Math1 gene therapy for kanamycin and furosemide-induced deaf guinea pigs / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To observe the morphology and function changes of cochlear hair cells before and after math1 gene injection into the cochlea of deaf guinea pigs which were induced by kanamycin and furosemide. To explore the feasibility of Math1 gene for medicine-induced deafness therapy.</p><p><b>METHODS</b>Kanamycin (500 mg/kg) and furosemide (50 mg/kg) were given to the healthy adult guinea pigs intramuscularly and intravenously to establish the deafness model. The guinea pigs whose auditory brainstem response (ABR) threshold > 95 dB SPL were randomly divided into five groups. Blank control group (without any treatment, n = 3), operation control group (right ear scala tympani operation, n = 3), artificial perilymph group (right ear scala tympani injection artificial perilymph, n = 3), virus vector group [right ear scala tympani injection adenovirus which carrying enhanced green fluorescent protein (EGFP) gene (Ad. EGFP) , n = 4], Math1 gene therapy group [right ear scala tympani injection adenovirus which carrying Math1 and EGFP gene (Ad. Math1-EGFP), n = 6]. Each animal received ABR test before and after injection. The cochlear tissue was observed by scanning electronic microscopy.</p><p><b>RESULTS</b>The ABR thresholds of tone burst( 4, 8, 16, 20 kHz ) were not statistically significant in different groups (P > 0.05). The number of hair cells increased in some of severe deaf guinea pigs after the injection of Ad. Math1-EGFP gene. However, there was no obvious difference with morphology and numbers of cochlea hair cells in other groups.</p><p><b>CONCLUSIONS</b>The injection of Math1 gene to cochlea can regenerate or repair the hair cells of medicine-induced deaf guinea pigs, but there was no improvement on the hearing loss.</p>

Histological changes of peripheral vestibular organs in the inner ears of Smad4 conditional knockout mice / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the histological changes in the vestibular endorgans of Smad4 gene conditional knockout mice and to explore the influence of the Smad4 gene on vestibular development.</p><p><b>METHODS</b>Histological changes of periphery vestibular organs in inner ear of Smad4 conditional knockout mice were investigated by frozen sections, immunofluorescence, confocal microscopy, scanning electron microscopy and transmission electron microscopy.</p><p><b>RESULTS</b>There was no Smad4 expression in the inner ear cartilage capsule of Smad4-/- mice. In Smad4+/- mice, Smad4 expression in the same cartilage capsule was positive, and it was strong positive in Smad4+/+ mice. Smad4 expression in vestibular sense epithelium, crista ampullaris and macula, was positive. And no difference was found among these three genotypes. Studying at scanning electron microscopy and transmission electron microscopy levels and anti-filament immunofluorescence showed that no pathological changes were observed in all the three genotype mice.</p><p><b>CONCLUSION</b>Although the Smad4 gene was knockout effectively in the auricular cartilage capsule of Smad4 conditional knockout mice,the histological changes of Smad4 conditional knockout mice in vestibulum auris internal were slightly.</p>

Death mode-dependent reduction in succinate dehydrogenase activity in hair cells of aging rat cochleae / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Our previous studies have shown that both apoptosis and necrosis are involved in hair cell (HC) pathogenesis in aging cochleae. To better understand the biological mechanisms responsible for the regulation of HC death, we examined the activity of succinate dehydrogenase (SDH), a mitochondrial bioenergetic enzyme, in the HCs of aging cochleae.</p><p><b>METHODS</b>The auditory brainstem response thresholds elicited by tone bursts at 4, 10 and 20 kHz were measured in both young (2-3 months) and aging (22-23 months) Wistar rats. SDH activity was evaluated with a colorimetric assay using nitroblue tetrazolium monosodium salt. The SDH-labeled organs of Corti were double stained with propidium iodide, a DNA intercalating fluorescent probe for illustration of HC nuclei. All the specimens were examined with fluorescence microscopy and confocal microscopy.</p><p><b>RESULTS</b>Aging rats exhibited a significant elevation of ABR thresholds with threshold shifts being 34 dB at 20 kHz, 28 dB at 10 kHz, and 25 dB at 4 kHz. Consistent with the reduction in the cochlear function, aging cochleae exhibited the reduction of SDH staining intensity in the apical and the basal ends of the cochleae, where a large number of apoptotic, necrotic, and missing HCs were evident. The reduction in SDH staining appeared in a cell-death-mode dependent fashion. Specifically, SDH labeling remained in apoptotic HCs. In contrast, SDH staining was markedly reduced or absent in necrotic HCs.</p><p><b>CONCLUSIONS</b>In the aging cochlea, SDH activity is preserved in HCs undergoing apoptosis, but is substantially reduced in necrosis. These results suggest that mitochondrial energetic function is involved in the regulation of cell death pathways in the pathogenesis of aging cochleae.</p>

