Neuronal MeCP2 in the dentate gyrus regulates mossy fiber sprouting of mice with temporal lobe epilepsy.

Sprouting of mossy fibers, one of the most consistent findings in tissue from patients with mesial temporal lobe epilepsy, exhibits several uncommon axonal growth features and has been considered a paradigmatic example of circuit plasticity that occurs in the adult brain. Clarifying the mechanisms responsible may provide new insight into epileptogenesis as well as axon misguidance in the central nervous system. Methyl-CpG-binding protein 2 (MeCP2) binds to methylated genomic DNA to regulate a range of physiological functions implicated in neuronal development and adult synaptic plasticity. However, exploring the potential role of MeCP2 in the documented misguidance of axons in the dentate gyrus has not yet been attempted. In this study, a status epilepticus-induced decrease of neuronal MeCP2 was observed in the dentate gyrus (DG). An essential regulatory role of MeCP2 in the development of functional mossy fiber sprouting (MFS) was confirmed through stereotaxic injection of a recombinant adeno-associated virus (AAV) to up- or down-regulate MeCP2 in the dentate neurons. Chromatin immunoprecipitation sequencing (ChIP-seq) was performed to identify the binding profile of native MeCP2 using micro-dissected dentate tissues. In both dentate tissues and HT22 cell lines, we demonstrated that MeCP2 could act as a transcription repressor on miR-682 with the involvement of the DNA methylation mechanism. Further, we found that miR-682 could bind to mRNA of phosphatase and tensin homolog (PTEN) in a sequence specific manner, thus leading to the suppression of PTEN and excessive activation of mTOR. This study therefore presents a novel epigenetic mechanism by identifying MeCP2/miR-682/PTEN/mTOR as an essential signal pathway in regulating the formation of MFS in the temporal lobe epileptic (TLE) mice. SIGNIFICANCE STATEMENT: Understanding the mechanisms that regulate axon guidance is important for a better comprehension of neural disorders. Sprouting of mossy fibers, one of the most consistent findings in patients with mesial temporal lobe epilepsy, has been considered a paradigmatic example of circuit plasticity in the adult brain. Although abnormal regulation of DNA methylation has been observed in both experimental rodents and humans with epilepsy, the potential role of DNA methylation in this well-documented example of sprouting of dentate axon remains elusive. This study demonstrates an essential role of methyl-CpG-binding protein 2 in the formation of mossy fiber sprouting. The underlying signal pathway has been also identified. The data hence provide new insight into epileptogenesis as well as axon misguidance in the central nervous system.

The effects of Helicobacter pylori eradication therapy on salivary pepsin concentration in patients with laryngopharyngeal reflux.

OBJECTIVE: To investigate the effect of Helicobacter pylori (HP) eradication therapy on salivary pepsin concentration in laryngopharyngeal reflux (LPR) patients with HP infection. MATERIALS AND METHODS: A total of 477 patients with suspected LPR were enrolled from June 2020 to September 2021. Reflux symptom index, reflux finding score, the positive rates and disintegrations per minute values of HP infection detected by 14C urea breath test and salivary pepsin concentrations analyzed using enzyme-linked immunosorbent assay were compared in LPR patients and non-LPR patients with or without HP infection. HP-positive patients were treated with HP eradication therapy while HP-negative patients with PPI therapy. RESULTS: The scores of nagging cough (0.88 vs. 0.50, P = 0.035), erythema or hyperemia (1.93 vs. 1.78, P = 0.035) and vocal fold edema (1.04 vs. 0.85, P = 0.025) were higher in the LPR (+) Hp (+) subgroup than in LPR (+) Hp (-) subgroup. The concentrations of salivary pepsin in the Hp (+) subgroup were higher than in the Hp (-) subgroup either in LPR patients (75.24 ng/ml vs. 61.39 ng/ml, P = 0.005) or the non-LPR patients (78.42 ng/ml vs. 48.96 ng/ml, P = 0.024). Compared to baseline (before treatment), scores of nagging cough (0.35 vs. 0.84, P = 0.019) and erythema or hyperemia (1.50 vs. 1.83, P = 0.039) and the concentrations of salivary pepsin (44.35 ng/ml vs. 74.15 ng/ml, P = 0.017) in LPR patients with HP infection decreased after HP treatment; yet, this was not observed for the LPR patients without HP infection treated with PPI only (P > 0.05). CONCLUSION: HP infection may aggravate the symptoms and signs of LPR patients, partly by increasing their salivary pepsin concentration.

