Identification of positionally distinct astrocyte subtypes whose identities are specified by a homeodomain code.

Astrocytes constitute the most abundant cell type in the central nervous system (CNS) and play diverse functional roles, but the ontogenetic origins of this phenotypic diversity are poorly understood. We have investigated whether positional identity, a fundamental organizing principle governing the generation of neuronal subtype diversity, is also relevant to astrocyte diversification. We identified three positionally distinct subtypes of white-matter astrocytes (WMA) in the spinal cord, which can be distinguished by the combinatorial expression of Reelin and Slit1. These astrocyte subtypes derive from progenitor domains expressing the homeodomain transcription factors Pax6 and Nkx6.1, respectively. Loss- and gain-of-function experiments indicate that the positional identity of these astrocyte subtypes is controlled by Pax6 and Nkx6.1 in a combinatorial manner. Thus, positional identity is an organizing principle underlying astrocyte, as well as neuronal, subtype diversification and is controlled by a homeodomain transcriptional code whose elements are reutilized following the specification of neuronal identity earlier in development.

Postoperative respiratory adverse events in children after endoscopic laryngeal cleft repair.

PURPOSE: Due to the serious nature of respiratory adverse events, understanding their incidence can help in decisions regarding safe postoperative disposition. There have been no studies, however, evaluating the risk of postoperative respiratory adverse events (PRAEs) in children undergoing endoscopic laryngeal cleft (LC) repair, which is the primary objective of this study. METHODS: We conducted a retrospective chart review of all patients who underwent LC repair at a large tertiary children's hospital from 2015 to 2020. PRAEs were defined as having at least one of the following: remained intubated, required reintubation, required positive pressure ventilation, required high flow O2 nasal cannula, or required more than one dose of racemic epinephrine. Univariate analyses compared demographic, preoperative characteristics, and intraoperative characteristics between those with and without a PRAE. RESULTS: Overall, 8/26 (31%) patients had a PRAE and there were no differences between patients who did and did not have a PRAE and most comorbidities. Younger age (p = 0.03), being male (p = 0.07), and being admitted preoperatively (p = 0.07) were potentially associated with PRAEs. Need for intraoperative intubation for any reason or duration was associated with increased incidence of PRAEs (p = 0.02). CONCLUSION: The overall 31% incidence of postoperative respiratory adverse events reaffirms the appropriateness of PICU disposition for a large proportion of children undergoing endoscopic LC repair. Further studies with increased sample sizes are needed to tease apart patient or procedure-specific factors that significantly increase the risk of respiratory adverse events to have more definitive evidence regarding safe postoperative disposition.

Perioperative respiratory adverse events in children undergoing triple endoscopy.

OBJECTIVES: Children with aerodigestive disorders often have many of the reported risk factors for development of perioperative respiratory adverse events. This study sought to evaluate the incidence of such events in this group of patients undergoing general anesthesia for "triple endoscopy" (flexible bronchoscopy with bronchoalveolar lavage, rigid laryngoscopy and bronchoscopy, and esophagogastroduodenoscopy) and to identify any patient-specific or procedure-specific risk factors associated with higher incidence of perioperative respiratory adverse events. METHODS: We performed a retrospective chart review of children 18 years or younger who underwent triple endoscopy as part of an aerodigestive evaluation. Data collected from medical records included: preoperative polysomnography, symptoms of acute respiratory illness, medical comorbidities, demographics, postoperative hospital or intensive care unit admission, and all respiratory events and interventions in the perioperative period. Patient-specific and procedure-specific factors were investigated via univariate analysis for any correlations with perioperative respiratory adverse events. RESULTS: Of the 122 patients undergoing triple endoscopy, 69 (57%) experienced a perioperative respiratory adverse event. We found no difference in the incidence of perioperative respiratory adverse events among children with documented lung disease compared with those with no lung disease (OR: 0.89, p = .8 95% CI: 0.43, 1.8), and no significant difference between those children who had a respiratory illness at the time of surgery, 1-2 weeks prior, 3-4 weeks prior, and those with no preceding respiratory illness. A higher percentage of males had a perioperative respiratory adverse event, compared with females (OR: 2.7, p = .01 95% CI: 1.3, 5.09). CONCLUSION: Patients undergoing triple endoscopy for evaluation of aerodigestive disorders at our institution experienced perioperative respiratory adverse events at a rate of 57%.

