The effects of intraperitoneal metoprolol administration on healing of bone defects in rat tibia: a pilot study.

OBJECTIVES: Metoprolol is a cardioselective competitive beta-1 adrenergic receptor antagonist with antihypertensive properties, devoid of intrinsic sympathomimetic activity. Various studies have suggested the effect of beta-blockers on bone remodeling. We aimed to investigate whether metoprolol affects bone remodeling by altering anti-inflammatory and pro-inflammatory cytokines. MATERIALS AND METHODS: Surgical defects of 3 mm diameter were created in tibiae of 72 Sprague-Dawley rats. Rats were randomly assigned to a control group without metoprolol treatment (n = 36), and a test group treated with 0.1 mg/kg/day metoprolol (n = 36). Six rats from each group were sacrificed at days 0, 1, 3, 5, 7, and 14. The percentages of cells, which showed positive immunohistochemical staining for IL-1ß, IL-6, IL-10, and RANKL, were assessed in the defect area. Differences in percentages of stained cells within each of the test and control groups over various time intervals were tested using one-way ANOVA test. A P value of < 0.05 was considered statistically significant. RESULTS: No significant differences in IL-1ß, IL-10, IL-6, and RANKL expressions were found between test and control groups at the same interval. Significant reduction was observed at different time intervals in the same group (P < 0.05). CONCLUSION: Metoprolol did not reduce bone-active cytokine: IL-1ß, IL-6, and RANKL. It also did not elevate IL-10 expression levels. Thus, it does not appear to decrease osteoclastogenesis. CLINICAL RELEVANCE: Results from this animal model help us understand any effect of metoprolol on bone healing by potential contribution to different real-world clinical research.

Dysplasia severity is associated with poor quality of life in patients with Barrett's esophagus referred for endoscopic eradication therapy.

Limited data exist regarding patient-reported outcomes and quality of life (QOL) experienced by patients with Barrett's esophagus (BE) referred for endoscopic eradication therapy (EET). Specifically, the impact of grade of dysplasia has not been explored. The purpose of this study is to measure patient-reported symptoms and QOL and identify factors associated with poor QOL in BE patients referred for EET. This was a prospective multicenter study conducted from January 2015 to October 2017, which included patients with BE referred for EET. Participants completed a set of validated questionnaires to measure QOL, symptom severity, and psychosocial factors. The primary outcome was poor QOL defined by a PROMIS score >12. Multivariable logistic regression analysis was performed to identify factors associated with poor QOL. In total, 193 patients participated (mean age 64.6 years, BE length 5.5 cm, 82% males, 92% Caucasians) with poor QOL reported in 104 (53.9%) participants. On univariate analysis, patients with poor QOL had lower use of twice daily proton pump inhibitor use (61.5% vs. 86.5%, P = 0.03), shorter disease duration (4.9 vs. 5.9 years, P = 0.04) and progressive increase in grade of dysplasia (high-grade dysplasia: 68.8% vs. 31.3%, esophageal adenocarcinoma: 75.5% vs. 24.5%, P < 0.001). Multivariate analysis demonstrated that high-grade dysplasia was independently associated with poor QOL (OR: 5.57, 95% CI: 1.05, 29.5, P = 0.04). In summary, poor QOL is experienced by the majority of patients with BE referred for EET and the degree of dysplasia was independently associated with poor QOL, which emphasizes the need to incorporate patient-centered outcomes when studying treatment of BE-related dysplasia.

Effects of dietary butyrate supplementation on intestinal integrity of heat-stressed cockerels.