A comparative analysis of laryngeal nerve damage in patients with idiopathic vocal cord paralysis exhibiting different paralytic sides.

Objective: To assess the extent of recurrent laryngeal nerve (RLN) and superior laryngeal nerve (SLN) damage in patients with idiopathic vocal cord paralysis (IVCP) exhibiting different paralytic sides. Methods: A total of 84 IVCP cases were evaluated using stroboscopic laryngoscopy, voice analysis, and laryngeal electromyography (LEMG). The results were compared between patients with left-sided paralysis and right-sided paralysis based on different disease courses (less than or more than 3 months). Results: Initially, the average age and disease progression of IVCP patients were found to be similar regardless of the side of paralysis (p > .05). Additionally, there were no significant variations in voice indicators, such as MPT, DSI, and VHI, between IVCP patients with left and right vocal cord paralysis (p > .05). Furthermore, no disparities were detected in the latencies and amplitudes of the paralyzed RLN and SLN, as well as the durations and amplitudes of the action potentials in the paralyzed TM and PCM, among IVCP patients with left and right vocal cord paralysis (p > .05). Notably, the amplitudes of the left paralytic CM were significantly lower than those of the right paralytic CM (0.45 vs. 0.53, Z = -2.013, p = .044). In addition, no disparities were observed in APDs and amplitudes between the ipsilateral PCM and TM, either for patients with left or right vocal fold paralysis (p > .05). Finally, all the IVCP patients were subdivided into two subgroups according to different disease course (less than or more than 3 months), and in each subgroup, the comparison of voice indicators and LEMG results in IVCP patients with left or right vocal fold paralysis were similar with the above findings (p > .05). Conclusion: Overall, the degree of RLN and SLN damage appeared to be similar in IVCP patients with left and right vocal cord paralysis, provided that the disease course was comparable. Level of Evidence: 4.

Submucosal Clustered Brownish Microvessels Based on NBI Endoscopy: A Characteristic of LPR.

OBJECTIVE: This study aimed to analyze the characteristics of laryngopharyngeal reflux (LPR) by using narrow band imaging (NBI) endoscopy. STUDY DESIGN: A prospective study. SETTING: A large-volume practice with tertiary care providers. METHODS: A total of 67 patients with suspected LPR who underwent 24-hour multichannel intraluminal impedance-pH monitoring were included from June 2020 to March 2022. Manifestations of NBI endoscopy included submucosal clustered brownish microvessels (CBMs), spotted brownish microvessels, and no special microvessels; the latter 2 formed the non-CBM group. The manifestations of all patients and their changes were observed after 8 weeks of proton pump inhibitor and symptomatic treatment for patients with LPR, and symptomatic treatment for patients without LPR. RESULTS: According to the results of 24-hour multichannel intraluminal impedance-pH monitoring, the incidence of submucosal CBMs was significantly higher in patients with LPR (30 cases) than in those without LPR (37 cases, P < .001), particularly in the posterior cricoid area (P < .001). Besides Reflux Finding Score, the incidence of signs such as subglottic edema and vocal fold edema was significantly higher in the CBM group than the non-CBM group (P < .05). Finally, 22 patients with LPR (91.7%) and only 2 patients without LPR (28.6%) underwent a transformation from CBMs to spotted brownish microvessels after continuous medication for 8 weeks in the CBM group (χ2 = 15.916, P < .001), while no significant change was observed in patients with or without LPR in the non-CBM group (P > .05). CONCLUSION: Submucosal CBMs in the posterior cricoid area under NBI endoscopy may be a characteristic of LPR.

