[18F]DPA-714 PET imaging for the quantitative evaluation of early spatiotemporal changes of neuroinflammation in rat brain following status epilepticus.

BACKGROUND: Most antiepileptic drug therapies are symptomatic and adversely suppress normal brain function by nonspecific inhibition of neuronal activity. In recent times, growing evidence has suggested that neuroinflammation triggered by epileptic seizures might be involved in the pathogenesis of epilepsy. Although the potential effectiveness of anti-inflammatory treatment for curing epilepsy has been extensively discussed, the limited quantitative data regarding spatiotemporal characteristics of neuroinflammation after epileptic seizures makes it difficult to be realized. We quantitatively analyzed the spatiotemporal changes in neuroinflammation in the early phase after status epilepticus in rats, using translocator protein (TSPO) positron emission tomography (PET) imaging, which has been widely used for the quantitative evaluation of neuroinflammation in several animal models of CNS disease. METHODS: The second-generation TSPO PET probe, [18F]DPA-714, was used for brain-wide quantitative analysis of neuroinflammation in the brains of rats, when the status epilepticus was induced by subcutaneous injection of kainic acid (KA, 15 mg/kg) into those rats. A series of [18F]DPA-714 PET scans were performed at 1, 3, 7, and 15 days after status epilepticus, and the corresponding histological changes, including activation of microglia and astrocytes, were confirmed by immunohistochemistry. RESULTS: Apparent accumulation of [18F]DPA-714 was observed in several KA-induced epileptogenic regions, such as the amygdala, piriform cortex, ventral hippocampus, mediodorsal thalamus, and cortical regions 3 days after status epilepticus, and was reversibly displaced by unlabeled PK11195 (1 mg/kg). Consecutive [18F]DPA-714 PET scans revealed that accumulation of [18F]DPA-714 was focused in the KA-induced epileptogenic regions from 3 days after status epilepticus and was further maintained in the amygdala and piriform cortex until 7 days after status epilepticus. Immunohistochemical analysis revealed that activated microglia but not reactive astrocytes were correlated with [18F]DPA-714 accumulation in the KA-induced epileptogenic regions for at least 1 week after status epilepticus. CONCLUSIONS: These results indicate that the early spatiotemporal characteristics of neuroinflammation quantitatively evaluated by [18F]DPA-714 PET imaging provide valuable evidence for developing new anti-inflammatory therapies for epilepsy. The predominant activation of microglia around epileptogenic regions in the early phase after status epilepticus could be a crucial therapeutic target for curing epilepsy.

Practical Synthesis of [18F]Pitavastatin and Evaluation of Hepatobiliary Transport Activity in Rats by Positron Emission Tomography.

We developed a practical synthetic method for fluorine-18 (18F)-labeled pitavastatin ([18F]PTV) as a positron emission tomography (PET) tracer to assess hepatobiliary transporter activity and conducted a PET scan as a preclinical study for proof-of-concept in rats. This method is a one-pot synthesis involving aromatic 18F-fluorination of an arylboronic acid ester followed by deprotection under acidic conditions, which can be reproduced in general clinical sites equipped with a standard radiolabeling system due to the simplified procedure. PET imaging confirmed that intravenously administered [18F]PTV was rapidly accumulated in the liver and gradually transferred into the intestinal lumen through the bile duct. Radiometabolite analysis showed that [18F]PTV was metabolically stable, and 80% of the injected dose was detected as the unchanged form in both blood and bile. We applied integration plot analysis to assess tissue uptake clearance (CLuptake, liver and CLuptake, kidney) and canalicular efflux clearance (CLint, bile), and examined the effects of inhibitors on membrane transport. Treatment with rifampicin, an organic anion transporting polypeptide inhibitor, significantly reduced CLuptake, liver and CLuptake, kidney to 44% and 64% of control, respectively. In contrast, Ko143, a breast cancer resistance protein inhibitor, did not affect CLuptake, liver but significantly reduced CLint, bile to 39% of control without change in [18F]PTV blood concentration. In addition, we found decreased CLuptake, liver and increased CLint, bile in Eisai hyperbilirubinemic rats in response to altered expression levels of transporters. We expect that [18F]PTV can be translated into clinical application, as our synthetic method does not need special apparatus in the radiolabeling system and PET scan with [18F]PTV can quantitatively evaluate transporter activity in vivo.