Differential epithelial growth in tissue-engineered larynx and trachea generated from postnatal and fetal progenitor cells.

Postnatal organ-specific stem and progenitor cells are an attractive potential donor cell for tissue-engineering because they can be harvested autologous from the recipient and have sufficient potential to regenerate the tissue of interest with less risk for ectopic growth or tumor formation compared to donor cells from embryonic or fetal sources. We describe the generation of tissue-engineered larynx and trachea (TELT) from human and mouse postnatal organoid units (OU) as well as from human fetal OU. Mouse TELT contained differentiated respiratory epithelium lining large lumens, cartilage and smooth muscle. In contrast, human postnatal TE trachea, formed small epithelial lumens with rare differentiation, in addition to smooth muscle and cartilage. Human fetal TELT contained the largest epithelial lumens with all differentiated cell types as well as smooth muscle and cartilage. Increased epithelial cytokeratin 14 was identified in both human fetal and postnatal TELT compared to native trachea, consistent with regenerative basal cells. Cilia in TELT epithelium also demonstrated function with beating movements. While both human postnatal and fetal progenitors have the potential to generate TELT, there is more epithelial growth and differentiation from fetal progenitors, highlighting fundamental differences in these cell populations.

Identifying Posterior Fossa Abnormality in Pediatric Aerodigestive Patients, a Case Series.

OBJECTIVES: Multidisciplinary clinics like Aerodigestive programs focus on issues associated with airway, pulmonary, and gastrointestinal issues. Rarely, significant neurological issues like posterior fossa abnormality are identified as the primary etiology. We describe 3 such patients and compare their clinical presentation to the other patients seen in Aerodigestive clinic. METHODS: A retrospective chart review was conducted to review the 3 posterior fossa patients and the remainder of children that were referred to the Aerodigestive Clinic at Children's Hospital Los Angeles from June 2016 to August 2018. Clinical characteristics including triple endoscopies and sleep studies were recorded. RESULTS: Of the 110 patients included for review, 3 patients (3%) had an underlying posterior fossa abnormality; all of whom had symptoms of sleep disordered breathing along with dysphagia compared with 30% incidence of this symptom profile in the remaining Aerodigestive population. CONCLUSION: Presence of sleep disordered breathing and dysphagia, with underlying vomiting history, warrants considering evaluation for posterior fossa abnormalities in addition to traditional workup for aerodigestive disorders. Due to the rarity of this presentation and small sample size, future studies with multicenter collaboration may help better describe identifiers to delineate this population with similar aerodigestive symptoms and clarify diagnostic algorithms.

Pediatric Invasive Fungal Rhinosinusitis: A Comprehensive Analysis of Prognostic Factors for Survival.

