inflammatory activity. However, its in vivo efficacy is constrained due to poor bioavailability. Herein, we report that CUR-encapsulated polysaccharide nanoparticles (CUR-PS-NPs) potently inhibit the release of cytokines, chemokines, and growth factors associated with damage of SARS-CoV-2 spike protein (CoV2-SP)-stimulated liver Huh7.5 and lung A549 epithelial cells. Treatment with CUR-PS-NPs effectively attenuated the interaction of ACE2 and CoV2-SP. The effects of CUR-PS-NPs were linked to reduced NF-κB/MAPK signaling which in turn decreased CoV2-SP-mediated phosphorylation of p38 MAPK, p42/44 MAPK, and p65/NF-κB as well as nuclear p65/NF-κB expression. The findings of the study strongly indicate that organic NPs of CUR can be used to control hyper-inflammatory responses and prevent lung and liver injuries associated with CoV2-SP-mediated cytokine storm.

Chemerin promotes proliferation and migration of ovarian cancer cells by upregulating expression of PD-L1 / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Ovarian cancer is the third-most-common malignant reproductive tumor in women. According to the American Cancer Society, it has the highest mortality rate of gynecological tumors. The five-year survival rate was only 29% during the period from 1975 to 2008 (Reid et al., 2017). In recent decades, the five-year survival rate of ovarian cancer has remained around 30% despite continuous improvements in surgery, chemotherapy, radiotherapy, and other therapeutic methods. However, because of the particularity of the volume and location of ovarian tissue, the early symptoms of ovarian cancer are hidden, and there is a lack of highly sensitive and specific screening methods. Most patients have advanced metastasis, including abdominal metastasis, when they are diagnosed (Reid et al., 2017). Therefore, exploring the mechanism of ovarian cancer metastasis and finding early preventive measures are key to improving the survival rate and reducing mortality caused by ovarian cancer.

Spleen tyrosine kinase facilitates neutrophil activation and worsens long-term neurologic deficits after spinal cord injury.

BACKGROUND: Spinal cord injury elicits widespread inflammation that can exacerbate long-term neurologic deficits. Neutrophils are the most abundant immune cell type to invade the spinal cord in the early acute phase after injury, however, their role in secondary pathogenesis and functional recovery remains unclear. We have previously shown that neutrophil functional responses during inflammation are augmented by spleen tyrosine kinase, Syk, a prominent intracellular signaling enzyme. In this study, we evaluated the contribution of Syk towards neutrophil function and long-term neurologic deficits after spinal cord injury. METHODS: Contusive spinal cord injury was performed at thoracic vertebra level 9 in mice with conditional deletion of Syk in neutrophils (Sykf/fMRP8-Cre). Hindlimb locomotor recovery was evaluated using an open-field test for 35 days following spinal cord injury. Long-term white matter sparing was assessed using eriochrome cyanide staining. Blood-spinal cord barrier disruption was evaluated by immunoblotting. Neutrophil infiltration, activation, effector functions, and cell death were determined by flow cytometry. Cytokine and chemokine expression in neutrophils was assessed using a gene array. RESULTS: Syk deficiency in neutrophils improved long-term functional recovery after spinal cord injury, but did not promote long-term white matter sparing. Neutrophil activation, cytokine expression, and cell death in the acutely injured spinal cord were attenuated by the genetic loss of Syk while neutrophil infiltration and effector functions were not affected. Acute blood-spinal cord barrier disruption was also unaffected by Syk deficiency in neutrophils. CONCLUSIONS: Syk facilitates specific neutrophil functional responses to spinal cord injury including activation, cytokine expression, and cell death. Long-term neurologic deficits are exacerbated by Syk signaling in neutrophils independent of acute blood-spinal cord barrier disruption and long-term white matter sparing. These findings implicate Syk in pathogenic neutrophil activities that worsen long-term functional recovery after spinal cord injury.

RIPK3 signaling and its role in the pathogenesis of cancers.

