Immunoglobulin A coating identifies colitogenic bacteria in inflammatory bowel disease.

Specific members of the intestinal microbiota dramatically affect inflammatory bowel disease (IBD) in mice. In humans, however, identifying bacteria that preferentially affect disease susceptibility and severity remains a major challenge. Here, we used flow-cytometry-based bacterial cell sorting and 16S sequencing to characterize taxa-specific coating of the intestinal microbiota with immunoglobulin A (IgA-SEQ) and show that high IgA coating uniquely identifies colitogenic intestinal bacteria in a mouse model of microbiota-driven colitis. We then used IgA-SEQ and extensive anaerobic culturing of fecal bacteria from IBD patients to create personalized disease-associated gut microbiota culture collections with predefined levels of IgA coating. Using these collections, we found that intestinal bacteria selected on the basis of high coating with IgA conferred dramatic susceptibility to colitis in germ-free mice. Thus, our studies suggest that IgA coating identifies inflammatory commensals that preferentially drive intestinal disease. Targeted elimination of such bacteria may reduce, reverse, or even prevent disease development.

The Wnt Antagonist Dickkopf-1 Promotes Pathological Type 2 Cell-Mediated Inflammation.

Exposure to a plethora of environmental challenges commonly triggers pathological type 2 cell-mediated inflammation. Here we report the pathological role of the Wnt antagonist Dickkopf-1 (Dkk-1) upon allergen challenge or non-healing parasitic infection. The increased circulating amounts of Dkk-1 polarized T cells to T helper 2 (Th2) cells, stimulating a marked simultaneous induction of the transcription factors c-Maf and Gata-3, mediated by the kinases p38 MAPK and SGK-1, resulting in Th2 cell cytokine production. Circulating Dkk-1 was primarily from platelets, and the increase of Dkk-1 resulted in formation of leukocyte-platelet aggregates (LPA) that facilitated leukocyte infiltration to the affected tissue. Functional inhibition of Dkk-1 impaired Th2 cell cytokine production and leukocyte infiltration, protecting mice from house dust mite (HDM)-induced asthma or Leishmania major infection. These results highlight that Dkk-1 from thrombocytes is an important regulator of leukocyte infiltration and polarization of immune responses in pathological type 2 cell-mediated inflammation.

Th9 Cells Drive Host Immunity against Gastrointestinal Worm Infection.

Type 2 inflammatory cytokines, including interleukin-4 (IL-4), IL-5, IL-9, and IL-13, drive the characteristic features of immunity against parasitic worms and allergens. Whether IL-9 serves an essential role in the initiation of host-protective responses is controversial, and the importance of IL-9- versus IL-4-producing CD4⁺ effector T cells in type 2 immunity is incompletely defined. Herein, we generated IL-9-deficient and IL-9-fluorescent reporter mice that demonstrated an essential role for this cytokine in the early type 2 immunity against Nippostrongylus brasiliensis. Whereas T helper 9 (Th9) cells and type 2 innate lymphoid cells (ILC2s) were major sources of infection-induced IL-9 production, the adoptive transfer of Th9 cells, but not Th2 cells, caused rapid worm expulsion, marked basophilia, and increased mast cell numbers in Rag2-deficient hosts. Taken together, our data show a critical and nonredundant role for Th9 cells and IL-9 in host-protective type 2 immunity against parasitic worm infection.

T regulatory cells maintain intestinal homeostasis by suppressing γδ T cells.

Immune tolerance against enteric commensal bacteria is important for preventing intestinal inflammation. Deletion of phosphoinositide-dependent protein kinase 1 (Pdk1) in T cells via Cd4-Cre induced chronic inflammation of the intestine despite the importance of PDK1 in T cell activation. Analysis of colonic intraepithelial lymphocytes of PDK1-deficient mice revealed markedly increased CD8α(+) T cell receptor (TCR)γδ(+) T cells, including an interleukin-17 (IL-17)-expressing population. TCRγδ(+) T cells were responsible for the inflammatory colitis as shown by the fact that deletion of Tcrd abolished spontaneous colitis in the PDK1-deficient mice. This dysregulation of intestinal TCRγδ(+) T cells was attributable to a reduction in the number and functional capacity of PDK1-deficient T regulatory (Treg) cells. Adoptive transfer of wild-type Treg cells abrogated the spontaneous activation and proliferation of intestinal TCRγδ(+) T cells observed in PDK1-deficient mice and prevented the development of colitis. Therefore, suppression of intestinal TCRγδ(+) T cells by Treg cells maintains enteric immune tolerance.