OBJECTIVE: Pediatric invasive fungal rhinosinusitis (IFS) is a devastating infection that manifests almost exclusively in immunocompromised children. The goal of this work was to determine which clinical features carry prognostic value for survival. METHODS: A retrospective review of children with a histopathological diagnosis of IFS was performed at an academic tertiary care institution from 1990 to 2021. Clinical variables were collected to generate survival and life-table estimators at 6-months and 1-year. RESULTS: Eighteen patients were included in this analysis, with a mean age of 9.8 years (range, 1-17 years). Most children were neutropenic (n = 15, 83.3%), with acute lymphoblastic leukemia (n = 10, 55.6%) representing the most common primary diagnosis. A mean of 3.2 operations (range 1-7 operations) was performed per patient for either mucormycosis (n = 10, 55.6%) or aspergillosis (n = 8, 44.4%). The mean time to absolute neutrophil count recovery was 65.8 days (range 20-137 days), with a 6-month and 1-year survival rate of 47.6% and 41.7%, respectively. Gross total resection (p = 0.006, p < 0.001), number of antifungals (p = 0.0004, p = 0.0003), and total operation number (p = 0.0032, p = 0.0035), served as positive prognostic factors for 6-month and 1-year survival. Conversely, altered mental status (p = 0.0026), cerebral involvement (p = 0.0010), cranial neuropathies (p < 0.0001), hyperglycemia (p = 0.0445, p = 0.0208), and intensive care unit status (p = 0.0013) served as negative prognostic factors for 6-month and 1-year survival. CONCLUSION: Several key elements were identified and found to play a vital role in influencing survival for pediatric IFS. Early diagnosis, prompt medical therapy, and aggressive surgical intervention remain at the forefront in the treatment of this complex opportunistic infection. LEVEL OF EVIDENCE: 4 Laryngoscope, 133:1239-1250, 2023.

Disparities in Access to Pediatric Otolaryngology Care During the COVID-19 Pandemic.

OBJECTIVE: Racial/ethnic minority pediatric otolaryngology patients experience health disparities, including barriers to accessing health care. Our hypothesis for this study is that Hispanic or economically disadvantaged patients would represent a larger percentage of missed appointments and report more barriers to receiving care during the COVID-19 pandemic. METHODS: A cross-sectional survey utilizing a modified version of the Barriers to Care Questionnaire was administered via telephone to no-show patients, and median income by zip code was collected. Chi-squared, logistic regression, and Student's t-tests were used to investigate any differences in those who did and did not keep their appointments as well as any differences in mean questionnaire scores. RESULTS: No-show patients were more likely to be Hispanic than not (OR 2.3, 95% CI: 1.3, 3.9, P = .002) and to live in a zip code that had a median income less than 200% of the federal poverty level (OR 1.7, 95% CI: 1.2, 2.4, P = .004). Respondents with a high school degree tended to report more barriers to care compared to those with less education. CONCLUSION: In our study, we identified ethnic, financial, and logistic concerns that may contribute to patients failing to keep their appointments with the otolaryngology clinic. Future studies are needed to assess the efficacy of measures aimed to reduce these barriers to care such as preventive plans to assist new patients and expanding telehealth services.

The effect of anesthesia without opioid on perioperative opioid demand in children with severe obstructive sleep apnea (OSA) for adenotonsillectomies - single-center retrospective observational study.

BACKGROUND: Children with severe obstructive sleep apnea (OSA) carry a higher risk of respiratory complications after adenotonsillectomy. Their altered sensitivity to opioids may be a significant contributor to respiratory morbidity. The purpose of this study was to identify how anesthesia without opioids affects perioperative opioid demand and postoperative course. METHODS: A chart review of children with severe OSA (apnea hypoxia index; AHI ≥ 10) undergoing adenotonsillectomies was performed. Comorbidities and perioperative medications were documented. Perioperative opioid doses within 48 h of procedure were calculated as morphine equivalents (mcg/kg). Pain scores, rescue medications, and postoperative complications in PICU and non-PICU settings were also documented. Anesthesia without opioid and with opioid groups were compared. RESULTS: The analysis included 225 children. A significantly higher percentage of children received no postoperative opioids in the anesthesia without opioid group compared to those with opioid (46 of 88 children vs. 43 of 137; P < 0.05). The incidence of severe postoperative pain between the two groups was not different in PICU (P = 0.88) or non-PICU setting (P = 0.84). Perioperative opioid administration was significantly lower in anesthesia without opioid (median, Q1, Q3: 0.0, 0.0, 83.0) compared to with opioid (144.4, 72.5, 222.2; P < 0.01). Anesthesia without opioid was one of the independent factors to achieve perioperative opioid avoidance (<50mcg/kg). CONCLUSIONS: Anesthesia without opioid for children with severe OSA for tonsillectomy significantly reduced perioperative demand for opioid and did not affect the occurrence of severe pain. Anesthesia without opioid is an effective strategy to minimalize opioid demand perioperatively for children with severe OSA for tonsillectomy.