RIPK3 (receptor-interacting protein kinase 3) is a serine/threonine-protein kinase. As a key component of necrosomes, RIPK3 is an essential mediator of inflammatory factors (such as TNFα-tumor necrosis factor α) and infection-induced necroptosis, a programmed necrosis. In addition, RIPK3 signaling is also involved in the regulation of apoptosis, cytokine/chemokine production, mitochondrial metabolism, autophagy, and cell proliferation by interacting with and/or phosphorylating the critical regulators of the corresponding signaling pathways. Similar to apoptosis, RIPK3-signaling-mediated necroptosis is inactivated in most types of cancers, suggesting RIPK3 might play a critical suppressive role in the pathogenesis of cancers. However, in some inflammatory types of cancers, such as pancreatic cancers and colorectal cancers, RIPK3 signaling might promote cancer development by stimulating proliferation signaling in tumor cells and inducing an immunosuppressive response in the tumor environment. In this review, we summarize recent research progress in the regulators of RIPK3 signaling, and discuss the function of this pathway in the regulation of mixed lineage kinase domain-like (MLKL)-mediated necroptosis and MLKL-independent cellular behaviors. In addition, we deliberate the potential roles of RIPK3 signaling in the pathogenesis of different types of cancers and discuss the potential strategies for targeting this pathway in cancer therapy.

Defective expansion and function of memory like natural killer cells in HIV+ individuals with latent tuberculosis infection.

PURPOSE: Tuberculosis (TB) is the leading cause of infectious disease related mortality, and only 10% of the infected individuals develop active disease. The likelihood of progression of latent tuberculosis infection (LTBI) to active TB disease is high in HIV infected individuals. Identification of HIV+ individuals at risk would allow treating targeted population, facilitating completion of therapy for LTBI and prevention of TB development. NK cells have an important role in T cell independent immunity against TB, but the exact role of NK cell subsets in LTBI and HIV is not well characterized. METHODS: In this study, we compared the expansion and function of memory like NK cells from HIV-LTBI+ individuals and treatment naïve HIV+LTBI+ patients in response to Mtb antigens ESAT-6 and CFP-10. RESULTS: In freshly isolated PBMCs, percentages of CD3-CD56+ NK cells were similar in HIV+LTBI+ patients and healthy HIV-LTBI+ individuals. However, percentages of CD3-CD56+CD16+ NK cells were higher in healthy HIV-LTBI+ individuals compared to HIV+LTBI+ patients. HIV infection also inhibited the expansion of memory like NK cells, production of IL-32α, IL-15 and IFN-γ in response to Mtb antigens in LTBI+ individuals. CONCLUSION: We studied phenotypic, functional subsets and activation of memory like-NK cells during HIV infection and LTBI. We observed that HIV+LTBI+ patients demonstrated suboptimal NK cell and monocyte interactions in response to Mtb, leading to reduced IL-15, IFN-γ and granzyme B and increased CCL5 production. Our study highlights the effect of HIV and LTBI on modulation of NK cell activity to understand their role in development of interventions to prevent progression to TB in high risk individuals.

Human Placental Trophoblasts Infected by Listeria monocytogenes Undergo a Pro-Inflammatory Switch Associated With Poor Pregnancy Outcomes.

The placenta controls the growth of the fetus and ensures its immune protection. Key to these functions, the syncytiotrophoblast (SYN) is a syncytium formed by fusion of underlying mononuclear trophoblasts. The SYN covers the placental surface and is bathed in maternal blood to mediate nutritional and waste exchanges between the mother and fetus. The bacterial pathogen Listeria monocytogenes breaches the trophoblast barrier and infects the placental/fetal unit resulting in poor pregnancy outcomes. In this work, we analyzed the L. monocytogenes intracellular lifecycle in primary human trophoblasts. In accordance with previous studies, we found that the SYN is 20-fold more resistant to infection compared to mononuclear trophoblasts, forming a protective barrier to infection at the maternal interface. We show for the first time that this is due to a significant reduction in L. monocytogenes uptake by the SYN rather than inhibition of the bacterial intracellular division or motility. We here report the first transcriptomic analysis of L. monocytogenes-infected trophoblasts (RNA sequencing). Pathway analysis showed that infection upregulated TLR2, NOD-like, and cytosolic DNA sensing pathways, as well as downstream pro-inflammatory circuitry (NF-κB, AP-1, IRF4, IRF7) leading to the production of mediators known to elicit the recruitment and activation of maternal leukocytes (IL8, IL6, TNFα, MIP-1). Signature genes associated with poor pregnancy outcomes were also upregulated upon infection. Measuring the release of 54 inflammatory mediators confirmed the transcriptomic data and revealed sustained production of tolerogenic factors (IL-27, IL-10, IL-1RA, TSLP) despite infection. Both the SYN and mononuclear trophoblasts produced cytokines, but surprisingly, some cytokines were predominantly produced by the SYN (IL-8, IL-6) or by non-fused trophoblasts (TNFα). Collectively, our data support that trophoblasts act as placental gatekeepers that limit and detect L. monocytogenes infection resulting in a pro-inflammatory response, which may contribute to the poor pregnancy outcomes if the pathogen persists.