Bacteriophage Lysins:A Novel Effective Antibacterial Agents / 微生物学通报

Lysins are efficient bacteria cell wall digesting enzymes encoded by DNA bacteriophage. Gram-positive bacteriophage lysins feature similar domain structure, high lytic efficiency, synergic antibacterial effect with antibiotics, rare neutralization by antibodies, less chance of developing drug-resistant strains, et al. The past decade has seen a considerable amount of research worldwidely focused on lysin, and lysins have been used successfully in a variety of animal models to control pathogenic antibiotic resistant bacteria found on mucosal surfaces and infected tissues. The great potential of lysins as an anti-infective agent prompted this review.

Relevance between clinical behavior and bionomical findings of acoustic neuromas / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To evaluate the relationship between labeling index (LI) Ki-67, proliferating cell nuclear antigen (PCNA) and transforming growth factor-beta1 (TGF-beta1) with the clinical behavior of acoustic neuroma.</p><p><b>METHODS</b>Expression of Ki-67, PCNA and TGF-beta1 was detected by immunohistochemistry in 53 specimens of acoustic neuromas. The relationship among tumor proliferation, histological representation, size of tumor, clinical proliferation index of tumor and tumor proliferation activity were analyzed.</p><p><b>RESULTS</b>In all 53 cases, the positive rate of Ki-67 was 77.4% (41/53) but the positive rate of PCNA was 84.9% (45/53). There was significant difference between the proliferate index, clinic growth rate and course of disease (t = 2.14, t = 2.70; P < 0.05). The positive rate of TGF-beta1 was 83.0% (44/53). The correlation of TGF-beta1 with LI (Ki-67) was significant difference (r = 0.36, P < 0.05). Cystic degeneration often occurred in large-size tumor (Z = 4.44, P < 0.05). There was no significant relationship between the expression of LI (Ki-67), LI (PCNA) and TGF-beta1 and the course of disease as well as between the cystic degeneration and the non-cystic degeneration. Although clinic growth rate of cystic degeneration was bigger than that of non-cystic degeneration, there was not statistically significant.</p><p><b>CONCLUSIONS</b>Ki-67 and PCNA are reflected proliferation activities of tumor cells in acoustic neuromas. Cell proliferation-labeling index LI (PCNA) was related with clinical growth rates. TGF-beta1 might participate in the biological behavior of acoustic neuroma. Cystic degeneration was one of special pattern of acoustic neuroma, however, tumor enlargement might due to the volume of the cystic but unrelated to fast proliferation of parenchyma cell.</p>

Electron microscopic observation of vocal fold polyps / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>The purpose of this study was to observe the ultrastructure of the fibroblasts, collagen and elastic fibers in vocal fold polyps.</p><p><b>METHODS</b>Ten vocal fold polyps and 3 normal vocal fold specimens obtained from total laryngectomy were studied by means of transmission electron microscope and scanning electron microscope.</p><p><b>RESULTS</b>The result showed that in vocal fold polyps, the quantity of fibroblasts increased and there were abundant organelles, suggesting that the fibroblast were in the status of activation. As the main cell to produce lamina propria extracellular matrix, the representation suggested that the extracellular matrix metabolism was active. Leucocytes soakage was observed, suggesting that the inflammation may play a role in the lesion. It was found by scanning electron microscopy that in case of lesions, collagen fibers and elastic fibers arrayed irregularly.</p><p><b>CONCLUSIONS</b>Under pathologic circumstance, fibroblasts, collagen and elastic fibers altered in morphology, which possibly induced the functional alteration.</p>

Ultrastructural localization of aquaporin 1 in endolymphatic sac of the mouse / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To study the ultrastructural localization of aquaporin 1 (AQP1) in the endolymphatic sac (ES) of the mouse inner ear and explore the function of the AQP1 in ES.</p><p><b>METHODS</b>The cellular localization of AQP1 in ES of the mouse inner ear was investigated by immunocytochemistry. The ultrastructural localization of AQP1 in the mouse inner ear was performed by immunogold electron microscopy which is characterized as cryoprotection and high sensitivity.</p><p><b>RESULTS</b>In the ES, strong AQP1 labeling was observed in the sub-epithelial connective tissue. Fibroblasts of sub-epithelial connective tissue of the ES present densely labeling of gold particles. But the epithelial cells of the ES were devoid of labeling. AQP1 was localized on the cell processes of the fibrocytes.</p><p><b>CONCLUSIONS</b>AQP1 in the ES may play an important role in absorbing water and regulate the balance of fluid and ion in the inner ear.</p>

