Motor endplate‐expressing cartilage‐muscle implants for reconstruction of a denervated hemilarynx

Objective

Tissue engineering of the larynx requires a complex, multiple tissue layer design. Additionally, spontaneous reinnervation of the larynx after recurrent laryngeal nerve (RLN) injury is often disorganized, resulting in subpar function. This study investigates use of tissue‐engineered cartilage and motor endplate‐expressing (MEE) tissue‐engineered skeletal muscle implants for laryngeal reconstruction and the promotion of organized reinnervation after RLN injury.

Methods

F344 rat primary muscle progenitor cells (MPCs) were isolated. Three‐dimensional muscle constructs were created by encapsulating MPCs in type I oligomeric collagen under passive tension. Constructs were then cultured in differentiation medium (MPC control constructs) or induced to form motor endplates (MEE constructs) with neurotrophic agents. Three‐dimensional cartilage constructs were created with adipose stem cells differentiated in chondrocyte medium. The muscle and cartilage constructs were implanted into surgically created myochondral defects in the F344 rat larynx with injured or intact (control) RLN. At 1‐, 3‐, and 6‐month timepoints, videolaryngoscopy, electromyography (EMG), histology, and immunohistochemistry were used to assess outcomes.

Results

At all timepoints, cartilage‐muscle implants were well integrated into host tissue. Functionally, there was increased vocal fold adduction and EMG activity in nerve‐injured rats treated with the MEE constructs when compared to those treated with the MPC control constructs. Motor endplate‐expressing constructs had increased myofiber cross‐sectional area compared to MPC control constructs.

Conclusion

Although our laboratory previously demonstrated that muscle and cartilage constructs could be used separately for hemilaryngeal reconstruction, this study suggests combining them with the modification of MEEs rather than MPCs, resulting in improved muscle recovery after recurrent laryngeal nerve injury.

Level of Evidence

NA. Laryngoscope, 2018

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Regarding Primary Surgery Versus Primary Radiation‐Based Treatment for Locally Advanced Oropharyngeal Cancer

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Real‐Time Light‐Guided Vocal Fold Injection: Ex Vivo Feasibility Study in a Canine Model

Objective

The transcricothyroid (CT) membrane approach is a good option for office‐based vocal fold injection (VFI). However, because the needle tip is invisible during injection using the CT approach, precise localization requires a high level of experience, and mastering this approach involves a steep learning curve. To overcome current limitations, we conceptualized a novel technique: real‐time light‐guided VFI (RL‐VFI), which enables simultaneous VFI under direct visualization of the lighted needle tip. Herein, we aimed to verify the feasibility of RL‐VFI in cadaveric canine model, simulating the setting of office‐based VFI, as well as to explore its clinical usefulness.

Study Design

Animal study.

Methods

A customized prototype device was developed. It consisted of three parts: light source, controller, and injector. Light source comprised laser diodes of two wavelengths (635 nanometers [nm], red; 532 nm, green). Four types of injector were developed using 40‐mm needles of 23‐ and 25‐gauge and optic fibers of 50 and 100 μm. ex vivo canine larynx was prepared for the experiment. Flexible laryngoscopy system was used to examine canine vocal folds.

Results

Various routes from three insertion points (3 mm, 10 mm, and 17 mm from the midline) were validated using the device. Regardless of the injection routes, the location of the needle tip was accurately indicated by light. RL‐VFI was feasible under light guidance without difficulties. Moreover, precise and simultaneous re‐injection could be performed at the intended point using the device.

Conclusion

We introduced RL‐VFI using our customized prototype device in an ex vivo canine larynx, simulating the setting of office‐based VFI. Clinical application of RL‐VFI will improve safety and precision of CT approach, as well as expand its applications in laryngology.

Level of Evidence

NA. Laryngoscope, 2018

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The history of chicken and other bird exploitation in Thailand: Preliminary analysis of bird remains from four archaeological sites

Abstract

The origin of the domestication of chicken Gallus gallus domesticus is still a subject of debate. It principally originates from the red junglefowl G. gallus, which is distributed throughout Southeast Asia and South China. However, the prehistoric exploitation of chicken and red junglefowl in Southeast Asia has remained unclear due to a small number of samples for bone analysis. In this study, we analyzed bird remains from four archaeological sites in Thailand: Ban Hua Ud (4000–3000 BP), Long Long Rak (1900–1600 BP), Ban Non Wat (3750–1500 BP), and Keed Kin (5^th–11^th Century AD). Bones from pheasants/fowls (Phasianidae), storks (Ciconiidae), cormorants (Phalacrocoracidae), crows (Corvidae), and passerines (Passeriformes) were found. A small number of stork bones were found at three lowland sites, with none found at Long Long Rak. Cormorant bones were dominant at Ban Hua Ud but were only found at that site. Pheasant/fowl bones were found at all sites except Ban Hua Ud, and dominated the number of identified specimens, suggesting that they were the most commonly used bird, at least after the Iron Age. Morphological bone identification criteria for the 24 Phasianidae species in Thailand have not been established; ancient DNA analysis is required to differentiate chicken/red junglefowl bone from other indigenous pheasants/fowls. It is obvious that mitochondrial DNA remained unchanged in the early stages of the domestication process. Further studies combining ancient DNA and morphological analyses are required to judge whether all of the candidate chicken bones are indeed from red junglefowl or chicken. While these results are regionally and temporally restricted, they give important quantitative information on bird exploitation during the Neolithic and early historic Dvaravadi periods in Thailand.