Inflammasome-mediated dysbiosis regulates progression of NAFLD and obesity.

Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of metabolic syndrome and the leading cause of chronic liver disease in the Western world. Twenty per cent of NAFLD individuals develop chronic hepatic inflammation (non-alcoholic steatohepatitis, NASH) associated with cirrhosis, portal hypertension and hepatocellular carcinoma, yet the causes of progression from NAFLD to NASH remain obscure. Here, we show that the NLRP6 and NLRP3 inflammasomes and the effector protein IL-18 negatively regulate NAFLD/NASH progression, as well as multiple aspects of metabolic syndrome via modulation of the gut microbiota. Different mouse models reveal that inflammasome-deficiency-associated changes in the configuration of the gut microbiota are associated with exacerbated hepatic steatosis and inflammation through influx of TLR4 and TLR9 agonists into the portal circulation, leading to enhanced hepatic tumour-necrosis factor (TNF)-α expression that drives NASH progression. Furthermore, co-housing of inflammasome-deficient mice with wild-type mice results in exacerbation of hepatic steatosis and obesity. Thus, altered interactions between the gut microbiota and the host, produced by defective NLRP3 and NLRP6 inflammasome sensing, may govern the rate of progression of multiple metabolic syndrome-associated abnormalities, highlighting the central role of the microbiota in the pathogenesis of heretofore seemingly unrelated systemic auto-inflammatory and metabolic disorders.

Membrane-bound Dickkopf-1 in Foxp3+ regulatory T cells suppresses T-cell-mediated autoimmune colitis.

Induction of tolerance is a key mechanism to maintain or to restore immunological homeostasis. Here we show that Foxp3+ regulatory T (Treg) cells use Dickkopf-1 (DKK-1) to regulate T-cell-mediated tolerance in the T-cell-mediated autoimmune colitis model. Treg cells from DKK-1 hypomorphic doubleridge mice failed to control CD4+ T-cell proliferation, resulting in CD4 T-cell-mediated autoimmune colitis. Thymus-derived Treg cells showed a robust expression of DKK-1 but not in naive or effector CD4 T cells. DKK-1 expression in Foxp3+ Treg cells was further increased upon T-cell receptor stimulation in vitro and in vivo. Interestingly, Foxp3+ Treg cells expressed DKK-1 in the cell membrane and the functional inhibition of DKK-1 using DKK-1 monoclonal antibody abrogated the suppressor function of Foxp3+ Treg cells. DKK-1 expression was dependent on de novo protein synthesis and regulated by the mitogen-activated protein kinase pathway but not by the canonical Wnt pathway. Taken together, our results highlight membrane-bound DKK-1 as a novel Treg-derived mediator to maintain immunological tolerance in T-cell-mediated autoimmune colitis.

The microbial metabolite butyrate regulates intestinal macrophage function via histone deacetylase inhibition.

Given the trillions of microbes that inhabit the mammalian intestines, the host immune system must constantly maintain a balance between tolerance to commensals and immunity against pathogens to avoid unnecessary immune responses against otherwise harmless bacteria. Misregulated responses can lead to inflammatory bowel diseases such as ulcerative colitis or Crohn's disease. The mechanisms by which the immune system maintains this critical balance remain largely undefined. Here, we demonstrate that the short-chain fatty acid n-butyrate, which is secreted in high amounts by commensal bacteria, can modulate the function of intestinal macrophages, the most abundant immune cell type in the lamina propria. Treatment of macrophages with n-butyrate led to the down-regulation of lipopolysaccharide-induced proinflammatory mediators, including nitric oxide, IL-6, and IL-12, but did not affect levels of TNF-α or MCP-1. These effects were independent of toll-like receptor signaling and activation of G-protein-coupled receptors, two pathways that could be affected by short-chain fatty acids. In this study, we provide several lines of evidence that suggest that these effects are due to the inhibition of histone deacetylases by n-butyrate. These findings elucidate a pathway in which the host may maintain tolerance to intestinal microbiota by rendering lamina propria macrophages hyporesponsive to commensal bacteria through the down-regulation of proinflammatory effectors.