RP11­156L14.1 regulates SSR1 expression by competitively binding to miR­548ao­3p in hypopharyngeal squamous cell carcinoma.

Emerging studies have demonstrated that long non­coding RNAs (lncRNAs) play essential roles in tumorigenesis. However, the role and function of lncRNAs in hypopharyngeal squamous cell carcinoma (HSCC) have not been completely elucidated. The present study explored the function of a novel lncRNA, RP11­156L14.1, in HSCC. RP11­156L14.1 was revealed to be highly expressed in HSCC tissues and cell lines. Knockdown of RP11­156L14.1 inhibited proliferation, migration, and invasion in HSCC cells. Furthermore, RP11­156L14.1 regulated epithelial­mesenchymal transition (EMT) by controlling EMT­related protein expression. Mechanistically, RP11­156L14.1 exerted its function as a competing endogenous RNA (ceRNA) and directly interacted with miR­548ao­3p. The present study also demonstrated that miR­548ao­3p regulated signal sequence receptor subunit 1 (SSR1) expression by targeting SSR1 3'­UTR. Moreover, the xenograft HSCC tumor model revealed that knockdown of RP11­156L14.1 markedly suppressed HSCC tumor growth in vivo. In summary, these findings indicated that the lncRNA RP11­156L14.1 functions as an oncogene in HSCC by competing with miR­548ao­3p in regulating SSR1 expression. The RP11­156L14.1/miR­548ao­3p/SSR1 axis could be utilized as a potential novel biomarker and therapeutic target for HSCC.

Self-protection of type III fibrocytes against severe 3-nitropropionic-acid-induced cochlear damage in mice.

After intense sound exposure, the lack of obvious degeneration in type III fibrocytes suggests that they might protect themselves against acoustic trauma. However, it is unknown whether and how type III fibrocytes play this role in other cochlear damage models. In this study, we investigated the self-protection of type III fibrocytes against severe cochlear energy failure induced by local administration of 3-nitropropionic acid to the inner ear. We detected that the type III fibrocytes did not degenerate significantly after 500 mM 3-nitropropionic acid application, and showed increased expression of proliferation marker Ki67. Moreover, low immunoreactivity for inducible nitric oxide synthase and cleaved caspase-3 was observed in type III fibrocytes 2 days after damage. These results indicate that after severe cochlear energy failure type III fibrocytes possess obvious proliferation activity, as well as strong antioxidant and antiapoptotic capacity, which can protect them from degeneration.

Effects of different bronchoalveolar lavage methods on tracheobronchial foreign body patients.

OBJECTIVES/HYPOTHESIS: To compare the different effects of bronchoalveolar lavage (BAL) with diverse combinations of lidocaine, epinephrine, and dexamethasone on pediatric patients with an inhaled tracheobronchial foreign body (TFB). STUDY DESIGN: Randomized controlled study. METHODS: Two hundred forty cases of pediatric patients with inhaled TFB were included in this study, and were randomly divided into four groups using three kinds of drugs for BAL, namely 0.9% saline (S) group, 2% lidocaine with diluted epinephrine (LE) group, 2% lidocaine with diluted epinephrine and 0.5% dexamethasone (LED), control group (C) without BAL. The incidences of intraoperative or postoperative complications and recovery periods were compared. Meanwhile, the concentrations of interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α in BAL fluids and plasma were evaluated by enzyme-linked immunosorbent assay. RESULTS: The incidences of bronchospasm, hypoxemia, and postoperative fever were significantly lower in the LED group than other groups (P < .001). Fever after the TFB removal procedure appeared later in the LED group than the other groups. The improvement and healing periods in the LE and LED groups were significantly shorter than those in the C and S groups (P < .001). The concentrations of IL-1ß, IL-6, and TNF-α in BAL fluids were significantly higher in the LE and LED groups than those in the S group (P < .001), but those in the plasma of the C and S groups were lower compared with the LE and LED groups (P < .001). CONCLUSIONS: BAL with lidocaine, epinephrine, and dexamethasone could promote recovery for TFB patients and reduce incidences of complications, possibly by regulating release of proinflammatory cytokines. LEVEL OF EVIDENCE: 1b.