Butyrate modulates intestinal epithelial cell structure and function. Three hundred and sixty Lohmann LSL-Classic layer cockerels were used to investigate the effect of butyrate on heat stress-induced intestinal injury and intestinal integrity. The experiment was conducted from day 85 to 105 of age. The birds were divided randomly into three treatments: control, heat stress (HS), and heat stress provided with butyrate (HSB) at a level of 0.35 g/kg of diet. The control was reared at 21 ± 1 °C throughout the experiment. The HS and HSB treatments were exposed to a cyclic HS (35 ± 1 °C from 09:00 to 13:00 and 21 ± 1 °C from 13:00 to 09:00). Intestinal and mucosal weights, villus height, villus surface area (VSA), absorptive epithelial cell area and intestinal beneficial bacteria were lower in the HS treatment than in the other two treatments (p < 0.05). Heat-stressed cockerels exhibited the highest (p < 0.05) villi injury scores and serum endotoxin levels compared with the other treatments. Dietary inclusion of butyrate increased (p < 0.05) intestinal and mucosal weights, villus height, VSA, absorptive epithelial cell area and intestinal beneficial bacteria counts and reduced (p < 0.05) HS-induced injury in intestinal epithelia as well as intestinal permeability to endotoxin. In conclusion, dietary butyrate exerted protective effects against intestinal damage induced by HS and improved intestinal health and integrity.

Three-dimensional printing in surgery: a review of current surgical applications.

BACKGROUND: Three-dimensional printing (3DP) is gaining increasing recognition as a technique that will transform the landscape of surgical practice. It allows for the rapid conversion of anatomic images into physical objects, which are being used across a variety of surgical specialties. It has been unclear which groups are leading the way in coming up with novel ways of using the technology and what specifically the technology is being used for. The aim of this article was to review the current applications of 3DP in modern surgical practice. MATERIALS AND METHODS: An electronic search was carried out in MEDLINE, EMBASE, and PsycINFO for terms related to 3DP. These were then screened for relevance and practical applications of the technology in surgery. RESULTS: Four hundred eighty-eight articles were initially found, and these were eventually narrowed down to 93 full-text articles. It was determined that there were three main areas in which the technology is being used to print: (1) anatomic models, (2) surgical instruments, and (3) implants and prostheses. CONCLUSIONS: Different specialties are at different stages in the use of the technology. The costs involved with implementing the technology and time taken for printing are important factors to consider before widespread use. For the foreseeable future, this is an exciting and interesting technology with the capacity to radically change health care and revolutionize modern surgery.

Flow cytometric sexing of spider sperm reveals an equal sperm production ratio in a female-biased species.

Producing equal amounts of male and female offspring has long been considered an evolutionarily stable strategy. Nevertheless, exceptions to this general rule (i.e. male and female biases) are documented in many taxa, making sex allocation an important domain in current evolutionary biology research. Pinpointing the underlying mechanism of sex ratio bias is challenging owing to the multitude of potential sex ratio-biasing factors. In the dwarf spider, Oedothorax gibbosus, infection with the bacterial endosymbiont Wolbachia results in a female bias. However, pedigree analysis reveals that other factors influence sex ratio variation. In this paper, we investigate whether this additional variation can be explained by the unequal production of male- and female-determining sperm cells during sperm production. Using flow cytometry, we show that males produce equal amounts of male- and female-determining sperm cells; thus bias in sperm production does not contribute to the sex ratio bias observed in this species. This demonstrates that other factors such as parental genes suppressing endosymbiont effects and cryptic female choice might play a role in sex allocation in this species.

[Modeling T2 high severe asthma using human induced pluripotent stem cells (hiPSC)]. / Modélisation de l'asthme sévère par la technologie des cellules humaines souches pluripotentes induites (hiPSC).

Severe asthma patients with persistent airflow obstruction are characterized by functional obstruction due to mucus plugs containing mucins, fibrin, and eosinophil derived Charcot- Leyden crystals. The molecular mechanisms underlying this endotype are not clearly understood. Developing new models is crucial to respiratory research insofar as critical differences exist between human and rodent airway epithelium. We (and other teams) have shown that it is possible to reconstitute in vitro a complex and functional airway epithelium displaying all the features described in vivo from human-induced pluripotent stem cells (hiPSC). Our aim is to establish a human in vitro model of severe asthma that will recapitulate airway epithelium remodeling and mucus plugs.