Microbiota-induced activation of epithelial IL-6 signaling links inflammasome-driven inflammation with transmissible cancer.

The microbiota is pivotal in the pathogenesis of inflammatory bowel disease (IBD)-associated inflammation-induced colorectal cancer (CRC), yet mechanisms for these effects remain poorly characterized. Here, we demonstrate that aberrant inflammasome-induced microbiota plays a critical role in CRC development, where mice deficient in the NOD-like receptor family pyrin domain containing 6 (NLRP6) inflammasome feature enhanced inflammation-induced CRC formation. Intriguingly, WT mice cohoused either with inflammasome-deficient mice or with mice lacking IL-18 feature exacerbated inflammation-induced CRC compared with singly housed WT mice. Enhanced tumorigenesis is dependent on microbiota-induced chemokine (C-C motif) ligand 5 (CCL5)-driven inflammation, which in turn promotes epithelial cell proliferation through local activation of the IL-6 pathway, leading to cancer formation. Altogether, our results mechanistically link the altered microbiota with the pathogenesis of inflammation-induced CRC and suggest that in some conditions, microbiota components may transfer CRC susceptibility between individuals.

IL-22 deficiency alters colonic microbiota to be transmissible and colitogenic.

IL-22 is a good candidate to play a critical role in regulating gut microbiota because it is an important inducer of antimicrobial peptides and mucins in the gut. However, whether IL-22 participates in immune homeostasis by way of modulating gut microbiota remains to be elucidated. In this study, we find, through 16S rRNA gene-pyrosequencing analysis, that healthy IL-22-deficient mice had altered colonic microbiota, notably with decreased abundance of some genera, including Lactobacillus, and increased levels of others. Mice harboring this altered microbiota exhibited more severe disease during experimentally induced colitis. Interestingly, this altered gut microbiota can be transmitted to cohoused wild-type animals along with the increased susceptibility to this colitis, indicating an important role for IL-22 in shaping the homeostatic balance between immunity and colonic microbiota for host health.

Ablation of IL-17A abrogates progression of spontaneous intestinal tumorigenesis.

The intrinsic role of endogenous IL-17A in spontaneous intestinal tumorigenesis has not been addressed previously to our knowledge. Ablation of IL-17A significantly reduced tumor development in mice bearing a heterozygote mutation in the adenomatous polyposis coli (APC) gene (Apc(Min/+) mice). There was also a decrease in inflammatory cytokines and proinflammatory mediators, reduced infiltration of lymphocytes including T cells, and preservation of intestinal architecture and the presence of APC protein in intestinal epithelial cells. Interestingly, IL-17A ablation also corrected immunological abnormalities such as splenomegaly and thymic atrophy in Apc(Min/+) mice. CD4 T cells from Apc(Min/+) mice showed hyperproliferative potential in vitro and in vivo and increased levels of IL-17A and IL-10. The effector CD4 T cells from Apc(Min/+) mice were more resistant to regulatory T cell-mediated suppression. Finally, these CD4 T cells induced colitis in immunodeficient mice upon adoptive transfer, whereas the ablation of IL-17A in CD4 T cells in Apc(Min/+) mice completely abolished this pathogenic potential in vivo. Taken together, our results show that CD4 T cell-derived IL-17A promotes spontaneous intestinal tumorigenesis with altered functions of CD4 T cells in Apc(Min/+) mice.

Induction and rescue of Nod2-dependent Th1-driven granulomatous inflammation of the ileum.

