Influence of CX3CR1 Deletion on Cochlear Hair Cell Survival and Macrophage Expression in Chronic Suppurative Otitis Media.

OBJECTIVE: Our objective was to determine whether the receptor CX3CR1 is necessary for the recruitment of macrophages to the cochlea in chronic suppurative otitis media (CSOM) and if its deletion can prevent hair cell loss in CSOM. BACKGROUND: CSOM is a neglected disease that afflicts 330 million people worldwide and is the most common cause of permanent hearing loss among children in the developing world. It is characterized by a chronically discharging infected middle ear. We have previously demonstrated that CSOM causes macrophage associated sensory hearing loss. The receptor CX3CR1 is expressed on macrophages, which have been shown to be increased at the time point of outer hair cell (OHC) loss in CSOM. METHODS: In this report, we examine the influence of CX3CR1 deletion (CX3CR1-/-) in a validated model of Pseudomonas aeruginosa (PA) CSOM. RESULTS: The data show no difference in OHC loss between the CX3CR1-/- CSOM group and CX3CR1+/+ CSOM group (p = 0.28). We observed partial OHC loss in the cochlear basal turn, no OHC loss in the middle and apical turns in both CX3CR1-/- and CX3CR1+/+ CSOM mice at 14 days after bacterial inoculation. No inner hair cell (IHC) loss was found in all cochlear turns in all groups. We also counted F4/80 labeled macrophages in the spiral ganglion, spiral ligament, stria vascularis and spiral limbus of the basal, middle, and apical turn in cryosections. We did not find a significant difference in the total number of cochlear macrophages between CX3CR1-/- mice and CX3CR1+/+ mice (p = 0.97). CONCLUSION: The data did not support a role for CX3CR1 macrophage associated HC loss in CSOM.

Silencing Nrf2 attenuates chronic suppurative otitis media by inhibiting pro-inflammatory cytokine secretion through up-regulating TLR4.

Compromised TLR-mediated chronic inflammation contributes to bacterial infection-caused chronic suppurative otitis media, but the mechanisms are unclear. The present study examined the expression status of nuclear erythroid 2-related factor 2 (Nrf2) and TLRs in human middle-ear mucosae tissues collected from patients with chronic suppurative otitis media, chronic otitis media and non-otitis media, and found that Nrf2 was high-expressed, whereas TLR4, instead of other TLRs, was low expressed in chronic suppurative otitis media compared to chronic otitis media and non-chronic otitis media groups. Consistently, inflammatory cytokines were significantly up-regulated in the chronic suppurative otitis media group, instead of the chronic otitis media and non-chronic otitis media groups. Next, LPS-induced acute otitis media and chronic suppurative otitis media models in mice were established, and high levels of inflammatory cytokines were sustained in the mucosae tissues of chronic suppurative otitis media mice compared to the non-otitis media and acute otitis media groups. Interestingly, continuous low-dose LPS stimulation promoted Nrf2 expression, but decreased TLR4 levels in chronic suppurative otitis media mice mucosae. In addition, knock-down of Nrf2 increased TLR4 expression levels in chronic suppurative otitis media mice, and both Nrf2 ablation and TLR4 overexpression inhibited the pro-inflammatory cytokine expression in chronic suppurative otitis media. Finally, we found that both Nrf2 overexpression and TLR4 deficiency promoted chronic inflammation in LPS-induced acute otitis media mice models. Taken together, knock-down of Nrf2 reversed chronic inflammation to attenuate chronic suppurative otitis media by up-regulating TLR4.

[The influence of interleukin gene polymorphism on the serum cytokine level in the patients presenting with chonic suppurative otitis media]. / Vliianie polimorfizma genov interleikinov i faktora nekroza opukholei na uroven' tsitokinov v syvorotke krovi u bol'nykh khronicheskim gnoinym srednim otitom.

The objective of the present work was to study the influence of allelic variant associations of 1-beta interleukin (C3953T, &511C, T31C), interleukin-6 (C174G), and tumour necrosis factor-alpha (G308A) gene polymorphisms on the serum cytokine level in the patients presenting with chronic suppurative otitis media. A total of 299 patients at the age varying from 16 to 55 years with this condition divided into three groups were examined. Group 1 was comprised of 146 patients suffering from the tubotympanic form of chronic suppurative otitis media (CSOM). Group 2 was composed of 153 patients with epitympanic antral form of this condition. The control group included 183 subjects who have never suffered pathological changes in the middle ear. Human genomic DNA was analyzed with the use of the polymerase chain reaction (PCR). The serum cytokine levels were measured by the solid-state enzyme immunoassay in the beginning and at the end of the treatment period. The study has demonstrated that 56.2% of the healthy residents of the trans-Baikal region had the C/T Il-1b (C3953T) genotype. 79.1% of the patients presenting with the carious carious-destructive form of chronic suppurative otitis media were the heterozygous carriers of the T511C gene of 1-beta interleukin and had the maximally high concentrations of this interleukin in the blood serum. A rise in the production of the pro-inflammatory mediator (IL-6) was found to be related to the severity of the inflammatory process in the middle ear. The TNF-alpha content in the patients with CSOM during the active period of the disease proved to increase by a factor of 6 in comparison with that in the subjects of the control group irrespective of the type of mutation.