The multi-faceted role of allergen exposure to the local airway mucosa.

Airway epithelial cells are the first to encounter aeroallergens and therefore have recently become an interesting target of many studies investigating their involvement in the modulation of allergic inflammatory responses. Disruption of a passive structural barrier composed of epithelial cells by intrinsic proteolytic activity of allergens may facilitate allergen penetration into local tissues and additionally affect chronic and ongoing inflammatory processes in respiratory tissues. Furthermore, the ability of rhinoviruses to disrupt and interfere with epithelial tight junctions may alter the barrier integrity and enable a passive passage of inhaled allergens through the airway epithelium. On the other hand, epithelial cells are no longer considered to act only as a physical barrier toward inhaled allergens, but also to actively contribute to airway inflammation by detecting and responding to environmental factors. Epithelial cells can produce mediators, which may affect the recruitment and activation of more specialized immune cells to the local tissue and also create a microenvironment in which these activated immune cells may function and propagate the inflammatory processes. This review presents the dual role of epithelium acting as a passive and active barrier when encountering an inhaled allergen and how this double role contributes to the start of local immune responses.

Contribution of regulatory T cells to alleviation of experimental allergic asthma after specific immunotherapy.

BACKGROUND: Allergen-specific immunotherapy (SIT) has been used since 1911, yet its mechanism of action remains to be elucidated. There is evidence indicating that CD4(+)FOXP3(+) regulatory T cells (Treg cells) are induced during SIT in allergic patients. However, the contribution of these cells to SIT has not been evaluated in vivo. OBJECTIVE: To evaluate the in vivo contribution of (i) CD4(+) CD25(+) T cells during SIT and of (ii) SIT-generated inducible FOXP3(+) Treg cells during allergen exposure to SIT-mediated suppression of asthmatic manifestations. METHODS: We used a mouse model of SIT based on the classical OVA-driven experimental asthma. Treg cells were quantified by flow cytometry 24 and 96 h post SIT treatment. We depleted CD4(+) CD25(+) T cells prior to SIT, and CD4(+)FOXP3(+) T cells prior to allergen challenges to study their contribution to the suppression of allergic manifestations by SIT treatment. RESULTS: Our data show that depletion of CD4(+)CD25(+) T cells at the time of SIT treatment reverses the suppression of airway hyperresponsiveness (AHR), but not of airway eosinophilia and specific IgE levels in serum. Interestingly, the number of CD4(+)CD25(+)FOXP3(+) T cells is transiently increased after SIT in the spleen and blood, suggesting the generation of inducible and presumably allergen-specific Treg cells during treatment. Depletion of CD4(+)FOXP3(+) Treg cells after SIT treatment partially reverses the SIT-induced suppression of airway eosinophilia, but not of AHR and serum levels of specific IgE. CONCLUSION AND CLINICAL RELEVANCE: We conclude that SIT-mediated tolerance induction towards AHR requires CD4(+)CD25(+) T cells at the time of allergen injections. In addition, SIT generates CD4(+)CD25(+)FOXP3(+) T cells that contribute to the suppression of airway eosinophilia upon allergen challenges. Therefore, enhancing Treg cell number or their activity during and after SIT could be of clinical relevance to improve the therapeutic effects of SIT.

Ursodeoxycholic acid suppresses eosinophilic airway inflammation by inhibiting the function of dendritic cells through the nuclear farnesoid X receptor.