Preoperative SARS-CoV-2 Screening Fails to Detect Viral Particles Prior to Airway Surgery.

OBJECTIVES/HYPOTHESIS: Children have higher rates of asymptomatic SARS-CoV-2 infections or milder courses of infection, and their carrier status may potentially impact viral transmission to those providing them care. The aim of this study is to compare the existing COVID-19 preoperative screening protocols to the detection of SARS-CoV-2 viral particles in surgical samples. STUDY DESIGN: Cross-sectional study. METHODS: We conducted a prospective study with consecutive convenience sampling of children undergoing adenoidectomy between January and April 2021. Total nucleic acid was extracted from adenoid tissue and real-time reverse transcription-polymerase chain reaction was conducted to test for the presence of SARS-CoV-2 viral particles. Univariate logistic regression was used to summarize the effect size of variables of interest on the odds of having SARS-CoV-2 positive adenoid tissue. RESULTS: Forty adenoid samples were collected and 11 (27.5%) had a positive SARS-CoV-2 reverse transcriptase-polymerase chain reaction. Patients with positive adenoids were older (11.8 vs. 7.9 years, odds ratio: 1.3, P = .01) and more likely to have had a positive nasopharyngeal swab in the previous 90 days (4/11 or 36% vs. 0). CONCLUSION: These data are the first report on the presence of SARS-CoV-2 particles in pediatric adenoidectomy specimens, with a high percentage of patients showing evidence of viral particles within the adenoid. This finding calls in to question the utility of preoperative COVID screening protocols which have yet to be rigorously validated in asymptomatic patients and have the potential to delay patients' surgical care. LEVEL OF EVIDENCE: 3 Laryngoscope, 132:1665-1667, 2022.

Risk factors for peri-operative respiratory adverse events after supraglottoplasty.

OBJECTIVE: To determine demographic and peri-operative characteristics that predict peri-operative respiratory adverse events (PRAE) requiring intensive care unit (ICU) level intervention after supraglottoplasty. STUDY DESIGN: Case series with chart review. SETTING: Tertiary care children's hospital. METHODS: Retrospective chart review was conducted of children with laryngomalacia who underwent supraglottoplasty between October 2014 and November 2019. PRAE were defined as any of the following events or requirements within 24 h of surgery: 1) failure to extubate, 2) reintubation, 3) positive pressure ventilation, 4) more than one dose of racemic epinephrine, or 5) greater than 4 L of oxygen via nasal cannula. RESULTS: Fifty-one subjects were enrolled, with a median age of 4.8 months and majority (62 %) Hispanic. Twenty-one (41 %) subjects experienced PRAE. After adjusting for age and gender on multivariate analysis, children admitted preoperatively to the ICU were more likely to have PRAE than those not admitted or admitted to a floor unit (OR 40.1, 95%CI: 4.1-388.6, p = 0.001). Additionally, children with intraoperative oxygen desaturations below 90 % for greater than 1 min were more likely to have PRAE than those who did not (OR 21.3, 95%CI: 2.4-189.9, p = 0.006). Other factors significantly associated with PRAE on univariate analysis included chronic lung disease, congenital cardiac abnormality, history of intubation, supplemental oxygen requirement, gastrostomy tube dependence, intraoperative intubation and longer surgery length. CONCLUSION: Preoperative ICU admission and intraoperative oxygen desaturations are independent risk factors for PRAE after supraglottoplasty. Results from this study can help inform decisions regarding the appropriate level of postoperative care required after supraglottoplasty. LEVEL OF EVIDENCE: IV.