Mutations in the NOD2 gene are strong genetic risk factors for ileal Crohn's disease. However, the mechanism by which these mutations predispose to intestinal inflammation remains a subject of controversy. We report that Nod2-deficient mice inoculated with Helicobacter hepaticus, an opportunistic pathogenic bacterium, developed granulomatous inflammation of the ileum, characterized by an increased expression of Th1-related genes and inflammatory cytokines. The Peyer's patches and mesenteric lymph nodes were markedly enlarged with expansion of IFN-gamma-producing CD4 and CD8 T cells. Rip2-deficient mice exhibited a similar phenotype, suggesting that Nod2 function likely depends on the Rip2 kinase in this model. Transferring wild-type bone marrow cells into irradiated Nod2-deficient mice did not rescue the phenotype. However, restoring crypt antimicrobial function of Nod2-deficient mice by transgenic expression of alpha-defensin in Paneth cells rescued the Th1 inflammatory phenotype. Therefore, through the regulation of intestinal microbes, Nod2 function in nonhematopoietic cells of the small intestinal crypts is critical for protecting mice from a Th1-driven granulomatous inflammation in the ileum. The model may provide insight into Nod2 function relevant to inflammation of ileal Crohn's disease.

Negative regulation of Toll-like receptor signaling plays an essential role in homeostasis of the intestine.

A healthy intestinal tract is characterized by controlled homeostasis due to the balanced interaction between commensal bacteria and the host mucosal immune system. Human and animal model studies have supported the hypothesis that breakdown of this homeostasis may underlie the pathogenesis of inflammatory bowel diseases. However, it is not well understood how intestinal microflora stimulate the intestinal mucosal immune system and how such activation is regulated. Using a spontaneous, commensal bacteria-dependent colitis model in IL-10-deficient mice, we investigated the role of TLR and their negative regulation in intestinal homeostasis. In addition to IL-10(-/-) MyD88(-/-) mice, IL-10(-/-) TLR4(-/-) mice exhibited reduced colitis compared to IL-10(-/-) mice, indicating that TLR4 signaling plays an important role in inducing colitis. Interestingly, the expression of IRAK-M, a negative regulator of TLR signaling, is dependent on intestinal commensal flora, as IRAK-M expression was reduced in mice re-derived into a germ-free environment, and introduction of commensal bacteria into germ-free mice induced IRAK-M expression. IL-10(-/-) IRAK-M(-/-) mice exhibited exacerbated colitis with increased inflammatory cytokine gene expression. Therefore, this study indicates that intestinal microflora stimulate the colitogenic immune system through TLR and negative regulation of TLR signaling is essential in maintaining intestinal homeostasis.

Interspecies commensal interactions have nonlinear impacts on host immunity.

The impacts of individual commensal microbes on immunity and disease can differ dramatically depending on the surrounding microbial context; however, the specific bacterial combinations that dictate divergent immunological outcomes remain largely undefined. Here, we characterize an immunostimulatory Allobaculum species from an inflammatory bowel disease patient that exacerbates colitis in gnotobiotic mice. Allobaculum inversely associates with the taxonomically divergent immunostimulatory species Akkermansia muciniphila in human-microbiota-associated mice and human cohorts. Co-colonization with A. muciniphila ameliorates Allobaculum-induced intestinal epithelial cell activation and colitis in mice, whereas Allobaculum blunts the A.muciniphila-specific systemic antibody response and reprograms the immunological milieu in mesenteric lymph nodes by blocking A.muciniphila-induced dendritic cell activation and T cell expansion. These studies thus identify a pairwise reciprocal interaction between human gut bacteria that dictates divergent immunological outcomes. Furthermore, they establish a generalizable framework to define the contextual cues contributing to the "incomplete penetrance" of microbial impacts on human disease.

Research Progress of Mesenchymal Stem Cell Therapy for Severe COVID-19.

Corona virus disease 2019 (COVID-19) refers to a type of pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Sixty million confirmed cases have been reported worldwide until November 29, 2020. Unfortunately, the novel coronavirus is extremely contagious and the mortality rate of severe and critically ill patients is high. Thus, there is no definite and effective treatment in clinical practice except for antiviral therapy and supportive therapy. Mesenchymal stem cells (MSCs) are not only characterized by low immunogenicity and homing but also have anti-inflammatory and immunomodulation characteristics. Furthermore, they can inhibit the occurrence and development of a cytokine storm, inhibit lung injury, and exert antipulmonary fibrosis and antioxidative stress, therefore MSC therapy is expected to become one of the effective therapies to treat severe COVID-19. This article will review the possible mechanisms of MSCs in the treatment of severe COVID-19.