In vitro culture of greater epithelial ridge cells from rat cochleae / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To establish in vitro culture systems of greater epithelial ridge (GER) cells from rat cochlear and to investigate the characterization, growth pattern and ultrastructure of GER cells.</p><p><b>METHODS</b>Using a combinatorial approach of enzymatic digestion and mechanical separation to allow isolation and culture of GER cells from P1 rat cochleae. The dissociated GER cells were cultured in serum-free or 10% fetal bovine serum DMEM respectively. BrdU, phalloidin, ZO1, calretinin and myosin VIIa immunostaining and scanning electron microscope observation were performed in GER cell cultures.</p><p><b>RESULTS</b>The dissociated GER cell cultures showed positive to ZO1, phalloidin and BrdU staining, but negative to myosin VIIa and calretinin. They assumed a polygonal morphology which was similar to epithelial cells and grew in islands-like patches in medium containing 10% fetal bovine serum while forming spheres in serum-free medium. The GER cells presented significant ability to proliferate in both conditions. Scanning electron microscope showed that there was microvillus and centre bodies but not hair cell specific stereociliary bundles on the surface of GER cultures.</p><p><b>CONCLUSIONS</b>The GER cell cultures showed significant ability to proliferate and grew in islands-like patches in medium containing 10% fetal bovine serum while forming spheres in serum-free medium. The dissociated GER cells expressed epithelial cell specific marker but not marker of hair cells.</p>

Effect of glutamate on distortion product otoacoustic emission and auditory brainstem response in guinea pigs / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the changes of the potentials and structure of the guinea pig cochlear during whole cochlear perfusion with glutamate.</p><p><b>METHODS</b>Cochlear microphonics (CM), compound action potential (CAP), distortion product otoacoustic emission (DPOAE) and auditory brainstem response (ABR) were measured to indicate the cochlear functional properties during whole cochlear perfusion. The morphology of the cochlear was monitored by transmission electron microscopy.</p><p><b>RESULTS</b>There were no significant DPOAE changes before and after glutamate perfusion. CM I/O function maintained a nonlinear characteristic during infusion. After glutamate perfusion, ABR latencies were delayed. There was significant difference in CAP threshold before and after glutamate perfusion. The average CAP threshold was elevated 35 dB. The OHCs appeared normal, but IHCs and afferent dendrites showed cytoplasmic blebs after glutamate infusion.</p><p><b>CONCLUSIONS</b>Glutamate is thought to be a primary amino acid neurotransmitter at the synapses formed by cochlear hair cells and spiral ganglion neurons. However, the excessive glutamate is neurotoxic for cells, and it can destroy the IHCs and spiral ganglion neurons. The present method can also be built up as an animal model of auditory neuropathy.</p>

The potential of avian cytokines as immunotherapeutics and vaccine adjuvants / 生物工程学报

With the imminent and widespread ban of the use of antibiotic feed additives and chemical antimicrobials in food production animals, alternative measures need to be sought to ensure that the livestock industry will not be adversely affected. Cytokines are proteins that control the type and extent of an immune response following infection or vaccination. They therefore represent excellent naturally occurring therapeutics. The identification, cloning and characterisation of cytokine genes in chickens have lagged somewhat behind similar work in mammals. Progress in isolating chicken homologues of mammalian cytokines has also been slowed by the generally low level of sequence similarity. Chicken cytokine genes that have been cloned to date include ChIFN-gamma, ChIL-1beta, ChIFN-alpha, ChIL-15, ChIL-18, ChIL-8, ChIL-2, ChIL-6, ChIL-16, SCF, MGF, TGFbeta, Lymphotactin, MIP-1beta, CXC and CC chemokines, so the use of cytokines in poultry has become more feasible with the discovery of a number of avian cytokine genes. The delivery methods for chicken cytokine are of prime importance and are required to be safe, easy to administer and cost-effective. Live viral vectors such as fowl adenovirus (FAV) expressing cytokine genes can be delivered via drinking water or aerosol sprays, making it very easy to administer. Since the immune system of chickens is similar to that of mammals, they offer an attractive model system to study the effectiveness of cytokine therapy in the control of disease in livestock. This review focus on the recent advances made in avian cytokines, with a particular focus on their assessment as therapeutic agents and vaccine adjuvants.