Tracheal squamous metaplasia in children with endotracheal intubation or tracheostomy.

OBJECTIVES: In the present study, tracheal epithelial biopsy samples between intubated children, children with tracheostomy and a control group of non-intubated children are compared with respect to their degree of normal differentiation versus the presence of squamous metaplasia. METHODS: Tracheal epithelial biopsies were obtained from intubated neonates undergoing tracheostomy, children with tracheostomy undergoing suprastomal granuloma excision and non-intubated control children undergoing laryngoscopy and bronchoscopy. Paraffin tissue blocks were sectioned at 5 µm thickness and subjected to both routine Hematoxylin and Eosin (H&E) staining and immunostained with the relevant antibodies for markers of epithelial differentiation including B-tubulin, CC10, Muc5ac, P63, keratin5 and keratin14. RESULTS: Squamous metaplasia was seen in 3/3 infants, all intubated and in 3/3 children with tracheostomy tubes in place undergoing excision of suprastomal granuloma. No metaplasia was observed in control tracheal epithelial biopsies in 7/7 non-intubated children. CONCLUSION: Our results demonstrate a clear association between intubation or tracheostomy and the presence of squamous metaplasia which is not otherwise encountered in control pediatric tracheal biopsies.

The transcription factor NFIA controls the onset of gliogenesis in the developing spinal cord.

The mechanisms controlling the transition from neurogenesis to gliogenesis in the vertebrate CNS are incompletely understood. We identified a family of transcription factors, called NFI genes, which are induced throughout the spinal cord ventricular zone (VZ) concomitantly with the induction of GLAST, an early marker of gliogenesis. NFIA is both necessary and sufficient for GLAST induction in the VZ. Unexpectedly, NFIA is also essential for the continued inhibition of neurogenesis in VZ progenitors. This function is mediated by the requirement of NFIA for the expression of HES5, a Notch effector. However, Notch effectors are unable to promote glial-fate specification in the absence of NFIA. Thus, NFIA links the abrogation of neurogenesis to a generic program of gliogenesis, in both astrocyte and oligodendrocyte VZ progenitors. At later stages, NFIA promotes migration and differentiation of astrocyte precursors, a function that is antagonized in oligodendrocyte precursors by Olig2.

Factors impacting participation in flexible endoscopic evaluation of swallowing in children.

OBJECTIVES: Flexible endoscopic evaluation of swallowing (FEES) is a common and useful tool for assessment of dysphagia and swallowing safety in children, however an important problem is limited participation in some children. We examine the factors associated with adequate participation in children undergoing FEES, including the completion rate, whether a clear diagnosis or feeding plan was made, and the incidence of adverse events (AEs). METHODS: We conducted a retrospective review of children undergoing FEES at an urban children's hospital. Data collected included age, gender, race, comorbidities, AEs and outcomes of FEES. RESULTS: Of the 130 patients, 46 (35%) were female with a median age of 2.6 years. 113 (87%) patients cooperated and obtained a result, while 15 (12%) patients did not cooperate and no result was obtained; there was no significant difference in gender, comorbidities, or race between those groups. The age distribution of FEES subjects in this study was skewed with nearly 75% below age 5. Patients who cooperated had a higher median age (3 years) than those who did not (1.2 years) and 47% of non-cooperators were less than 1 year of age. There were no significant AEs, the most common mild AE was excessive crying (34 subjects, 26%). In a multivariate model controlling for age and gender, excessive crying was associated with a decreased odds of cooperation (OR: 0.16, p = 0.004, 95% CI: 0.04, 0.54). CONCLUSION: Children who undergo FEES have an overall favorable completion rate and no serious adverse events, however its utility is limited in cases where children refuse to participate. Understanding the factors associated with failure to cooperate with FEES is important in developing strategies to improve participation. Excessive crying is identified as such a factor in this study.