Mitochondrial DNA deletions in patients with chronic suppurative otitis media.

The aim of this study was to investigate the 4977 and 7400 bp deletions of mitochondrial DNA in patients with chronic suppurative otitis media and to indicate the possible association of mitochondrial DNA deletions with chronic suppurative otitis media. Thirty-six patients with chronic suppurative otitis media were randomly selected to assess the mitochondrial DNA deletions. Tympanomastoidectomy was applied for the treatment of chronic suppurative otitis media, and the curettage materials including middle ear tissues were collected. The 4977 and 7400 bp deletion regions and two control regions of mitochondrial DNA were assessed by using the four pair primers. DNA was extracted from middle ear tissues and peripheral blood samples of the patients, and then polymerase chain reactions (PCRs) were performed. PCR products were separated in 2 % agarose gel. Seventeen of 36 patients had the heterozygote 4977 bp deletion in the middle ear tissue but not in peripheral blood. There wasn't any patient who had the 7400 bp deletion in mtDNA of their middle ear tissue or peripheral blood tissue. The patients with the 4977 bp deletion had a longer duration of chronic suppurative otitis media and a higher level of hearing loss than the others (p < 0.01). Long time chronic suppurative otitis media and the reactive oxygen species can cause the mitochondrial DNA deletions and this may be a predisposing factor to sensorineural hearing loss in chronic suppurative otitis media. An antioxidant drug as a scavenger agent may be used in long-term chronic suppurative otitis media.

Polymorphisms in Toll-like receptors 2 and 4 genes and their expression in chronic suppurative otitis media.

OBJECTIVE: Toll-like receptors (TLRs) have a prominent role in inducing innate immune response. It has been suggested that regulation of TLRs is involved in the pathogenesis of chronic otitis media. TLR 2 and TLR 4 polymorphisms were connected with susceptibility to acute otitis and chronic otitis with effusion. The objective of this study was to establish expression of TLR 2 and 4 on middle ear mucosa in different types of chronic suppurative otitis media (CSOM), and the influence of gene polymorphisms TLR 2 Arg753Gln and TLR 4 Thr399Ile and Asp299Gly to susceptibility to CSOM. MATERIAL AND METHODS: Middle ear mucosa and full blood samples were obtained from 85 patients with chronic suppurative otitis media with and without cholesteatoma. Control group for mucosal TLR expression consisted of 71 samples of middle ear mucosa taken from patients with otosclerosis, and control group for DNA polymorphism consisted of 100 full blood samples in healthy subjects. DNA polymorphism detection was done with restriction fragment length polymorphism in RT PCR. Expression of TLR 2 and 4 was determined with immunohistochemical staining. RESULTS: TLR 2 and TLR 4 expression on the middle ear mucosa was not influenced by age of the patients with chronic otitis media. Incidence of TLR 2 Arg753Gln polymorphism was significantly higher in patients with chronic otitis media, compared to control group. Significant association between TLR 2 Arg753Gln polymorphism and different types of mucosal changes in patients with chronic otitis media was established. TLR 2 and 4 expression on experimental group mucosa was significantly different compared to control group, where there was no expression (p=0.000). Strong dependence of TLR 2 and TLR 4 expression on middle ear mucosa with different mucosal changes and immunohistochemical activity after staining was detected. CONCLUSION: Certain polymorphisms in TLR genes could be indicative for susceptibility to chronic otitis media. Expression of TLR 2 and 4 on middle ear mucosa was more dependable on different types of mucosal changes and type of CSOM than on bacteria found in the specimens. This can indicate that the type of mucosal changes are closely correlated with TLRs activity in middle ear.

Attenuated TLRs in middle ear mucosa contributes to susceptibility of chronic suppurative otitis media.

The variability in the recovery of otitis media (OM) is not well understood. Recent data have shown a critical role for toll-like receptors (TLRs) in inflammatory responses to bacteria. It remains unclear whether TLRs-mediated mucosal immunity plays a role in the OM recovery. The etiology, pathological profile, expression levels of TLR2, TLR4, TLR5, TLR9 and proinflammatory cytokines were measured in human middle-ear mucosae sampled from three subject groups: non-OM group, chronic otitis-media (COM) group, and chronic suppurative otitis-media (CSOM) group. Of the 72 ears, 86.11% CSOM patients were positive for bacteria. The cellular makeup of the middle ear mucosa differs among the three groups. Mucosae from the CSOM group presented chronic inflammation or suppurative inflammation in the rudimentary stroma, mainly with infiltration of monocytes and macrophages. The mRNA and protein levels of TLR2, TLR4, and TLR5 exhibited no difference between the non-OM and COM groups but were significantly lower in the CSOM group. Conversely, there was no significant difference in the TLR9 level among the three groups. Furthermore, proinflammatory cytokines TNF-α, IL-1ß, IFN-γ, IL-6 were up-regulated in the CSOM group. This study provides evidence that the variability in clinical otitis media recovery might be associated with the variability in the expression of mucosal TLRs. Reduced TLR levels in the middle-ear mucosa might cause weak host response to bacteria, persistent inflammation and susceptibility to CSOM.