Pepsin deteriorates prognosis of children with otitis media with effusion who undergo myringotomy or tympanostomy tube insertion.

OBJECTIVE: To investigate the concentrations of pepsin and pepsinogen within the middle ear cavity and determine whether pepsin and pepsinogen affect the prognosis of children with otitis media with effusion (OME). METHODS: All middle-ear lavage fluid from patients with OME undergoing myringotomy (M subgroup) or tympanostomy tube insertion (T subgroup) was collected and pepsin and pepsinogen were detected using enzyme-linked immunosorbent assay. After close follow-up over 2 years, the effects of pepsin and pepsinogen on the prognosis of the patients with OME in the M and T subgroups were analyzed. RESULTS: The average pepsin and pepsinogen concentrations were significantly lower in the M subgroup (n=54; 24.38±16.10mg/mL and 286.49±91.95mg/mL, respectively) than in the T subgroup (n=55; 45.56±16.60mg/mL and 664.92±107.06mg/mL; t=2.484, P=0.018 and t=2.670, P=0.011, respectively). In the M subgroup, the average time to tympanic membrane healing and tympanic pressure restoration to normal was much longer in pepsin(+) patients (17.0±2.0 days and 26.0±2.5 days, respectively) than in pepsin(-) patients (14.0±1.1 days and 22.0±1.0 days; t=3.871, P=0.001 and t=5.734, P=0.000, respectively), and the hearing level of pepsin(+) patients with OME ascended to 13.08±1.19dB, which was much lower than that of pepsin(-) patients (18.29±1.27dB; t=11.001, P=0.000). In the T subgroup, the complication rate including otorrhea and myringosclerosis was much higher in patients with high pepsin concentrations than in those with low pepsin concentrations (P<0.05). Finally, in both subgroups, the recurrence rates of OME in pepsin(+) or patients with high pepsin concentrations (34.6% [9/26] and 28.6% [10/35]) were significantly higher than those in pepsin(-) or low pepsin concentrations (10.7% [3/28] and 5.0% [1/20]; χ(2)=4.456, P=0.035 and χ(2)=4.420, P=0.036). However, pepsinogen had no significant effect on OME prognosis or recurrence. CONCLUSION: Pepsin but not pepsinogen could postpone tympanic membrane healing and pressure restoration in children with OME undergoing myringotomy and increase the incidence of recurrence and complications including otorrhea and myringosclerosis for those undergoing tympanostomy tube insertion. Therefore, pepsin could be considered a poor prognostic factor for OME, further emphasizing the important role of pepsin in OME pathogenesis.

MiR-139 targets CXCR4 and inhibits the proliferation and metastasis of laryngeal squamous carcinoma cells.

Our previous studies have showed that chemokine receptor 4 (CXCR4) was over-expressed in laryngeal squamous cell carcinoma (LSCC). However, the mechanism underlying aberrant CXCR4 expression remains unclear. To investigate the roles played by miRNAs in CXCR4 over-expression in LSCC, putative miR-139 was predicted through computational algorithms, including TargetScan, PicTar and miRBase, and luciferase reporter assay was explored to confirm that whether CXCR4 was directly regulated by miR-139. Then, quantitative real-time PCR, immunohistochemistry and in situ hybridization methods were employed to detect the expression of miR-139 and CXCR4 in primary LSCC tissues, normal adjacent mucosal tissues and metastatic lesions derived from 40 LSCC patients in the Second Hospital, Xi'An JiaoTong University. Finally, gain- and loss-of-function assays were adopted to explore the effects of miR-139 and CXCR4 on proliferation, invasion and metastasis of the human LSCC cell line Hep-2 in vitro and in vivo. Our results showed that miR-139 dampened CXCR4 expression, and CXCR4 was directly targeted by miR-139. Additionally, the expression of miR-139 was reduced in alignment with the progression of primary to metastatic LSCC. Moreover, an inverse correlation was observed between miR-139 and CXCR4 protein levels in LSCC specimens. Functional analyses demonstrated that ectopic expression of miR-139 inhibited cell proliferation, migration and metastasis of Hep-2 cells in vitro and in vivo. Similar to the observations seen in restoring miR-139 expression, dampening of CXCR4 expression inhibited cell growth, migration and invasion, whereas miR-139 over-expression reversed the pro-metastatic effect of CXCR4. Taken together, we conclude that miR-139 targets CXCR4 and inhibits proliferation and metastasis of LSCC.

