Establishment of a Large Animal Model for Eustachian Tube Functional Study in Miniature Pigs.

This study was performed to investigate whether miniature pigs are a suitable animal model for studies of the Eustachian tube (ET). Sixteen Chinese experimental miniature pigs were used in this investigation. Ten animals were used for anatomical and morphometric analyses to obtain qualitative and quantitative information regarding the ET. Three animals were used for histological analysis to determine the fine structure of ET cross-sections. Three animals were used to investigate the feasibility of balloon dilation of the Eustachian tube (BDET). The anatomical study indicated that the pharyngeal orifice and tympanic orifice of the miniature pig ET are located at the posterior end of the nasal lateral wall and anterior wall of the middle ear cavity, respectively. The cartilaginous tube was seen to pass through the whole length of the ET, the length of the cartilaginous part of the ET and the diameter of the isthmus were similar between humans and miniature pigs. The inclination of the ET in miniature pigs was larger than that in humans. The gross histology seemed to be slightly different between miniature pig and human, but the fine structures were essentially the same in both species. BDET experiments verified that the miniature pig model is suitable as a model for clinical operations. The miniature pig ET corresponds very well to that of humans. In addition, the miniature pig ET is suitable as a model for clinical operations. Therefore, the miniature pig is a valid animal model for ET study. Anat Rec, 302:1024-1038, 2019. © 2019 Wiley Periodicals, Inc.

Crocetin downregulates the proinflammatory cytokines in methylcholanthrene-induced rodent tumor model and inhibits COX-2 expression in cervical cancer cells.

The effect of crocetin (C20H24O4) on methylcholanthrene- (MCA-) induced uterine cervical cancer in mice was studied in this paper. After the mice were treated orally with crocetin, maleic dialdehyde (MDA), polymorphonuclear cells (PMN), interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) were examined by ELISA or immunohistochemistry. The inducible nitric oxide synthase (iNOS) activation in HeLa cells was analyzed using fluorescence microscopy for light microscopic examination. The MCA mice showed a significant increase in plasma MDA, PMN, IL-1ß, TNF-α, and nitrates levels. At the same time, the mRNA level of COX-2 in HeLa cells was also significantly increased. These changes were attenuated by crocetin supplementation in the MCA mice. Crocetin supplementation in the MCA mice also showed protection against cervical cancer. These results suggest that crocetin may act as a chemopreventive and an anti-inflammatory agent.

[Endolymphatic sac tumor with von Hippel-Lindau disease: report of two cases with testing of von Hippel-Lindau gene].

OBJECTIVE: Endolymphatic sac tumors (ELSTs) are rare in the general population with much higher prevalence in von Hippel-Lindau(VHL) disease. The purpose of this study is to present two cases of endolymphatic sac tumor with VHL disease with analysis of VHL gene and to explore their association with VHL disease using molecular analysis. METHODS: Clinical data of these two patients from different VHL families were studied. DNAs extracted from peripheral bloods were amplified by the polymerase chain reaction using oligonucleotide primers corresponding to the VHL gene, then compared the mutations with the Human Gene Mutation Database. RESULTS: In case 1, 6 family members were enrolled in the study. Among them, three had been identified to have a germline missense point mutation at codon 194 of the VHL gene exon 1 (p.S65W). The little sister of the patient (case 1) underwent vitrectomy for retinal hemangioblastoma 5 years ago in another hospital. The mother of the patient (case 1) was further diagnosed to have a cerebellar hemangioblastoma and renal carcinoma in the following physical examination. Case 2 with her parents were also tested. Codon 499 of the VHL gene exon 3 (p.R167W) were detected in case 2 and her mother, but the mother refused further examination. CONCLUSIONS: The genetic diagnosis plays an important role in early detection of symptomatic patients and suspected patients. Clinical screening for members of the VHL families, and close follow-up of carriers allow an early detection of tumors and the metastasis, which is the most common cause of death of these patients.

Type I hair cell regeneration induced by Math1 gene transfer following neomycin ototoxicity in rat vestibular sensory epithelium.