BACKGROUND: Ursodeoxycholic acid (UDCA) is the only known beneficial bile acid with immunomodulatory properties. Ursodeoxycholic acid prevents eosinophilic degranulation and reduces eosinophil counts in primary biliary cirrhosis. It is unknown whether UDCA would also modulate eosinophilic inflammation outside the gastrointestinal (GI) tract, such as eosinophilic airway inflammation seen in asthma. The working mechanism for its immunomodulatory effect is unknown. METHODS: The immunosuppressive features of UDCA were studied in vivo, in mice, in an ovalbumin (OVA)-driven eosinophilic airway inflammation model. To study the mechanism of action of UDCA, we analyzed the effect of UDCA on eosinophils, T cells, and dendritic cell (DCs). DC function was studied in greater detail, focussing on migration and T-cell stimulatory strength in vivo and interaction with T cells in vitro as measured by time-lapse image analysis. Finally, we studied the capacity of UDCA to influence DC/T cell interaction. RESULTS: Ursodeoxycholic acid treatment of OVA-sensitized mice prior to OVA aerosol challenge significantly reduced eosinophilic airway inflammation compared with control animals. DCs expressed the farnesoid X receptor for UDCA. Ursodeoxycholic acid strongly promoted interleukin (IL)-12 production and enhanced the migration in DCs. The time of interaction between DCs and T cells was sharply reduced in vitro by UDCA treatment of the DCs resulting in a remarkable T-cell cytokine production. Ursodeoxycholic acid-treated DCs have less capacity than saline-treated DCs to induce eosinophilic inflammation in vivo in Balb/c mice. CONCLUSION: Ursodeoxycholic acid has the potency to suppress eosinophilic inflammation outside the GI tract. This potential comprises to alter critical function of DCs, in essence, the effect of UDCA on DCs through the modulation of the DC/T cell interaction.

The neuropeptide calcitonin gene-related peptide affects allergic airway inflammation by modulating dendritic cell function.

BACKGROUND: The neuropeptide calcitonin gene-related peptide (CGRP) is released in the lung by sensory nerves during allergic airway responses. Pulmonary dendritic cells (DC) orchestrating the allergic inflammation could be affected by CGRP. OBJECTIVE: To determine the immunomodulatory effects of CGRP on DC function and its impact on the induction of allergic airway inflammation. METHODS: CGRP receptor expression on lung DC was determined by RT-PCR and immunofluorescence staining. The functional consequences of CGRP receptor triggering were evaluated in vitro using bone marrow-derived DC. DC maturation and the induction of ovalbumin (OVA)-specific T cell responses were analysed by flow cytometry. The in vivo relevance of the observed DC modulation was assessed in a DC-transfer model of experimental asthma. Mice were sensitized by an intrapharyngeal transfer of OVA-pulsed DC and challenged with OVA aerosol. The impact of CGRP pretreatment of DC on airway inflammation was characterized by analysing differential cell counts and cytokines in bronchoalveolar lavage fluid (BALF), lung histology and cytokine responses in mediastinal lymph nodes. RESULTS: RT-PCR, immunofluorescence and cAMP assay demonstrated the expression of functionally active CGRP receptors in lung DC. RT-PCR revealed a transcriptional CGRP receptor down-regulation during airway inflammation. CGRP specifically inhibited the maturation of in vitro generated DC. Maturation was restored by blocking with the specific antagonist CGRP(8-37) . Consequently, CGRP-pretreated DC reduced the activation and proliferation of antigen-specific T cells and induced increased the numbers of T regulatory cells. The transfer of CGRP-pretreated DC diminished allergic airway inflammation in vivo, shown by reduced eosinophil numbers and increased levels of IL-10 in BALF. CONCLUSIONS AND CLINICAL RELEVANCE: CGRP inhibits DC maturation and allergen-specific T cell responses, which affects the outcome of the allergic airway inflammation in vivo. This suggests an additional mechanism by which nerve-derived mediators interfere with local immune responses. Thus, CGRP as an anti-inflammatory mediator could represent a new therapeutic tool in asthma therapy.

Dendritic cells and airway epithelial cells at the interface between innate and adaptive immune responses.