Novel assisted cough system based on simulating cough airflow dynamics.

ABSTRACT: Cough is a defensive behavior that protects the respiratory system from infection and clears airway secretions. Cough airflow dynamics have been analyzed by a variety of mathematical and experimental tools. In this paper, the cough airflow dynamics of 42 subjects were obtained and analyzed. An identification model based on piecewise Gauss function for cough airflow dynamics is proposed through the dimensionless method, which could achieve over 90% identification accuracy. Meanwhile, an assisted cough system based on pneumatic flow servo system is presented. The vacuum situation and feedback control are used to increase the simulated peak cough flow rate, which are important for airway secretion clearance and to avoid airway collapse, respectively. The simulated cough peak flow could reach 5 L/s without the external assistance such as manual pressing, patient cooperation and other means. Finally, the backstepping control is developed to generate a simulated cough airflow that closely mimics the natural cough airflow of humans. The assisted cough system opens up wide opportunities of practical application in airway secretion clearance for critically ill patients with COVID 2019 and other pulmonary diseases.

Mechanical ventilation strategy for pulmonary rehabilitation based on patient-ventilator interaction.

Mechanical ventilation is an effective medical means in the treatment of patients with critically ill, COVID-19 and other pulmonary diseases. During the mechanical ventilation and the weaning process, the conduct of pulmonary rehabilitation is essential for the patients to improve the spontaneous breathing ability and to avoid the weakness of respiratory muscles and other pulmonary functional trauma. However, inappropriate mechanical ventilation strategies for pulmonary rehabilitation often result in weaning difficulties and other ventilator complications. In this article, the mechanical ventilation strategies for pulmonary rehabilitation are studied based on the analysis of patient-ventilator interaction. A pneumatic model of the mechanical ventilation system is established to determine the mathematical relationship among the pressure, the volumetric flow, and the tidal volume. Each ventilation cycle is divided into four phases according to the different respiratory characteristics of patients, namely, the triggering phase, the inhalation phase, the switching phase, and the exhalation phase. The control parameters of the ventilator are adjusted by analyzing the interaction between the patient and the ventilator at different phases. A novel fuzzy control method of the ventilator support pressure is proposed in the pressure support ventilation mode. According to the fuzzy rules in this research, the plateau pressure can be obtained by the trigger sensitivity and the patient's inspiratory effort. An experiment prototype of the ventilator is established to verify the accuracy of the pneumatic model and the validity of the mechanical ventilation strategies proposed in this article. In addition, through the discussion of the patient-ventilator asynchrony, the strategies for mechanical ventilation can be adjusted accordingly. The results of this research are meaningful for the clinical operation of mechanical ventilation. Besides, these results provide a theoretical basis for the future research on the intelligent control of ventilator and the automation of weaning process.

Genetic repression of mouse VEGF expression regulates coagulation cascade.

VEGF is an important growth factor in embryonic development and functional maintenance of adult organs. Single allele inactivation of VEGF leads to embryonic lethality and many conditional disruptions show dramatic phenotypes or reduced viabilities. To overcome this burden, a reversible system, which was used in the regulation of VEGF expression and functional studies of many tissues and organs in adult, was developed. VEGF is known for expression in many tissues and high expression in platelets. Repression of VEGF expression showed significant defect in bleeding time and clotting time. In mouse model of LPS-induced blood hypercoagulation, significant reduction of plasma thrombin-antithrombin formation was observed in VEGF repressed mice, and tissues expressed different modes of damages. These results indicate that VEGF regulates coagulation cascade and may affect sepsis. This is the first genetic evidence of direct link between VEGF transcription and coagulation cascade. Studies using this system may help us to find out how VEGF regulates coagulation cascade and provide new strategies in treating coagulation related diseases.