CONCLUSION: In the current study, hair cells of vestibular terminal organs in rats were completely eliminated with trans-scala vestibuli injection of neomycin, and then the Math1 gene was transferred. It was shown that type I vestibular hair cells were regenerated and synapses were formed. OBJECTIVES: The objective of this study was to identify the cell type of the regenerated vestibular hair cells and relative innervation and synaptic linkage after hair cells of vestibular terminal organs in rats were completely eliminated. METHODS: Neomycin injection was used to eliminate all the vestibular terminal organs, and then the animals were treated with an injection of Ad-Math1-EGFP in the scala vestibuli of the cochlea. RESULTS: Math1 gene transfer into the inner ear induced type I hair cell regeneration and synaptic formation. However, neither the number nor the appearance of the hair cells was normal.

Surgical treatment of endolymphatic sac tumor.

The objective of this study was to understand the clinical characteristics of endolymphatic sac tumor and to optimize its diagnosis and treatment. We carried out a retrospective review of 11 patients diagnosed as having endolymphatic sac tumor based on operative findings and pathological features, and their clinical manifestations, differential diagnosis, and surgical approaches are discussed in detail. The lesions of 10 cases were completely surgically resected, two cases via the mastoid approach, 8 cases via the oto-cervical or cranio-oto-cervical combined approach. In one case the tumor was partially removed and the patient received adjuvant radiotherapy. In operation, four cases had facial-hypoglossal neural anastomosis, two cases had great auricular nerve graft, and in four cases the facial nerve integrity remained. Survival follow-up data range from 14 months to 10 years. We conclude that endolymphatic sac tumor is very rare and easily misdiagnosed. Reasonable surgical treatment can provide a good prognosis.

The role of Smad4 in vestibular development in mice.

The regulation of the bone morphogenetic protein (BMP) signal transduction pathway is important in the development of the inner ear and vestibular system. We reported previously that small mothers against decapentaplegic homolog-4 (Smad4) is required for inner ear cochlear development and normal auditory function in mammals; however, the distribution and functional mechanisms of Smad4 at various stages of vestibular development remained unclear. To investigate the relationship between the Smad4 gene and vestibular organ development, we measured changes in the expression of BMP4 and Smad4 during vestibular development in C57BL/6 mice. In addition, vestibular structures, pathologic changes, and the vestibular function of chondrocyte-specific Smad4 knockout mice were compared to those of the control group. We found that the expression of Smad4 in the inner ear was delayed compared with that of BMP4. Moreover, chondrocyte-specific Smad4 knockout homozygous mice showed stunted growth and partial vestibular deformities, but it showed less histologic changes in the vestibular end-organs and saccule dysfunction. These results suggest that Smad4 participates in late-stage shaping of the configuration of the vestibule and development of vestibular functional, but a Smad4-independent pathway for the inner ear vestibular BMP4 signal transduction could not be rule out.

Combined application of oto-endoscopes and nasal endoscopes for resection of dermoid tumor in eustachian tube.

Dermoid tumor in the eustachian tube (DTIET) is a congenital disease. Our patient had the symptom of discharging pus in the left ear when he was about 1 year old. Due to lack of understanding of the disease at that time, he was misdiagnosed as having cholesteatoma, for which he underwent three surgical operations in the middle ear. Recently, with detailed preoperative imaging estimation and full consideration of surgical risks, under the guidance of TV, using oto-endoscopes and nasal endoscopes jointly, the DTIET was resected completely via the combined approach of pharyngeal and tympanic openings of the eustachian tube. This case indicated again that it was a space-occupying lesion in the eustachian tube. On the basis of the characteristic expressions of CT and MRI, the possibility of DTIET should be fully considered. Endoscopic technology has advantages for surgical operation in the eustachian tube, which might remove the mass and achieve the aim of minimal trauma.

Chondrocyte-specific Smad4 gene conditional knockout results in hearing loss and inner ear malformation in mice.