Long-term follow-up of chronic suppurative otitis media in a high-risk children cohort.

OBJECTIVE: Chronic suppurative otitis media (CSOM) is the leading cause of mild to moderate hearing impairment in children worldwide and a major public health problem in many indigenous populations. There is a lack of basic epidemiological facts and knowledge on the development of CSOM, as the disease primarily affects developing countries where research capacities often are limited. The purpose of this study was to determine the long-term outcome of CSOM in a high-risk population and to identify risk factors. METHODS: Follow-up study (2008) on a population-based cohort of 465 children in Greenland, initially examined (1996-8) between the ages 0 and 4 years. Follow-up was attempted among 307 children living in the two major towns. Binomial logistic regression analysis was made to identify risk factors for developing CSOM and for maintaining disease in to adolescence (odds ratios). Log linear binomial regression was used to estimate risk ratios and absolute risks. RESULTS: At follow-up 236 participated (77% of those available). The prevalence of CSOM was 32/236 (14%) at age group 0-4 years and 21/236 (9%) at age group 11-15 years. Thirteen had disease debut after the initial study. Of those with CSOM in the initial study 24/32 (75%) healed spontaneously. Risk factors for the development of CSOM at any time in childhood was the mother's history of CSOM OR 2.55 (95% CI 1.14-5.70; p=0.02), and mothers with low levels of schooling OR 1.57 (1.03-2.40; p=0.04). Once CSOM had developed boys were more likely to have persistent disease OR 5.46 (95% CI 1.47-20.37; p=0.01). The absolute risk of CSOM if the mother had both a history of CSOM and low schooling was for boys 45.4% (95% CI 26.5-77.7) and for girls 30.7% (95% CI 17.8-53.10). The cumulative risk of CSOM was 19% at follow-up. CONCLUSIONS: Even though a large number of CSOM cases seemed to heal spontaneously, the prevalence of untreated CSOM among school-age children in Greenland remained high as new cases were found at follow-up. Increased focus on prevention and identification of children at special risk could reduce the high prevalence of CSOM.

Otorhinolaringologic manifestation of Smith-Magenis syndrome.

Smith-Magenis syndrome (SMS) is a multiple congenital anomaly/mental retardation (MCA/MR) syndrome link to a contiguous-gene deletion syndrome, involving chromosome 1 7p 11.2,whose incidence is estimated to be 1:25,000 livebirth. SMS is characterised by a specific physical, behavioural and developmental pattern. The main clinical features consist of a broad flat midface with brachycefaly, broad nasal bridge, brachydactily, speech delay, hoarse deep voice and peripheral neuropathy. Behavioural abnormalities include hypermotility, self-mutilation and sleep disturbance. This report defines the otorhinolaryngological aspects of a new case of SMS, confirmed by cytogenetic-molecular analysis, in a 9 year old girl affected by chronic otitis media, deafness and sinusitis, who presented with typical clinical signs and symptoms.

p73-deficient mice have neurological, pheromonal and inflammatory defects but lack spontaneous tumours.

p73 (ref. 1) has high homology with the tumour suppressor p53 (refs 2-4), as well as with p63, a gene implicated in the maintenance of epithelial stem cells. Despite the localization of the p73 gene to chromosome 1p36.3, a region of frequent aberration in a wide range of human cancers, and the ability of p73 to transactivate p53 target genes, it is unclear whether p73 functions as a tumour suppressor. Here we show that mice functionally deficient for all p73 isoforms exhibit profound defects, including hippocampal dysgenesis, hydrocephalus, chronic infections and inflammation, as well as abnormalities in pheromone sensory pathways. In contrast to p53-deficient mice, however, those lacking p73 show no increased susceptibility to spontaneous tumorigenesis. We report the mechanistic basis of the hippocampal dysgenesis and the loss of pheromone responses, and show that new, potentially dominant-negative, p73 variants are the predominant expression products of this gene in developing and adult tissues. Our data suggest that there is a marked divergence in the physiological functions of the p53 family members, and reveal unique roles for p73 in neurogenesis, sensory pathways and homeostatic control.

[Blood group-dependent efficiency of immunological correction in patients with chronic purulent otitis media]. / Zavisimost' éffektivnosti immunokorrektsii ot grupp krovi u bol'nykh khronicheskim gnoinym srednim otitom.

Clinical and immunological investigations were performed in 300 patients with otitis media chronica purulenta. The patients, aged 16-65 years, received immunocorrectors in addition to basic postoperative therapy consisting of antibiotics and antihistamine drugs. The highest immunocorrective effect was registered in patients with blood group AI, the weakest in those with blood group AB(IV). Blood group B(III) and A(II) were the second and the third.