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Novel GLI2 mutations identified in patients with Combined Pituitary Hormone Deficiency (CPHD): evidence for a pathogenic effect by functional characterization

Abstract

Context

The Gli‐family of zinc‐finger transcription factors regulates the Sonic Hedgehog (Shh) signalling pathway that plays a key role in early pituitary and ventral forebrain development. Heterozygous GLI2 loss of function mutations in humans have been reported in holoprosencephaly (HPE), HPE‐like phenotypes associated with pituitary anomalies and combined pituitary hormone deficiency with or without other extra pituitary findings.

Objective

The aim of this study was the search for GLI2 mutations in a cohort of Italian CPHD patients and the assessment of a pathogenic role for the identified variants through in vitro studies.

Patients

One hundred‐forty‐five unrelated CPHD patients diagnosed with or without extra pituitary manifestations were recruited from different Italian centres.

Methods

The GLI2 mutation screening was carried out through direct sequencing of all the 13 exons and intron‐exon boundaries. Luciferase reporter assays were performed to evaluate the role of the detected missense variants.

Results

Five different novel heterozygous non‐synonymous GLI2 variants were identified in 5 patients. The mutations were three missense (p.Pro386Leu, p.Tyr575His, p.Ala593Val), one frameshift (p.Val1111Glyfs*19) and one nonsense (p.Arg1226X). The latter two mutants are likely pathogenic since they lead to a truncated protein. The in‐vitro functional study of the plasmids bearing two of the three missense variants (namely p.Tyr575His and p.Ala593Val) revealed a significant reduction in transcriptional activity.

Conclusion

In conclusion, the analysis of GLI2 in individuals with CPHD led to the identification of five variations with a likely negative impact on the GLI2 protein, confirming that GLI2 is an important causative gene in CPHD. The functional in vitro study analysis performed on the missense variations were useful to strengthen the hypothesis of pathogenicity.

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Quality of life is significantly impaired in both secretory and non‐functioning pituitary adenomas

Summary

Objective

To evaluate the quality of life (QoL) in patients with pituitary adenomas in comparison with healthy Mexican population QoL scores.

Design & Measurements

Cross‐sectional study using the short form 36 questionnaire (SF‐36) in 175 patients with pituitary adenomas grouped by adenoma subtype and disease activity, and compared them with the healthy Mexican population normative QoL scores.

Patients

44 patients with non‐functioning pituitary adenomas (NFPA), 48 with acromegaly, 53 with prolactinomas and 30 with Cushing disease (CD) were enrolled in this study.

Results

Mental and physical components scores (MCS & PCS) of SF‐36 questionnaire were lower in patients with active disease in all adenoma subtypes (p<0.03). A significant negative relationship between prolactin levels and MCS (r= ‐0.30, p<0.01) and PCS (r=‐0.41, p<0.01) were found in prolactinomas. Patients with CD showed 24h urine free cortisol levels negatively correlated with MCS (r=‐0.43, p <0.01) but not with PCS. No significant correlation was found between IGF‐1ULN and QoL scores in acromegaly. NFPA patients had lower QoL scores than patients with controlled CD, acromegaly or prolactinoma (p <0.02). Active CD and prolactinoma have lower QoL scores in comparison of NFPA (p <0.05). Having an adenoma, secretory or non‐functional, decrease QoL scores in comparison of results in the healthy Mexican population register. Using an adjusted‐multivariate model, we confirmed that disease activity in all secretory adenomas is an independent risk factor, reducing SF‐36 scores significantly.

Conclusion

Activity in all secretory pituitary adenomas' patients decrease mental and physical QoL. However, independently of disease activity, secretory and NFPA significantly decrease QoL in comparison with healthy Mexican population QoL register.

This article is protected by copyright. All rights reserved.

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Effect of cosmetic chemical preservatives on resident flora isolated from healthy facial skin

Abstract

Background/Aims

Healthy skin harbors numerous microbes known to maintain its health and prevent attacks from external pathogens. The influence of chemical preservatives commonly used in cosmetic products on facial resident flora remains poorly characterized. In this study, we aimed to investigate the antibacterial activity of five such preservatives on in vitro cultivated skin‐resident bacteria.

Methods

Both Gram‐positive and Gram‐negative bacteria were isolated on blood agar, tryptic soy agar, and nutrient agar; Gram‐negative bacteria were then selected on Hank's balanced salt solution containing antibiotics and Reasoner's 2A. The minimum inhibitory concentrations (MICs) of methylisothiazolinone (MTI), iodopropynyl butylcarbamate (IPBC), ethylhexylglycerin (EHG), methylparaben (MP), and phenoxyethanol (PE) were estimated for nine facial resident bacteria, Escherichia coli, and Staphylococcus aureus using serial broth dilution in vitro.

Results

The maximum test concentrations coincided with the upper limits set by the "Cosmetic Safety and Technical Specification" (2015 edition, China). Nine facial resident bacteria were isolated from 14 healthy adults: Staphylococcus epidermidis, Staphylococcus capitis, Kocuria, Micrococcus luteus, Bacillus, Acinetobacter, Pseudomonas parafulva, Pseudomonas oleovorans, and Roseomonas cervicalis. MTI and IPBC displayed the strongest effect on all tested strains (MICs ≤0.01%), followed by EHG and MP (MICs ≤0.3%), and finally PE with the weakest effect (MIC ≤1%).

Conclusion

The five chemical preservatives assayed inhibited survival of the nine facial resident bacteria isolates, when tested at the maximum allowed limit. The corresponding MICs will provide a reference for the effective utilization of these compounds in product formulations.

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