Super enhancer regulation of cytokine-induced chemokine production in alcoholic hepatitis.

Alcoholic hepatitis (AH) is associated with liver neutrophil infiltration through activated cytokine pathways leading to elevated chemokine expression. Super-enhancers are expansive regulatory elements driving augmented gene expression. Here, we explore the mechanistic role of super-enhancers linking cytokine TNFα with chemokine amplification in AH. RNA-seq and histone modification ChIP-seq of human liver explants show upregulation of multiple CXCL chemokines in AH. Liver sinusoidal endothelial cells (LSEC) are identified as an important source of CXCL expression in human liver, regulated by TNFα/NF-κB signaling. A super-enhancer is identified for multiple CXCL genes by multiple approaches. dCas9-KRAB-mediated epigenome editing or pharmacologic inhibition of Bromodomain and Extraterminal (BET) proteins, transcriptional regulators vital to super-enhancer function, decreases chemokine expression in vitro and decreases neutrophil infiltration in murine models of AH. Our findings highlight the role of super-enhancer in propagating inflammatory signaling by inducing chemokine expression and the therapeutic potential of BET inhibition in AH treatment.

Irradiation Accelerates Plaque Formation and Cellular Senescence in Flow-Altered Carotid Arteries of Apolipoprotein E Knock-Out Mice.

Background Chronic inflammation through cellular senescence, known as the senescence-associated secretory phenotype, is a mechanism of various organ diseases, including atherosclerosis. Particularly, ionizing radiation (IR) contributes to cellular senescence by causing DNA damage. Although previous clinical studies have demonstrated that radiotherapy causes atherosclerosis as a long-term side effect, the detailed mechanism is unclear. This study was conducted to investigate the relationship between radiation-induced atherosclerosis and senescence-associated secretory phenotype in murine carotid arteries. Methods and Results Partial ligation of the left carotid artery branches in 9-week-old male apolipoprotein E-deficient mice was performed to induce atherosclerosis. The mice received total body irradiation at a dose of 6 Gy using gamma rays at 2 weeks post operation. We compared the samples collected 4 weeks after IR with unirradiated control samples. The IR and control groups presented pathologically progressive lesions in 90.9% and 72.3% of mice, respectively. Plaque volume, macrophage accumulation, and phenotype switching of vascular smooth muscle cells were advanced in the IR group. Irradiated samples showed increased persistent DNA damage response (53BP1 [p53 binding protein 1]), upregulated cyclin-dependent kinase inhibitors (p16INK4a and p21), and elevated inflammatory chemokines expression (monocyte chemotactic protein-1, keratinocyte-derived chemokine, and macrophage inflammatory protein 2). Conclusions IR promoted plaque growth in murine carotid arteries. Our findings support the possibility that senescence-associated secretory phenotype aggravates atherogenesis in irradiated artery. This mice model might contribute to mechanism elucidation of radiation-induced atherosclerosis.

IFN-γ is essential for alveolar macrophage-driven pulmonary inflammation in macrophage activation syndrome.

Macrophage activation syndrome (MAS) is a life-threatening cytokine storm complicating systemic juvenile idiopathic arthritis (SJIA) driven by IFN-γ. SJIA and MAS are also associated with an unexplained emerging inflammatory lung disease (SJIA-LD), with our recent work supporting pulmonary activation of IFN-γ pathways pathologically linking SJIA-LD and MAS. Our objective was to mechanistically define the potentially novel observation of pulmonary inflammation in the TLR9 mouse model of MAS. In acute MAS, lungs exhibit mild but diffuse CD4-predominant, perivascular interstitial inflammation with elevated IFN-γ, IFN-induced chemokines, and alveolar macrophage (AMϕ) expression of IFN-γ-induced genes. Single-cell RNA sequencing confirmed IFN-driven transcriptional changes across lung cell types with myeloid expansion and detection of MAS-specific macrophage populations. Systemic MAS resolution was associated with increased AMϕ and interstitial lymphocytic infiltration. AMϕ transcriptomic analysis confirmed IFN-γ-induced proinflammatory polarization during acute MAS, which switches toward an antiinflammatory phenotype after systemic MAS resolution. Interestingly, recurrent MAS led to increased alveolar inflammation and lung injury, and it reset AMϕ polarization toward a proinflammatory state. Furthermore, in mice bearing macrophages insensitive to IFN-γ, both systemic features of MAS and pulmonary inflammation were attenuated. These findings demonstrate that experimental MAS induces IFN-γ-driven pulmonary inflammation replicating key features of SJIA-LD and provides a model system for testing potentially novel treatments directed toward SJIA-LD.

