Danielle L. Germundson, "Chronic Allergen Exposure Results in Microgliosis and Increased Blood Brain Barrier Permeability in a Mouse Model of Cow’s Milk Allergy"
Prolonged peripheral inflammation is thought to pose a risk for the development of neuropsychiatric and neurodegenerative disorders through sustained neuroinflammation. Such chronic inflammation may occur by repeated exposure to an allergen in sensitized individuals. For example, a mildly food-allergic individual who does not experience anaphylaxis may continue to consume the allergen. Under the hypothesis that long-term exposure to an allergen by an individual with subclinical food allergy results in neuroinflammation and associated neuropathology, we investigated glial reactivity, blood-brain barrier integrity, and peripheral immune cell migration into the brain in a mouse model of mild cow’s milk allergy. We sensitized 4-week-old male C57BL/6J mice to a vehicle or the vehicle containing a bovine whey allergen, β-lactoglobulin (BLG), for 5 weeks and placed the mice on a whey-containing diet for 2 weeks to simulate repeated allergen exposure. While health and growth were not overtly affected by the sensitization protocol, sensitized mice had significantly increased levels of BLG-specific IgE compared to the vehicle-sensitized sham mice. Immunostaining the brains of sensitized mice for Iba1 showed a greater number of microglia with reactive morphology throughout. Vascular permeability was assessed by the seepage of IgG from blood vessels into brain parenchyma by immunohistochemistry. Diffused IgG staining indicative of a ‘leaky’ blood brain barrier was observed in BLG-sensitized mice while more defined staining outlined blood vessels of sham mice. Interestingly, we also observed increased numbers of brain mast cells localized around vascularized regions of the brain in BLG-sensitized mice. Taken together, these results indicate that chronic exposure to an allergen, even without overt anaphylaxis, can produce observable neuropathology suggestive of neuroinflammation. Furthermore, our experimental paradigm establishes a novel tool to study food-allergy associated changes in the brain and highlights the possibility of allergen avoidance to prevent the development of neuropsychiatric or neurodegenerative disorders in susceptible individuals.
Junguk Hur, Ph.D., "Literature mining-based analysis of host-coronavirus gene-gene interactions"
Human coronaviruses have given rise to a series of major crises in global public health. The COVID-19 pandemic, caused by Severe Acute Respiratory Syndrome Coronavirus 2, has caused over 24.9 million confirmed cases with 838 thousand deaths worldwide and 5.0 million confirmed cases with 183 thousand deaths in the USA alone. Numerous research on COVID-19 is ongoing and their novel findings are published daily. To develop efficient mechanism-based therapeutics against COVID-19, it is critical to systematically study the interactions between hosts and coronaviruses. In this study, we used our in-house centrality- and network-based literature mining analysis platform, SciMiner, to perform a preliminary gene-interaction analysis of COVID-19. The abstracts of 32,363 COVID-19-related articles, compiled in LitCovid, a curated literature hub containing up-to-date scientific information about the 2019 novel Coronavirus. SciMiner identified 103 host genes and 19 coronavirus genes, represented in Ontology of Genes and Genomes, and 226 unique interactions among them, detected at the sentence level. These interactions were clearly separated into two clusters, one related to viral invasion and another one associated with host-immune responses. This comprehensive literature-derived COVID-19 gene interaction network may facilitate the generation of novel hypotheses about pathogenic mechanisms and the identification of potential therapeutic targets.
Harpreet Kaur, "Impact of modulating gut bacteria using antibiotic, probiotic, and prebiotic interventions in AppNL-G-F mouse model of Alzheimer disease"
Background: Alzheimer's disease (AD) is a multifactorial, neurodegenerative disorder and several factors are known to contribute to its etiology. Evidence suggests that AD may be associated with a dysbiosis of the intestinal microbiota but whether is it is a cause, or an effect is yet to be understood.
Methods: In order to understand the effect of gut bacteria on the pathophysiology of AD, we used two approaches to manipulate gut bacteria in mice; probiotic and antibiotic therapies. Female wild type control C57BL/6 mice were compared to a mouse line that has the human Aβ sequence knocked into the mouse APP gene along with three disease causing mutations (AppNL-G-F). The animals at 2.5 months of age were randomly divided into five groups and three groups were treated with a cocktail of non-absorbable antibiotics (ABX) in drinking water and two groups were given water only for 10 days. Following ABX treatment, animals were either given normal diet, a probiotic (VSL#3) containing diet, or a diet containing both probiotic and prebiotic for 2 months. Upon completion, learning and memory was assessed along with brain Aβ and cytokines levels. GFAP and Iba-1 immunohistochemistry was performed on brain sections.