Because they can recognize and sample inhaled allergens, dendritic cells (DC) have been shown to be responsible for the initiation and maintenance of adaptive Th2 responses in asthma. It is increasingly clear that DC functions are strongly influenced by a crosstalk with neighboring cells like epithelial cells. Whereas the epithelium was initially considered only as a barrier, it is now seen as a central player in controlling the function of lung DCs through release of innate cytokines-promoting Th2 responses. Clinically relevant allergens, as well as known environmental and genetic risk factors for allergy and asthma, often interfere directly or indirectly with the innate immune functions of airway epithelial cells and DC. A better understanding of these interactions might lead to a better prevention and ultimately to new treatments for asthma.

Alternatively activated macrophages and impaired phagocytosis of S. aureus in chronic rhinosinusitis.

BACKGROUND: Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by biased Th2 inflammation and CRS without nasal polyps (CRSsNP) by a Th1 immune response. Colonization by Staphylococcus aureus is increased in CRSwNP. We aimed to determine macrophage phenotypes in nasal mucosa of CRSwNP and CRSsNP and to examine phagocytosis of S. aureus in these pathologies. METHODS: Macrophage phenotyping was performed by immunohistochemical staining on nasal mucosa sections from 28 patients; in addition flow cytometry analysis was performed. Tissue homogenate protein levels of IFN-γ, IL-5, IL-6, IL-1ß, TGF-ß, eosinophil cationic protein (ECP) and total IgE were analyzed and correlated with macrophage subtypes. Phagocytosis of S. aureus was analyzed by flow cytometry. Survival of S. aureus in Thp1 cells in the presence of polarizing cytokines was studied in vitro. RESULTS: By immunohistochemical analysis more M2 macrophages were present in CRSwNP than in CRSsNP. This also was positively correlated with increased levels of IL-5, ECP and locally produced IgE and decreased levels of IL-6, IL-1ß and IFN-γ. FACS analysis of dissociated nasal tissue confirmed the presence of increased numbers of M2 macrophages (CD206(+) HLADR(+) CD14(+) CD11c(+) CD20(-) ) in CRSwNP as compared to controls, while the number of M1 macrophages (CD206(-) HLADR(+) CD14(+) CD11c(int) CD16(-) CD20(-) ) was not different. Phagocytosis of S. aureus by human tissue derived macrophages was reduced in CRSwNP as compared to macrophages from the control inferior turbinates. CONCLUSIONS: Decreased phagocytosis of S. aureus and an M2 activation phenotype in CRSwNP could potentially contribute to persistence of chronic inflammation in CRSwNP.

MeDALL (Mechanisms of the Development of ALLergy): an integrated approach from phenotypes to systems medicine.

The origin of the epidemic of IgE-associated (allergic) diseases is unclear. MeDALL (Mechanisms of the Development of ALLergy), an FP7 European Union project (No. 264357), aims to generate novel knowledge on the mechanisms of initiation of allergy and to propose early diagnosis, prevention, and targets for therapy. A novel phenotype definition and an integrative translational approach are needed to understand how a network of molecular and environmental factors can lead to complex allergic diseases. A novel, stepwise, large-scale, and integrative approach will be led by a network of complementary experts in allergy, epidemiology, allergen biochemistry, immunology, molecular biology, epigenetics, functional genomics, bioinformatics, computational and systems biology. The following steps are proposed: (i) Identification of 'classical' and 'novel' phenotypes in existing birth cohorts; (ii) Building discovery of the relevant mechanisms in IgE-associated allergic diseases in existing longitudinal birth cohorts and Karelian children; (iii) Validation and redefinition of classical and novel phenotypes of IgE-associated allergic diseases; and (iv) Translational integration of systems biology outcomes into health care, including societal aspects. MeDALL will lead to: (i) A better understanding of allergic phenotypes, thus expanding current knowledge of the genomic and environmental determinants of allergic diseases in an integrative way; (ii) Novel diagnostic tools for the early diagnosis of allergy, targets for the development of novel treatment modalities, and prevention of allergic diseases; (iii) Improving the health of European citizens as well as increasing the competitiveness and boosting the innovative capacity of Europe, while addressing global health issues and ethical issues.