Physiology of Cultured Human Microglia Maintained in a Defined Culture Medium.

Microglia are the primary immune cell of the CNS, comprising 5-20% of the ∼60 billion neuroglia in the human brain. In the developing and adult CNS, they preferentially target active neurons to guide synapse maturation and remodeling. At the same time, they are the first line of defense against bacterial, fungal, and viral CNS infections. Although an extensive literature details their roles in rodents, less is known about how they function in humans because of the difficulty in obtaining tissue samples and the understandable inability to extensively study human microglia in situ. In this study, we use recent advances in the study of brain microenvironments to establish cultures of primary human microglia in a serum-free medium. Postsurgical samples of human brain were enzymatically and mechanically dissociated into single cells, and microglia were isolated at high purity by positive selection using CD11b Ab-coated microbeads. The CD11b+ cells were plated on poly-l-lysine-coated surfaces and bathed in serum-free DMEM/F12 supplemented with three essential components (TGF-ß, IL-34, and cholesterol). Under these conditions, microglia assumed a ramified morphology, showed limited proliferation, actively surveyed their surroundings, and phagocytosed bacterial microparticles. In the presence of LPS, they assumed a more compact shape and began production of proinflammatory cytokines and reactive oxygen species. LPS on its own triggered release of TNF-α, whereas release of IL-1ß required costimulation by ATP. Thus, human microglia maintained in a defined medium replicate many of the characteristics expected of native cells in the brain and provide an accessible preparation for investigations of human microglial physiology, pharmacology, and pathophysiology.

Studies of molecular pathways associated with blood neutrophil corticosteroid insensitivity in equine asthma.

Severe equine asthma is characterized by airway hyperresponsiveness, neutrophilic inflammation and structural alterations of the lower airways. In asthmatic horses with neutrophilic inflammation, there is insensitivity to corticosteroids characterized by the persistence of neutrophils within the airways with therapy. We hypothesized that hypoxia or oxidative stress in the microenvironment of the lung contributes to this insensitivity of neutrophils to corticosteroids in asthmatic horses. Blood neutrophils isolated from horses with severe asthma (N = 8) and from healthy controls (N = 8) were incubated under different cell culture conditions simulating hypoxia and oxidative stress and, in the presence, or absence of dexamethasone. The pro-inflammatory gene and protein expression of neutrophils were studied. In both groups, pyocyanin-induced oxidative stress increased the mRNA expression of IL-8, IL-1ß, and TNF-α. While IL-1ß and TNF-α were downregulated by dexamethasone under these conditions, IL-8 was not. Simulated hypoxic conditions did not enhance pro-inflammatory gene expression in neutrophils from either group of horses. In conclusion, oxidative stress but not hypoxia may contribute to corticosteroid insensitivity via a selective gene regulation pathway. Equine neutrophil responses were similar in both heathy and asthmatic horses, indicating that it is not specific to asthmatic inflammation.

Expression of Resistin, Chemerin, and Chemerin's Receptor in the Unstable Carotid Atherosclerotic Plaque.