Safety and efficacy of Early injection laryngoplasty in pediatric patients.

OBJECTIVE: The major morbidity of unilateral vocal fold immobility (UVFI) in children is due to aspiration. Early injection laryngoplasty (IL) can decrease aspiration risk; however, this has not been well studied in pediatric otolaryngology. This study examines safety and efficacy of early IL in children. METHODS: Retrospective review of pediatric patients undergoing IL with any injectate between 2006 and 2017 within 6 months of onset of UVFI. Outcomes included diet pre- and postprocedure, incidence of aspiration-related sequelae, and adverse events. RESULTS: Seventeen patients met eligibility criteria. Ten (58.8%) were males. Median age was 8 months (interquartile range, 2 months-11.5 years). All patients had prior surgeries; the largest subgroup (11 patients, 64.7%) had UVFI after repair of a congenital cardiac defect. Other causes included thyroidectomy, high vagal injury, and prolonged intubation. Sixteen patients underwent swallowing evaluation prior to IL and 14 patients required dietary modifications due to aspiration risk. Consistency and/or volume of oral intake improved after IL in 10 (71.4%) of them. Five patients underwent gastrostomy tube placement for significant oromotor incoordination. Children with congenital cardiac defects had more previous surgeries (3.0 ± 0.4 vs. 1.2 ± 0.2, P = .006) and were more likely to require G-tube placement due to poor feeding despite IL (45% vs. 0%, P = .05). No patients experienced adverse events due to IL; in particular, none experienced airway symptoms requiring intubation. CONCLUSION: Early IL in pediatric patients with UVFI is safe and can reduce aspiration and improve oral intake. Future studies should elucidate patient subgroups most likely to benefit from this intervention. LEVEL OF EVIDENCE: 4 Laryngoscope, 129:1699-1705, 2019.

Geographic health disparities in the Los Angeles pediatric neck abscess population.

OBJECTIVE: To assess geographical sociodemographic differences in neck abscesses that require surgical drainage in Los Angeles. STUDY DESIGN: retrospective review. METHODS: We reviewed the medical records of 119 consecutive pediatric patients at Children's Hospital Los Angeles (CHLA) from 2014 to 2017 with a diagnosis of a neck abscess requiring incision and drainage. Sociodemographic information including zip code of residence was extracted and analyzed with Chi-square, Fisher's exact test, and multivariate logistic regression. RESULTS: The average age of patients with a neck abscess in this study was 3.4 years old, 53.8% were female, 54.6% were Hispanic, and 82.5% had public health insurance. 79% of patients had an abscess located in the superficial neck, and 10.1% had an abscess located in the retropharyngeal space. There were no significant differences in gender, race, type of health insurance, or income between patients that lived within 10 miles of CHLA versus those that lived farther than 10 miles. On multivariate analysis, zip codes with a high volume of neck abscesses were more likely to be lower income neighborhoods. Gender, race, type of health insurance, and distance from CHLA were not associated with zip codes with a high volume of neck abscesses. CONCLUSION: Geographic areas in the greater Los Angeles community with a high volume of neck abscesses requiring incision and drainage at our institution were associated with lower income neighborhoods. LEVEL OF EVIDENCE: IV.

Geographic health disparities in the Los Angeles pediatric esophageal foreign body population.