Results: AppNL-G-F mice showed a significant deficit in memory performance compared to controls. An increase in pro-inflammatory cytokines and Aβ along with increased GFAP and Iba-1 immunoreactivity was observed in AppNL-G-F brains. No change was observed in intestinal permeability across all groups. Two months of probiotic supplementation improved memory and decreased plaque load in AppNL-G-F mice. Consistent with this, probiotic feeding attenuated GFAP and Iba-1 immunoreactivity in AppNL-G-F brains. On other hand, antibiotic treatment alone or in combination with probiotic/prebiotics appeared to increase microgliosis and astrogliosis in AppNL-G-F brains.
Conclusion: These results demonstrate that transiently depleting gut bacteria with antibiotic treatment has a long-term effect on worsening AD-associated parameters in AppNL-G-F brains. On the other hand, modulating gut bacteria using probiotic feeding decreases plaque load and gliosis. This data suggests that gut microbiota modulate host brains and probiotics might be useful as supplementary therapy in AD.
Suba Nookala, Ph.D., "Synergistic effect of combined clindamycin and NLRP3 inflammasome inhibitor in a murine model of Group A Streptococcal Necrotizing Fasciitis"
Group A Streptococcus (GAS) is an important human pathogen that can cause a spectrum of diseases ranging from mild pharyngitis and impetigo to deeper, life-threatening invasive infections such as streptococcal toxic shock syndrome and necrotizing fasciitis (NF), a subgroup of Necrotizing Soft Tissue Infections (NSTI). An overarching pro-inflammatory cytokine storm and rapidly progressive tissue destruction lead to significant morbidity and lethal outcomes of GAS NF. Previously, we established that the pro-inflammatory mediator, IL-1β is a key upstream regulator of inflammatory pathways during GAS NSTI. Considering its pleiotropic role in host protection versus pro-inflammatory potential, the induction, expression, and secretion of biologically active IL1β are tightly regulated and are coordinated canonically by the activation of the NLRP3 inflammasome in a caspase 1-dependent manner. Despite extraordinary success by targeting IL-1β through IL-1rα (Rilonacept) and IL-1rN (receptor antagonist, Anakinra) in the treatment of other inflammatory conditions, these IL-1β blocking approaches disproportionately increase the risk for life-threatening and invasive GAS infections suggesting that blocking the IL-1β receptor is detrimental to the host. Therefore, we explored the possibility to target upstream of IL-1β secretion using a synthetic small molecular compound, MCC950, an inhibitor exhibiting selective activity against the NLRP3 inflammasome, as a logical therapeutic approach for controlling disease severity and outcomes of GAS NSTI. We studied the synergistic effect of MCC950 combined with current standard therapy, clindamycin (CLN) in our established mouse model of GAS NSTI. Preliminary findings show that while there were no differences in bacterial burden at the site of infection or dissemination into organs between the two treatments, compared to treatment with CLN alone, CLN MCC intervention resulted in significantly reduced skin lesions and plasma levels of the pro-inflammatory mediators, IL-1b, IL-6, and TNF-a as early as 24h post-infection, and IL-10 and IFN-g by 72h post-infection. Consistent with the significant reduction/absence of skin lesions, RT-qPCR analysis of skin lesions show that the mRNA transcripts of IL-1b, TNF-a, caspase 1, and P2XR were downregulated in CLN MCC treated mice. Further characterization of circulating myeloid-derived suppressor cells (MDSCs) by flow cytometry showed that the subset of CD11b+ Ly6cHigh monocytic-MDSCs was significantly elevated in mice treated with CLN MCC highlighting the emerging and important role of immunoprotective MDSCs in GAS NF. Although the molecular mechanisms of synergy are not yet understood, our findings highlight the utility of MCC950 for interrogating the role of NLRP3 inflammasome and IL-1b release during GAS NSTI, and may prove useful in the development of effective host-directed approaches that are amenable with existing therapeutic strategies to alleviate tissue destruction and disease severity during GAS NF.