Background and Purpose: Unstable carotid plaques are a common cause of ischemic strokes. Identifying markers that reflect/contribute to plaque instability has become a prominent focus in cardiovascular research. The adipokines, resistin and chemerin, and ChemR23 (chemerin receptor), may play a role in carotid atherosclerosis, making them potential candidates to assess plaque instability. However, the expression and interrelationship of resistin and chemerin (and ChemR23) protein and mRNA within the carotid atherosclerotic plaque remains elusive. Thus, we investigated herein, the association between plaque mRNA and protein expression of resistin and chemerin (and ChemR23) and carotid plaque instability in humans, and whether sex differences exist in the relationship between these adipokines and plaque instability. Methods: Human carotid plaques were processed for immunohistochemical/mRNA analysis of resistin, chemerin, and ChemR23. Plaque instability was assessed by gold-standard histological classifications. A semi-quantitative scoring system was used to determine the intensity of adipokine expression on macrophages/foam cells, as well as the percentage of inflammatory cells stained positive. Plaque adipokine protein expression was also digitally quantified and mRNA expression was assessed by qRT-PCR. Results: Resistin and chemerin mRNA expression was 80% and 32% lower, respectively, in unstable versus stable plaques (P<0.05), while no difference in ChemR23 mRNA expression was observed. In contrast, greater resistin staining intensity and percentage of cells stained positive were detected in unstable versus stable plaques (P<0.01). Similarly, chemerin and ChemR23 staining intensity and percentage of cells stained were positively associated with plaque instability (P<0.05). No strong sex-specific relationship was observed between adipokines and plaque instability. Conclusions: This study examined the relationship between resistin, chemerin, and ChemR23, and carotid plaque instability, with a specific analysis at the plaque level. We reported a positive association between plaque instability and protein levels of resistin, chemerin, and ChemR23 but a negative association with resistin and chemerin mRNA expression. This suggests these adipokines exert proinflammatory roles in the process of carotid atherosclerosis and may be regulated via a negative feedback regulatory mechanism.

Performance of capecitabine in novel combination therapies in colorectal cancer.

Colorectal cancer is one of the most common cancers throughout the world, and no definitive cure has ever been found. Perhaps a new insight into the effectiveness of chemotherapy drugs could help better treat patients. Targeted therapies have significantly improved the median overall survival of colorectal cancer patients. One of the standard chemotherapy regimens used for colorectal cancer is capecitabine, which is important in monotherapy and combination therapies. Capecitabine, with other chemotherapeutic agents (irinotecan, oxaliplatin, perifosine, 17-allylamino-17-demethoxygeldanamycin, aspirin, celecoxib, statins, quinacrine, inositol hexaphosphate and inositol, cystine/theanine, curcumin, and isorhamnetin), and biological ones (antibodies) plays an important role in the inhibition of some signaling pathways, increasing survival, reducing tumor growth and side effects of capecitabine. However, some drugs, such as proton pump inhibitors, are negatively related to capecitabine; therefore, the purpose of this work is to review and discuss the performance of capecitabine combination therapies in colorectal cancer.

Conversion of AML-blasts to leukemia-derived dendritic cells (DCleu) in 'DC-culture-media' shifts correlations of released chemokines with antileukemic T-cell reactions.

Dendritic cells (DC) and T-cells are mediators of CTL-responses. Autologous (from patients with acute myeloid leukaemia (AML) or myelodysplasia (MDS)) or allogeneic (donor)-T-cells stimulated by DCleu, gain an efficient lysis of naive blasts, although not in every case. CXCL8, -9, -10, CCL2, -5 and Interleukin (IL-12) were quantified by Cytometric Bead Array (CBA) in supernatants from 5 DC-generating methods and correlated with AML-/MDS-patients' serum-values, DC-/T-cell-interactions/antileukemic T-cell-reactions after mixed lymphocyte culture (MLC) and patients' clinical course. The blast-lytic activity of T-cells stimulated with DC or mononuclear cells (MNC) was quantified in a cytotoxicity assay. Despite great variations of chemokine-levels, correlations with post-stimulation (after stimulating T-cells with DC in MLC) improved antileukemic T-cell activity were seen: higher released chemokine-values correlated with improved T-cells' antileukemic activity (compared to stimulation with blast-containing MNC) - whereas with respect to the corresponding serum values higher CXCL8-, -9-, and -10- but lower CCL5- and -2-release correlated with improved antileukemic activity of DC-stimulated (vs. blast-stimulated) T-cells. In DC-culture supernatants higher chemokine-values correlated with post-stimulation improved antileukemic T-cell reactivity, whereas higher serum-values of CXCL8, -9, and -10 but lower serum-values of CCL5 and -2 correlated with post-stimulation improved antileukemic T-cell-reactivity. In a context of 'DC'-stimulation (vs serum) this might point to a change of (CCL5 and -2-associated) functionality from a more 'inflammatory' or 'tumor-promoting' to a more 'antitumor'-reactive functionality. This knowledge could contribute to develop immune-modifying strategies that promote antileukemic (adaptive) immune-responses.

Novel signaling pathways regulate SARS-CoV and SARS-CoV-2 infectious disease.

