Proteome of normal human perilymph and perilymph from people with disabling vertigo.

The vast majority of hearing loss, the most common sensory impairment, and vertigo, which commonly causes falls, both reflect underlying dysfunction of inner ear cells. Perilymph sampling can thus provide molecular cues to hearing and balance disorders. While such "liquid biopsy" of the inner ear is not yet in routine clinical practice, previous studies have uncovered alterations in perilymph in patients with certain types of hearing loss. However, the proteome of perilymph from patients with intact hearing has been unknown. Furthermore, no complete characterization of perilymph from patients with vestibular dysfunction has been reported. Here, using liquid-chromatography with tandem mass spectrometry, we analyzed samples of normal perilymph collected from three patients with skull base meningiomas and intact hearing. We identified 228 proteins that were common across the samples, establishing a greatly expanded proteome of the previously inferred normal human perilymph. Further comparison to perilymph obtained from three patients with vestibular dysfunction with drop attacks due to Meniere's disease showed 38 proteins with significantly differential abundance. The abundance of four protein candidates with previously unknown roles in inner ear biology was validated in murine cochleae by immunohistochemistry and in situ hybridization: AACT, HGFAC, EFEMP1, and TGFBI. Together, these results motivate future work in characterizing the normal human perilymph and identifying biomarkers of inner ear disease.

Immediate and delayed cochlear neuropathy after noise exposure in pubescent mice.

Moderate acoustic overexposure in adult rodents is known to cause acute loss of synapses on sensory inner hair cells (IHCs) and delayed degeneration of the auditory nerve, despite the completely reversible temporary threshold shift (TTS) and morphologically intact hair cells. Our objective was to determine whether a cochlear synaptopathy followed by neuropathy occurs after noise exposure in pubescence, and to define neuropathic versus non-neuropathic noise levels for pubescent mice. While exposing 6 week old CBA/CaJ mice to 8-16 kHz bandpass noise for 2 hrs, we defined 97 dB sound pressure level (SPL) as the threshold for this particular type of neuropathic exposure associated with TTS, and 94 dB SPL as the highest non-neuropathic noise level associated with TTS. Exposure to 100 dB SPL caused permanent threshold shift although exposure of 16 week old mice to the same noise is reported to cause only TTS. Amplitude of wave I of the auditory brainstem response, which reflects the summed activity of the cochlear nerve, was complemented by synaptic ribbon counts in IHCs using confocal microscopy, and by stereological counts of peripheral axons and cell bodies of the cochlear nerve from 24 hours to 16 months post exposure. Mice exposed to neuropathic noise demonstrated immediate cochlear synaptopathy by 24 hours post exposure, and delayed neurodegeneration characterized by axonal retraction at 8 months, and spiral ganglion cell loss at 8-16 months post exposure. Although the damage was initially limited to the cochlear base, it progressed to also involve the cochlear apex by 8 months post exposure. Our data demonstrate a fine line between neuropathic and non-neuropathic noise levels associated with TTS in the pubescent cochlea.

FGF23 deficiency leads to mixed hearing loss and middle ear malformation in mice.

Fibroblast growth factor 23 (FGF23) is a circulating hormone important in phosphate homeostasis. Abnormal serum levels of FGF23 result in systemic pathologies in humans and mice, including renal phosphate wasting diseases and hyperphosphatemia. We sought to uncover the role FGF23 plays in the auditory system due to shared molecular mechanisms and genetic pathways between ear and kidney development, the critical roles multiple FGFs play in auditory development and the known hearing phenotype in mice deficient in klotho (KL), a critical co-factor for FGF23 signaling. Using functional assessments of hearing, we demonstrate that Fgf[Formula: see text] mice are profoundly deaf. Fgf[Formula: see text] mice have moderate hearing loss above 20 kHz, consistent with mixed conductive and sensorineural pathology of both middle and inner ear origin. Histology and high-voltage X-ray computed tomography of Fgf[Formula: see text] mice demonstrate dysplastic bulla and ossicles; Fgf[Formula: see text] mice have near-normal morphology. The cochleae of mutant mice appear nearly normal on gross and microscopic inspection. In wild type mice, FGF23 is ubiquitously expressed throughout the cochlea. Measurements from Fgf[Formula: see text] mice do not match the auditory phenotype of Kl-/- mice, suggesting that loss of FGF23 activity impacts the auditory system via mechanisms at least partially independent of KL. Given the extensive middle ear malformations and the overlap of initiation of FGF23 activity and Eustachian tube development, this work suggests a possible role for FGF23 in otitis media.