Role of pepsin and pepsinogen: linking laryngopharyngeal reflux with otitis media with effusion in children.

OBJECTIVES/HYPOTHESIS: To analyze the relationship between laryngopharyngeal reflux (LPR) represented by pepsin and pepsinogen, and pathogenesis of otitis media with effusion (OME). STUDY DESIGN: Prospective case-control study. METHODS: Children with OME who required adenoidectomy and tympanostomy/tympanostomy tubes placement were enrolled in OME group, whereas children with adenoid hypertrophy (AH) who required adenoidectomy and individuals who required cochlear implantation (CI) were enrolled in AH and CI groups, respectively. Pepsinogen mRNA and protein levels were assessed by real-time fluorescence-based quantitative polymerase chain reaction and immunohistochemistry in adenoid specimens from the OME and AH groups. Pepsin and pepsinogen concentrations were evaluated by enzyme-linked immunosorbent assay in middle ear fluid and plasma from the OME and CI groups. RESULTS: The levels of pepsinogen protein expressed in cytoplasm of epithelial cells and clearance under epithelial cells in adenoid specimens from the OME group were significantly higher than those in the AH group. Furthermore, the concentrations of pepsin and pepsinogen in the OME group were 51.93±11.58 ng/mL and 728±342.6 ng/mL, respectively, which were significantly higher than those in the CI group (P<.001). In addition, the concentrations of pepsin in dry ears were significantly lower than those in serous and mucus ears in the OME group (F=22.77, P<.001).Finally, the concentration of pepsinogen in middle ear effusion was positively correlated with the expression intensity of pepsinogen protein in cytoplasm of epithelial cells (r=0.73, P<.05) in the OME group. CONCLUSIONS: Pepsin and pepsinogen in middle ear effusion are probably caused by LPR and may be involved in the pathogenesis of OME. LEVEL OF EVIDENCE: 3b.

[Expression of human beta-defensin-2 in laryngeal squamous cell carcinoma and its correlation with CD1a(+) dendritic cells infiltrating in tumor.].

OBJECTIVE: To investigate the expression of human beta-defensin-2 (HBD-2) in laryngeal squamous cell carcinoma (LSCC), and reveal the immune significance of HBD-2 in LSCC. METHODS: The expression of HBD-2 and the infiltration of CD1a(+) dendritic cells were verified by immunohistochemistry strept actividin-biotin complex (SABC) method in thirty-four cases of LSCC's paraffin sections, and the expression of HBD-2 mRNA was detected in eighteen cases of postoperative tumor specimens and five cases of normal paracarcinoma tissue using cDNA probe by in situ hybridization. Based on the type and characteristic of datas, SPSS13.0 program package was used for statistical calculation. RESULTS: (1) The difference of expression of HBD-2 was significant among the grades of malignant cell differentiation (F = 6.809, P < 0.05), the expression of HBD-2 in well differentiated group was significantly increased compared with moderately and poorly differentiated group (P < 0.05), there wasn't significant difference with the clinical type, T status and lymphatic metastasis in the expression of HBD-2. (2) The difference of expression of HBD-2 mRNA among different grades of malignant cell differentiation was significant (F = 16.391, P < 0.05), less or no positive signal was observed in poorly differentiated group. (3) The infiltration of CD1a(+) dendritic cells had positive correlation with the expression of HBD-2 (r = 0.343, P < 0.05). CONCLUSIONS: The expression of HBD-2 was related to the grade of malignant cell differentiation in LSCC, which was regulated at the level of transcription and translation. HBD-2 might play a role in anti-tumor immunity by chemo-attractiving and activating of immature dendritic cells.