ABSTRACT: Severe acute respiratory syndrome coronavirus (SARS-CoV)-2 induces severe infection, and it is responsible for a worldwide disease outbreak starting in late 2019. Currently, there are no effective medications against coronavirus. In the present study, we utilized a holistic bioinformatics approach to study gene signatures of SARS-CoV- and SARS-CoV-2-infected Calu-3 lung adenocarcinoma cells. Through the Gene Ontology platform, we determined that several cytokine genes were up-regulated after SARS-CoV-2 infection, including TNF, IL6, CSF2, IFNL1, IL-17C, CXCL10, and CXCL11. Differentially regulated pathways were detected by the Kyoto Encyclopedia of Genes and Genomes, gene ontology, and Hallmark platform, including chemokines, cytokines, cytokine receptors, cytokine metabolism, inflammation, immune responses, and cellular responses to the virus. A Venn diagram was utilized to illustrate common overlapping genes from SARS-CoV- and SARS-CoV-2-infected datasets. An Ingenuity pathway analysis discovered an enrichment of tumor necrosis factor- (TNF-) and interleukin (IL)-17-related signaling in a gene set enrichment analysis. Downstream networks were predicted by the Database for Annotation, Visualization, and Integrated Discovery platform also revealed that TNF and TNF receptor 2 signaling elicited leukocyte recruitment, activation, and survival of host cells after coronavirus infection. Our discovery provides essential evidence for transcript regulation and downstream signaling of SARS-CoV and SARS-CoV-2 infection.

Use of selective PGE2 receptor antagonists on human endometriotic stromal cells and peritoneal macrophages.

Non-hormonal therapeutic strategies for endometriosis are needed. The aim of this study was to characterize the effects of prostaglandin (PG)E2 receptor inhibitors to explore their potential as novel therapeutic strategies for endometriosis. The expression of PGE2 receptors (EP2 and EP4) in donated tissues from human ovarian endometriosis, adenomyosis and peritoneal endometriosis was examined using immunohistochemistry. Human endometriotic stromal cells (ESC) isolated from ovarian endometriotic tissue and peritoneal macrophages were treated with EP2 and EP4 antagonists. cAMP accumulation and the effect of EP antagonists were measured using cAMP assays. DNA synthesis in ESC was detected using bromodeoxyuridine incorporation analysis. Interleukin (IL)-6 and IL-8 protein levels in ESC supernatants were measured using ELISAs. mRNA expression level for aromatase by ESC, and selected cytokines by peritoneal macrophages was measured using RT-PCR. EP2 and EP4 receptors were expressed in cells derived from control and diseased tissue, ovarian endometriotic, adenomyotic and peritoneal lesions. A selective EP2 antagonist reduced DNA synthesis, cAMP accumulation and IL-1ß-induced proinflammatory cytokine secretion and aromatase expression. A selective EP4 antagonist negated IL-1ß-induced IL-6 secretion and aromatase expression. In peritoneal macrophages, EP expression was elevated in endometriosis samples but the EP4 antagonist reduced cAMP levels and expression of vascular endothelial growth factor, chemokine ligand 2 and chemokine ligand 3 mRNA. EP2 and EP4 are functioning in endometriosis lesions and peritoneal macrophages, and their selective antagonists can reduce EP-mediated actions, therefore, the EP antagonists are potential therapeutic agents for controlling endometriosis.

Kidney Subcapsular Allograft Transplants as a Model to Test Virus-Derived Chemokine-Modulating Proteins as Therapeutics.

Solid tissue transplant is a growing medical need that is further complicated by a limited donor organ supply. Acute and chronic rejection occurs in nearly all transplants and reduces long-term graft survival, thus increasing the need for repeat transplantation. Viruses have evolved highly adapted responses designed to evade the host's immune defenses. Immunomodulatory proteins derived from viruses represent a novel class of potential therapeutics that are under investigation as biologics to attenuate immune-mediated rejection and damage. These immune-modulating proteins have the potential to reduce the need for traditional posttransplant immune suppressants and improve graft survival. The myxoma virus-derived protein M-T7 is a promising biologic that targets chemokine and glycosaminoglycan pathways central to kidney transplant rejection. Orthotopic transplantations in mice are prohibitively difficult and costly and require a highly trained microsurgeon to successfully perform the procedure. Here we describe a kidney-to-kidney subcapsular transplant model as a practical and simple method for studying transplant rejection, a model that requires fewer mice. One kidney can be used as a donor for transplants into six or more recipient mice. Using this model there is lower morbidity, pain, and mortality for the mice. Subcapsular kidney transplantation provides a first step approach to testing virus-derived proteins as new potential immune-modulating therapeutics to reduce transplant rejection and inflammation.