A sub-nW 2.4 GHz Transmitter for Low Data-Rate Sensing Applications.

This paper presents the design of a narrowband transmitter and antenna system that achieves an average power consumption of 78 pW when operating at a duty-cycled data rate of 1 bps. Fabricated in a 0.18 µm CMOS process, the transmitter employs a direct-RF power oscillator topology where a loop antenna acts as a both a radiative and resonant element. The low-complexity single-stage architecture, in combination with aggressive power gating techniques and sizing optimizations, limited the standby power of the transmitter to only 39.7 pW at 0.8 V. Supporting both OOK and FSK modulations at 2.4 GHz, the transmitter consumed as low as 38 pJ/bit at an active-mode data rate of 5 Mbps. The loop antenna and integrated diodes were also used as part of a wireless power transfer receiver in order to kick-start the system power supply during energy harvesting operation.

A 1.1nW Energy Harvesting System with 544pW Quiescent Power for Next Generation Implants.

This paper presents a nW power management unit (PMU) for an autonomous wireless sensor that sustains itself by harvesting energy from the endocochlear potential (EP), the 70-100 mV electrochemical bio-potential inside the mammalian ear. Due to the anatomical constraints inside the inner ear, the total extractable power from the EP is limited to 1.1-6.25 nW. A nW boost converter is used to increase the input voltage (30-55 mV) to a higher voltage (0.8 to 1.1 V) usable by CMOS circuits in the sensor. A pW Charge Pump circuit is used to minimize the leakage in the boost converter. Further, ultra-low-power control circuits consisting of digital implementations of input impedance adjustment circuits and Zero Current Switching circuits along with Timer and Reference circuits keep the quiescent power of the PMU down to 544 pW. The designed boost converter achieves a peak power conversion efficiency of 56%. The PMU can sustain itself and a duty-cyled ultra-low power load while extracting power from the EP of a live guinea pig. The PMU circuits have been implemented on a 0.18µm CMOS process.

Loss of osteoprotegerin expression in the inner ear causes degeneration of the cochlear nerve and sensorineural hearing loss.

Osteoprotegerin (OPG) is a key regulator of bone remodeling. Mutations and variations in the OPG gene cause many human diseases that are characterized by not only skeletal abnormalities but also poorly understood hearing loss: Paget's disease, osteoporosis, and celiac disease. To gain insight into the mechanisms of hearing loss in OPG deficiency, we studied OPG knockout (Opg(-/-)) mice. We show that they develop sensorineural hearing loss, in addition to conductive hearing loss due to abnormal middle-ear bones. OPG deficiency caused demyelination and degeneration of the cochlear nerve in vivo. It also activated ERK, sensitized spiral ganglion cells (SGC) to apoptosis, and inhibited proliferation and survival of cochlear stem cells in vitro, which could be rescued by treatment with exogenous OPG, an ERK inhibitor, or bisphosphonate. Our results demonstrate a novel role for OPG in the regulation of SGC survival, and suggest a mechanism for sensorineural hearing loss in OPG deficiency.

Quantitative polarized light microscopy of unstained mammalian cochlear sections.

Hearing loss is the most common sensory deficit in the world, and most frequently it originates in the inner ear. Yet, the inner ear has been difficult to access for diagnosis because of its small size, delicate nature, complex three-dimensional anatomy, and encasement in the densest bone in the body. Evolving optical methods are promising to afford cellular diagnosis of pathologic changes in the inner ear. To appropriately interpret results from these emerging technologies, it is important to characterize optical properties of cochlear tissues. Here, we focus on that characterization using quantitative polarized light microscopy (qPLM) applied to unstained cochlear sections of the mouse, a common animal model of human hearing loss. We find that the most birefringent cochlear materials are collagen fibrils and myelin. Retardance of the otic capsule, the spiral ligament, and the basilar membrane are substantially higher than that of other cochlear structures. Retardance of the spiral ligament and the basilar membrane decrease from the cochlear base to the apex, compared with the more uniform retardance of other structures. The intricate structural details revealed by qPLM of unstained cochlear sections ex vivo strongly motivate future application of polarization-sensitive optical coherence tomography to human cochlea in vivo.