Efficacy of the combination of minimally invasive CO2 laser incision with photodynamic therapy for infected epidermoid cysts.

BACKGROUND: There are three main techniques for the removal of epidermoid cysts: traditional wide excision, minimal excision, and punch biopsy excision. For inflamed cysts, the wall is more friable and, therefore, more difficult to remove completely. The classic surgical excision always leads to a long scar or high rate of recurrence. CO2 laser has been proven to result in minimal incision, less bleeding, no suture, and a smaller or no scar. Photodynamic therapy (PDT) has been proposed as an antimicrobial alternative for common and drug-resistant bacteria in nonspecific and multiple sites. It was also shown to be effective in accelerating healing and inhibiting excessive proliferation of hyperplastic scar. Thus, we combined minimally invasive CO2 laser incision with PDT for epidermoid cysts with infection. METHODS: Thirty-three patients had a total of 39 infectious cysts. Two of the patients withdrew due to the high cost after 1 treatment session. After local injection of anesthesia, a hole measuring 2-3 mm was made at the pore in the upper part of the cyst along skin texture by CO2 laser (power 5 W, surgical pattern). The contents of the cyst were extracted through the hole using a curette and compression with gauze. PDT was then performed immediately. A total of 3 PDT sessions were recommended. The overall clinical effects, recurrence rates, cosmetic outcomes, adverse events, and patient satisfaction were assessed. RESULTS: We achieved a 97% success rate in 31 patients with 34 lesions using a combination of minimally invasive CO2 laser incision with PDT. At the 6- to 12-month follow-up, 30 of the patients had excellent cosmetic outcomes and satisfactory therapeutic effect. Pain during the illumination process, which can be relieved by dynamic cold air, was the primary adverse event. CONCLUSION: Our results demonstrate promise for the combination of minimally invasive CO2 laser incision with PDT as a safe and effective therapy for epidermoid cysts with infection. This treatment can inactivate a wide range of microbes including gram-positive and -negative bacteria, without developing drug resistance. Furthermore, it can promote fast wound healing and reduce scar formation.

Numerical Analysis of Airway Mucus Clearance Effectiveness Using Assisted Coughing Techniques.

Cough is a protective respiratory reflex used to clear respiratory airway mucus. For patients with cough weakness, such as chronic obstructive pulmonary disease, neuromuscular weakness disease and other respiratory diseases, assisted coughing techniques are essential to help them clear mucus. In this study, the Eulerian wall film model was applied to simulate the coughing clearance process through a computational fluid dynamics methodology. Airway generation 0 to generation 2 based on realistic geometry is considered in this study. To quantify cough effectiveness, cough efficiency was calculated. Moreover, simulations of four different coughing techniques applied for chronic obstructive pulmonary disease and neuromuscular weakness disease were conducted. The influences of mucus film thickness and mucus viscosity on cough efficiency were analyzed. From the simulation results, we found that with increasing mucus film thickness and decreasing mucus viscosity, cough efficiency improved accordingly. Assisted coughing technologies have little influence on the mucus clearance of chronic obstructive pulmonary disease models. Finally, it was observed that the cough efficiency of the mechanical insufflation-exsufflation technique (MIE) is more than 40 times the value of an unassisted coughing technique, which indicates that the MIE technology has a great effect on airway mucus clearance for neuromuscular weakness disease models.

Satisfactory response of a back carbuncle to 5-aminolevulinic acid (ALA) photodynamic therapy: A case report.

Carbuncle, a collection of interconnected furuncles with multiple pustular openings, is usually caused by Staphylococcus aureus (S. aureus). In this condition, both skin and subcutaneous tissue of the lesion show severe inflammation. It often occurs in immunocompromized patients such as those with diabetes, nephritis, malnutrition, heart failure, hypogammaglobulinemia, exfoliative dermatitis, or pemphigus or those using corticosteroids for long-term. Antibiotics and aggressive debridement are the primary recommended treatments for carbuncle. We report a case of carbuncle that received satisfactory response, in which the inflammation subsided and the wound healed after the administration of ALA photodynamic therapy for three times.