OBJECTIVE: To assess geographical sociodemographic differences in the pediatric esophageal foreign body population of Los Angeles. METHODS: We retrospectively reviewed the medical records of 128 consecutive pediatric patients at Children's Hospital Los Angeles (CHLA) from 2014 to 2017 with a diagnosis of a retained foreign body in the esophagus removed by rigid or flexible esophagoscopy. Sociodemographic information including zip code of residence was extracted and analyzed with Chi-square, Fisher's exact test, and multivariable logistic regression. RESULTS: The average age of patients with a retained esophageal foreign body in this study was 2.5 years old, 52.3% were male, 91.4% had no past medical history, 53.1% were Hispanic, 82.0% had public health insurance, and 63.3% were transfers from an outside hospital. The most common foreign body removed was a coin. There were no significant differences in gender, race, type of health insurance, or income between patients that lived within 10 miles of CHLA versus farther than 10 miles. On multivariable analysis, zip codes with a high volume of esophageal foreign bodies were more likely to be lower income neighborhoods. Gender, race, type of health insurance, and distance from CHLA were not risk factors for zip codes with a high volume of esophageal foreign bodies. CONCLUSION: Geographic areas in the greater Los Angeles community with a high volume of retained esophageal foreign bodies requiring endoscopic removal at our institution are associated with lower income neighborhoods. Further studies should be performed to better understand health disparities within the U.S. pediatric esophageal foreign body population.

Establishing Proximal and Distal Regional Identities in Murine and Human Tissue-Engineered Lung and Trachea.

The cellular and molecular mechanisms that underpin regeneration of the human lung are unknown, and the study of lung repair has been impeded by the necessity for reductionist models that may exclude key components. We hypothesized that multicellular epithelial and mesenchymal cell clusters or lung organoid units (LuOU) could be transplanted to recapitulate proximal and distal cellular structures of the native lung and airways. Transplantation of LuOU resulted in the growth of tissue-engineered lung (TELu) that contained the necessary cell types consistent with native adult lung tissue and demonstrated proliferative cells at 2 and 4 weeks. This technique recapitulated important elements of both mouse and human lungs featuring key components of both the proximal and distal lung regions. When LuOU were generated from whole lung, TELu contained key epithelial and mesenchymal cell types, and the origin of the cells was traced from both ActinGFP and SPCGFP donors to indicate that the cells in TELu were derived from the transplanted LuOU. Alveolar epithelial type 2 cells (AEC2s), club cells, ciliated cells marked by beta-tubulin IV, alveolar epithelial type I cells, Sox-2-positive proximal airway progenitors, p63-positive basal cells, and CGRP-positive pulmonary neuroendocrine cells were identified in the TELu. The mesenchymal components of peribronchial smooth muscle and nerve were identified with a CD31-positive donor endothelial cell contribution to TELu vasculature. TELu successfully grew from postnatal tissues from whole murine and human lung, distal murine lung, as well as murine and human trachea. These data support a model of postnatal lung regeneration containing the diverse cell types present in the entirety of the respiratory tract.

Bacterial biofilms and increased bacterial counts are associated with airway stenosis.

OBJECTIVES: Most airway stenoses are acquired secondary to the use of prolonged endotracheal intubation. Antibiotics have been shown to decrease local inflammation and granulation tissue formation in the trachea. However, antibiotic therapy is not 100% effective in preventing or treating granulation tissue formation. Development of bacterial biofilms may explain this finding. This study evaluates the difference between tracheal stenotic segments and normal trachea in terms of (1) presence of bacterial biofilms, (2) quantitative bacterial counts, and (3) inflammatory markers. STUDY DESIGN: Cross-sectional study. SETTING: Tertiary care academic medical center. SUBJECTS: A total of 12 patients were included in the study. Tissue from stenotic segments from 6 patients with airway stenosis undergoing open airway procedures were compared with tracheal tissue from 6 patients without airway stenosis undergoing tracheostomy. METHODS: Scanning electron microscopy for biofilm detection, quantitative polymerase chain reaction for quantitative analysis of bacterial count, and immunohistochemistry were performed for inflammatory markers transforming growth factor ß1 (TGF-ß1) and SMAD3. RESULTS: Compared with the patients without airway stenosis, patients in the airway stenosis group showed presence of bacterial biofilms, a significantly higher expression of 16S rRNA gene copies per microgram of tissue (187.5 vs 7.33, P = .01), and higher expression of TGF-ß1 (91% vs 8%, P < .001) and SMAD3 (83.5% vs 17.8%, P < .001). CONCLUSION: Bacterial biofilms, increased bacterial counts, and higher expression of TGF-ß1 and SMAD3 are associated with airway stenosis.