Nicholas Smith, "Differential brain pathway activation in a non-anaphylactic mouse model of cow’s milk allergy"
Food allergy has been associated with various neuropsychiatric disorders, such as anxiety, attention deficit hyperactivity, and autism spectrum disorders, although, how the peripheral immune disorder affects mood and behavior is not well understood. To utilize this association for potential clinical treatments of behavioral disorders, a specific mechanism must be elucidated. We have previously demonstrated that sensitization of C57BL/6J mice to a milk allergen, beta-lactoglobulin (BLG; Bos d 5), results in anxiety-like behavior and gut dysbiosis without anaphylactic response to allergen challenge in male mice. We hypothesize that in response to food allergy, mice would express an increase in circulating cytokines and stimulate pathways relating to inflammation and glial cell function within the brain. To evaluate pathogenic influence of food allergy on brain function, we performed transcriptomic analysis of different regions of the brain in a mouse model of cow’s milk allergy. Male C57BL/6J mice were orally inoculated with a buffer containing 1 mg BLG with adjuvant, cholera toxin, or the adjuvant alone for 5-weeks to establish BLG- and sham-sensitized mice. All mice were challenged with 50 mg BLG in the 6th week and their clinical reactions were observed. Mice were euthanized the following day, and blood and various regions of the brains were harvested to perform ELISA for BLG-specific IgE and RNA sequencing for brain region-specific transcriptomics, respectively. Despite the lack of clinical symptoms upon BLG challenge in sensitized mice, increased serum BLG-specific IgE was observed, confirming their acquired immunity to the allergen. Increased plasma eotaxin-2, CCL9, and CXCL4 were also detected, while IL-6, CXCL5, and CCL5 were reduced. Transcriptomics analysis using the Ingenuity Pathway Analysis platform generated a small list of core regulators that are implicated in brain pathology including Apex1, Fmr1, Dio2, Slc16a2, Bdnf, Psen1, and Eomes. In the brain region that included the striatum, synaptogenesis, eNOS signaling, white adipose tissue browning, endocannabinoid neuron developing, and G protein αs pathways were differentially activated. In a region containing the thalamus and hypothalamus, netrin and Th17 signaling pathways were activated in BLG-sensitized mice, while the Fcε receptor and amyotrophic lateral sclerosis signaling pathways were highlighted in the midbrain region. Overall assessment of functional and disease pathways across brain regions from sensitized mice substantiated the regulation of movement, differentiation, and neurite formation of cells in the brain. Our results demonstrate allergic biomarkers without classical symptom presentation, despite the lack of symptoms mice had altered cytokine abundances and changes in pathway activation associated brain function. The results indicate neuroinflammation caused by allergy potentially via an IgE-Fcε mechanism. The resulting inflammation likely causes changes in number and structure of glia and neurons in addition to altering neuronal signaling causing differences in behavior.
Sattya Talukdar, "RSV does not induce EMT in airway epithelium"
Viral infections are known to induce Epithelial-mesenchymal transition (EMT). Many respiratory viruses, including influenza virus and rhinovirus, are involved in developing pulmonary fibrosis which is commonly observed in asthma, COPD, and cystic fibrosis. Respiratory Syncytial Virus (RSV) causes severe pneumonia and bronchiolitis and is responsible for enormous child mortality and morbidity. The host-RSV interaction is not clearly explored yet and it is still unclear whether RSV infection induces EMT in the infected airway epithelium. The present study was designed to evaluate the role of RSV infection in EMT induction in vitro. We used an immortalized lung epithelial cell line (A549 cells) and primary normal human bronchial epithelial (NHBE) cells in monolayer or air-liquid interface culture (ALI) culture. For determining EMT, we compared the expression of at least two EMT markers (E-cadherin and Vimentin) by western blotting and immunofluorescence. In contrast to transforming growth factor β (TGF-β)-mediated EMT, RSV did not induce EMT in A549 cells. Remarkably, RSV did not induce EMT in both NHBE monolayer and differentiated airway epithelium. In addition, RSV infection did not cause any impact on epithelial barrier integrity (confirmed by quantifying trans-epithelial electrical resistance using a volt-ohm meter) and ciliary function of infected cells (confirmed by quantifying ciliary beat frequency using high-speed video microscopy). Thus, our results suggest that RSV infection does not induce EMT in the airway epithelium.
Zhihan Wang, "Drug Repurposing for Various Courses of COVID-19 Based on Single-cell RNA Sequencing Data"
With more than 24 million people suffered from Coronavirus Disease 2019 (COVID-19), caused by novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), poses a critical threat to human health. There is no specific drug to date, highlighting the urgent need for rapid development of therapeutics for COVID-19. To identify potentially repurposable drugs, we employed a systematic approach to mine candidates from U.S. FDA approved drugs and pre-clinical small-molecule compounds by integrating the gene expression perturbation data by chemicals from the Library of Integrated Network-Based Cellular Signatures (LINCS) project with publicly available single-cell RNA sequencing dataset from mild and severe COVID-19 patients. We identified 281 FDA approved drugs that have the potential to be effective against SARS-CoV-2 infection, 10 of which are currently undergoing clinical trials to evaluate their efficacy against COVID-19. In conclusion, we have identified a list of repurposable anti-SARS-CoV-2 drugs using a systems biology approach. This project was supported by NIH P20GM113123 Pilot grant.