Effects of topically applied agents on intra-oral wound healing in a rat model: a clinical and histomorphometric study.

OBJECTIVES: Topically applied chlorhexidine and hyaluronan have many studies supporting their use to enhance oral wound healing. Allantoin is widely used topically to promote epithelial proliferation and wound healing, with very little scientific evidence to support such uses. This study investigated and compared the influence of these agents on the healing of intra-oral excisional wounds with large epithelial and connective tissue defects. METHODS: Excisional wounds, 3 mm in diameter, were made at the centre of the palate of 125 Wistar male albino rats. Five animals constituted the baseline group at time 0. The remaining animals were divided into four experimental and one control groups, in which chlorhexidine digluconate gel 0.2% (Perio.Kin®), hyaluronan gel (Gengigel®), allantoin 0.5% in vehicle gel, vehicle gel alone and nothing were applied daily to the wounds. The wound areas were measured photographically and the epithelialization rates were determined histologically at 0, 3, 7, 14 and 21 days post-surgery. RESULTS: The mean wound area and mean distance between the epithelial margins decreased significantly with time in all experimental and control groups (P < 0.05). A significant rate of wound area reduction was observed following the use of Perio.Kin® and Gengigel® at 7 and 14 days. Perio.Kin® showed a significant rate of wound epithelialization at 7 days. Allantoin did not positively or negatively affect wound healing. CONCLUSIONS: None of the tested agents had a negative effect on the rate of wound healing when applied on an excisional wound with epithelial and connective tissue defect. Positive results were achieved with Perio.Kin® and Gengigel®.

Blockade of CCR4 in a humanized model of asthma reveals a critical role for DC-derived CCL17 and CCL22 in attracting Th2 cells and inducing airway inflammation.

BACKGROUND: As Th2 type lymphocytes orchestrate the cardinal features of allergic asthma, inhibiting their recruitment to the lungs could be of therapeutic benefit. Although human Th2 cells express the CCR4 chemokine receptor and increased production of CCR4 ligands has been found in asthmatic airways, studies in animals have reached contradictory conclusions on whether blocking this pathway would be beneficial. OBJECTIVE: As a lack of efficacy might be due to differences between mouse and man, we readdressed this question using a humanized severe combined immunodeficiency model of asthma. METHODS: Mice received peripheral blood mononuclear cells from house dust mite (HDM) allergic asthmatic patients and then underwent bronchial challenge with HDM. RESULTS: This resulted in marked allergic inflammation and bronchial hyper-reactivity. Administration of CCR4 blocking antibody abolished the airway eosinophilia, goblet cell hyperplasia, IgE synthesis and bronchial hyper-reactivity. In this chimeric system, human CD11c+ dendritic cells (DCs) were the predominant source of CCR4 ligands, suggesting that DC-derived chemokines attract Th2 cells. In separate experiments using human DCs, in vitro exposure to HDM of DCs from HDM allergic patients but not healthy controls caused CCL17 and CCL22 release that resulted in chemoattraction of polarized human Th2 cells in a CCR4-dependent way. CONCLUSIONS: Taken together, our data provide proof of concept that CCR4 blockade inhibits the salient features of asthma and justify further clinical development of CCR4 antagonists for this disease.

Lung dendritic cells: targets for therapy in allergic disease.

Dendritic cells (DCs) are crucial in determining the functional outcome of allergen encounter in the lung. Antigen presentation by myeloid DCs leads to Th2 sensitization typical of allergic disease, whereas antigen presentation by plasmacytoid DCs serves to dampen inflammation. It is increasingly clear that DCs have an antigen presenting function beyond sensitization. DCs therefore constitute a novel target for the development of antiallergic therapy aimed at the origin of the inflammatory cascade. A careful study of DC biology and of the receptors expressed by lung DCs has provided a framework for the discovery of novel antiallergic compounds.