Energy extraction from the biologic battery in the inner ear.

Endocochlear potential (EP) is a battery-like electrochemical gradient found in and actively maintained by the inner ear. Here we demonstrate that the mammalian EP can be used as a power source for electronic devices. We achieved this by designing an anatomically sized, ultra-low quiescent-power energy harvester chip integrated with a wireless sensor capable of monitoring the EP itself. Although other forms of in vivo energy harvesting have been described in lower organisms, and thermoelectric, piezoelectric and biofuel devices are promising for mammalian applications, there have been few, if any, in vivo demonstrations in the vicinity of the ear, eye and brain. In this work, the chip extracted a minimum of 1.12 nW from the EP of a guinea pig for up to 5 h, enabling a 2.4 GHz radio to transmit measurement of the EP every 40-360 s. With future optimization of electrode design, we envision using the biologic battery in the inner ear to power chemical and molecular sensors, or drug-delivery actuators for diagnosis and therapy of hearing loss and other disorders.

Proteome of human perilymph.

Current diagnostic tools limit a clinician's ability to discriminate between many possible causes of sensorineural hearing loss. This constraint leads to the frequent diagnosis of the idiopathic condition, leaving patients without a clear prognosis and only general treatment options. As a first step toward developing new diagnostic tools and improving patient care, we report the first use of liquid chromatography-tandem mass-spectrometry (LC-MS/MS) to map the proteome of human perilymph. Using LC-MS/MS, we analyzed four samples, two collected from patients with vestibular schwannoma (VS) and two from patients undergoing cochlear implantation (CI). For each cohort, one sample contained pooled specimens collected from five patients and the second contained a specimen obtained from a single patient. Of the 271 proteins identified with high confidence among the samples, 71 proteins were common in every sample and used to conservatively define the proteome of human perilymph. Comparison to human cerebrospinal fluid and blood plasma, as well as murine perilymph, showed significant similarity in protein content across fluids; however, a quantitative comparison was not possible. Fifteen candidate biomarkers of VS were identified by comparing VS and CI samples. This list will be used in future investigations targeted at discriminating between VS tumors associated with good versus poor hearing.

The role of surface species in chemical vapor deposited carbon nanotubes.

Chemical vapor deposition (CVD) of carbon nanotubes (CNTs) has been investigated using a coupled gas phase and surface chemistry model. This model successfully bridged the gap between the reactor and molecular length scales and allowed individual surface kinetic processes to be identified as growth limiting directly from reactor scale parameters. Carbon nanotube growth rate is a function of the reactor wall temperature such that deposition would occur in the transition region between the hydrogen abstraction and hydrocarbon adsorption limited regimes. Deposition was limited under low reactor temperatures or rich methane conditions by hydrogen abstraction from surface bound hydrocarbons due to the availability of gaseous H(1). At high reactor temperatures and rich hydrogen conditions, the deposition reaction was shown to be limited by hydrocarbon adsorption onto the nanoparticle surface. Optimal process conditions for efficient CNT production are discussed, as well as identifying the limiting reaction steps for the surface chemistry.

Modeling of the carbon nanotube chemical vapor deposition process using methane and acetylene precursor gases.

Chemical vapor deposition of carbon nanotubes (CNTs) in a horizontal tube-flow reactor has been investigated with a fully coupled reactor-scale computational model. The model combined conservation of mass, momentum, and energy equations with gas-phase and surface chemical reactions to describe the evolution of a hydrogen and hydrocarbon feed-stream as it underwent heating and reactions throughout the reactor. Investigation was directed toward steady state deposition onto iron nanoparticles via methane and hydrogen as well as feed-streams consisting of acetylene and hydrogen. The model determines gas-phase velocity, temperature, and concentration profiles as well as surface concentrations of adsorbed species and CNT growth rate along the entire length of the reactor. The results of this work determine deposition limiting regimes for growth via methane and acetylene, demonstrate the need to tune reactor wall temperature to specific inlet molar ratios to achieve optimal CNT growth, and demonstrate the large effect that active site specification can have on calculated growth rate.