Relationship of eosinophils and plasma cells to biofilm in chronic rhinosinusitis.

BACKGROUND: This study investigates the relationship of eosinophils and plasma cells to biofilm in chronic rhinosinusitis (CRS). A prospective observational study was performed at the Keck Hospital, University of Southern California, Department of Otolaryngology, Los Angeles, CA. METHODS: A total of 29 patients, 20 undergoing endoscopic sinus surgery for CRS and 9 control patients undergoing septoplasty for nasal obstruction without history or evidence of CRS, were included in this study. Contiguous sinonasal mucosa sample sections were examined by hematoxylin and eosin (H&E), fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC) for biofilm, microbes, eosinophil major basic protein (EMBP), and cluster designation 27 (CD27). EMBP and CD27 were used as eosinophil and plasma cell markers, respectively. RESULTS: Biofilm was visualized in 15 of 20 patients with CRS on H&E sections, confirmed by microbial presence using FISH. Biofilm was not identified in tissue samples of the nine control patients. On IHC analysis, CD27 and EMBP expression were significantly higher in patients with CRS compared with control (p < 0.05) and had greater expression in biofilm-positive patients compared with biofilm-negative patients. Nasal polyps correlated with higher expression of CD27 and EMBP, but in CRS patients without polyps CD27 and EMBP was also significantly greater in biofilm-positive specimens compared with biofilm-negative specimens. CONCLUSION: Biofilm presence in CRS appears to correlate to host inflammatory response involving plasma cell and eosinophil recruitment.

Gold nanorod-sphingosine kinase siRNA nanocomplexes: a novel therapeutic tool for potent radiosensitization of head and neck cancer.

Radiation therapy (RT) is an important treatment modality used against a number of human cancers, including head and neck squamous cell carcinoma (HNSCC). However, most of these cancers have an inherent anti-apoptotic mechanism that makes them resistant to radiation therapy. This radioresistance of cancer cells necessitates the irradiation of tumor areas with extremely high doses of radiation to achieve effective therapy, resulting in damage to normal tissues and leading to several treatment related side effects. These side effects significantly impair the quality of life of treated patients, and preclude the possibility of repeat radiation treatment in patients with tumor recurrence. Our previous research has correlated the upregulation of the anti-apoptotic sphingosine kinase (SphK1) gene in HNSCC cells with their radioresistance properties. In the current study, we hypothesized that by downregulating the SphK1 gene using nanotechnology mediated gene silencing, we can render these cells more vulnerable to radiation therapy by enabling apoptosis at lower radiation doses. We have employed biocompatible gold nanorods (GNRs) as carriers of short interfering RNA (siRNA) targeting the SphK1 gene. GNRs play a critical role in protecting the siRNA molecules against physiological degradation, as well as delivering them inside target cells. Following their synthesis and characterization, these nanoplexes were applied to HNSCC cells in culture, resulting in the radiosensitization of the treated cells. Furthermore, the GNR-siRNA nanoplexes were injected intratumorally into subcutaneous HNSCC tumors grown in mice, prior to the initiation of radiation therapy in vivo. Subsequent exposure of GNR-SphK1siRNA nanoplex-treated tumors to radiation (GNR-SphK1siRNA + IRRA) resulted in over 50% tumor regression compared to control GNR-GFPsiRNA nanoplex and radiation treated tumors (GNR-GFPsiRNA + IRRA). In addition, we were able to induce this tumor regression in nanoplex treated tumors with radiation doses much lower than those commonly required in clinical RT. These experiments lay the foundation for the development of a nanotechnology-mediated gene silencing tool for more potent radiation therapy of a number of human cancers, with minimal, if any, toxic side effects.