Smad4 is the central intracellular mediator of transforming growth factor-beta (TGF-beta) signaling, which plays crucial roles in tissue regeneration, cell differentiation, embryonic development, and regulation of the immune system. Conventional Smad4 gene knockout results in embryonic lethality, precluding its use in studies of the role of Smad4 in inner ear development. We used chondrocyte-specific Smad4 knockout mice (Smad4Co/Co) to investigate the function of Smad4 in inner ear development. Smad4Co/Co mice were characterized by a smaller cochlear volume, bone malformation, and abnormalities of the osseous spiral lamina and basilar membrane. The development of the hair cells was also abnormal, as evidenced by the disorganized stereocilia and reduced density of the neuronal processes beneath the hair cells. Auditory function tests revealed the homozygous Smad4Co/Co mice suffered from severe sensorineural hearing loss. Our results suggest that Smad4 is required for inner ear development and normal auditory function in mammals.

Smad5 haploinsufficiency leads to hair cell and hearing loss.

The Smads are a group of related intracellular proteins critical for transmitting the signals to the nucleus from the transforming growth factor-beta superfamily at the cell surface. Knockout of the Smad5 is embryonic lethal. However, the Smad5 knockout of single allele (+/-) could survive. We used Smad5 heterozygous knockout (+/-) to determine the role of Smad5 in the development of inner ear morphology and function. In situ hybridization showed that Smad5 was expressed predominantly in hair cells, spiral ganglion, and supporting cells. Measurements of hearing thresholds using auditory brainstem response showed that Smad5 defect resulted in progressive hearing loss between 4 and 24 weeks after birth. Morphological examination revealed apoptosis in the inner ear, with significant loss of outer hair cells in adult Smad5 mutant mice. Our results indicated that deficiency in the Smad5-mediated signaling resulted in apoptosis of hair cells, suggesting Smad5 is a gene that may be related with presbycusis.

(2Z)-Ethyl 5-(4-methoxy-phen-yl)-7-methyl-3-oxo-2-(3,4,5-trimethoxy-benzyl-idene)-3,5-dihydro-2H-thia-zolo[3,2-a]pyrimidine-6-carboxyl-ate.

In the title compound, C(27)H(28)N(2)O(7)S, the dihedral angles between the thia-zole ring and the mono- and tris-ubstituted benzene rings are 87.8 (2) and 17.9 (3)°, respectively. The dihydro-pyrimidine ring adopts a flattened boat conformation. In the crystal structure, π-π stacking occurs [centroid-centroid separation = 3.6611 (11) Å].

Ethyl 5-(4-chloro-phen-yl)-2-[(Z)-(methoxy-carbon-yl)methyl-ene]-7-methyl-3-oxo-3,5-dihydro-2H-thia-zolo[3,2-a]pyrimi-dine-6-carboxyl-ate.

The title compound, C(19)H(17)ClN(2)O(5)S, was synthesized by the reaction of ethyl 6-(4-chloro-phen-yl)-2-mercapto-4-methyl-1,6-dihydro-pyrimidine-5-carboxyl-ate and dimethyl acetyl-ene-dicarboxyl-ate in methanol. In the mol-ecule, the nearly planar thia-zole ring, with a mean deviation from the plane of 0.0108 (3) Å, is fused with a dihydro-pyrimidine ring in a flattened half-chair conformation.

rac-(4R,17S,18R,26R)-Ethyl 4'-methoxy-carbonyl-5''-(4-methoxy-phen-yl)-1'-methyl-2,3''-dioxo-2'',3''-dihydro-indoline-3-spiro-2'-pyrrolidine-3'-spiro-2''-thia-zolo[3,2-a]pyrimidine-6''-carboxyl-ate.

In the title compound, C(30)H(30)N(4)O(7)S, the two spiro junctions link a planar 2-oxindole ring [with a mean deviation from the plane of 0.0319 (3) Å, a pyrrolidine ring in an envelope conformation and a thia-zolo[3,2-a]pyrimidine system. Two mol-ecules are connected into a dimer by two N-H⋯O hydrogen bonds, forming an R(2) (2)(8) graph-set motif. The title compound has four stereogenic centers and appears as a racemic mixture of one single diastereoisomer (RSRR/SRSS).