PET imaging of 11C-labeled coenzyme Q10: Comparison of biodistribution between [11C]ubiquinol-10 and [11C]ubiquinone-10.

Coenzyme Q10 (CoQ10) plays a key role not only as an essential electron carrier in the mitochondrial electron transport chain, but also as an antioxidant to protect cells from oxidative stress. CoQ10 supplementation is expected to be effective for a variety of diseases. The predominant forms of CoQ10 are the ubiquinol-10 (reduced form) and ubiquinone-10 (oxidized form). Both forms of CoQ10 supplements are commercially available, however, their kinetic difference is still unclear. In order to conduct in vivo analysis of the kinetics of ubiquinol-10 and ubiquinone-10, we succeeded in synthesizing 11C-labeled ubiquinol-10 ([11C]UQL) and ubiquinone-10 ([11C]UQN), respectively. In the present study, we aimed to investigate the kinetics of [11C]UQL and [11C]UQN, both of which were administered via the tail vein of 8-week-old male Sprague-Dawley rats. Whole-body positron emission tomography (PET) imaging was performed to follow the time course of accumulation in the liver, spleen, brain, and other organs. Then, at the two typical time points at 20 or 90 min after injection, we conducted the biodistribution study. Various organs/tissues and blood were collected, weighed and counted with a gamma counter. Percent injected dose per gram of tissue (%ID/g) was calculated as the indicator of the accumulation of each compound. As the results, at both time points, %ID/g of [11C]UQL in the cerebrum, cerebellum, white adipose tissue, muscle, kidney, and testis were higher (P < 0.05) than that of [11C]UQN: at 90-min time point, %ID/g of [11C]UQL in the brown adipose tissue was higher (P < 0.05) than that of [11C]UQN: on the contrary, %ID/g of [11C]UQL in the spleen was lower (P < 0.05) than that of [11C]UQN at 90 min. In a separate study of the metabolite analysis in the plasma, UQL injected into the tail vein of rats was almost unchanged during the PET scanning time, but UQN was gradually converted to the reduced form UQL. Therefore, the uptake values of UQL into the tissues and organs were rather accurate but those of UQN might be the sum of UQN uptake and partly converted UQL uptake. These studies suggested that the accumulation level of administered CoQ10 differs depending on its redox state, and that CoQ10 redox state could be crucial for optimization of the effective supplementation.

A Clinical Quantitative Evaluation of Hepatobiliary Transport of [11C]Dehydropravastatin in Humans Using Positron Emission Tomography.

Various positron emission tomography (PET) probes have been developed to assess in vivo activities in humans of drug transporters, which aid in the prediction of pharmacokinetic properties of drugs and the impact of drug-drug interactions. We developed a new PET probe, sodium (3R, 5R)-3, 5-dihydroxy-7-((1S, 2S, 6S, 8S)-6-hydroxy-2-methyl-8- ((1-[11C]-(E)-2-methyl-but-2-enoyl) oxy) -1, 2, 6, 7, 8, 8a-hexahydronaphthalen-1-yl) heptanoate ([11C]DPV), and demonstrated its usefulness for the quantitative investigation of Oatps (gene symbol SLCO) and Mrp2 (gene symbol ABCC2) in rats. To further analyze the species differences and verify the pharmacokinetic parameters in humans, serial PET scanning of the abdominal region with [11C]DPV was performed in six healthy volunteers with and without an OATP1Bs and MRP2 inhibitor, rifampicin (600 mg, oral), in a crossover fashion. After intravenous injection, [11C]DPV rapidly distributed to the liver and kidney followed by secretion into the bile and urine. Rifampicin significantly reduced the liver distribution of [11C]DPV 3-fold, resulting in a 7.5-fold reduced amount of excretion into the bile and the delayed elimination of [11C]DPV from the blood circulation. The hepatic uptake clearance (CLuptake, liver) and canalicular efflux clearance (CLint, bile) of [11C]DPV (544 ± 204 and 10.2 ± 3.5 µl/min per gram liver, respectively) in humans were lower than the previously reported corresponding parameters in rats (1800 and 298 µl/min per gram liver, respectively) (Shingaki et al., 2013). Furthermore, rifampicin treatment significantly reduced CLuptake, liver and CLint, bile by 58% and 44%, respectively. These results suggest that PET imaging with [11C]DPV is an effective tool for quantitatively characterizing the OATP1Bs and MRP2 functions in the human hepatobiliary transport system.

Human organic anion transporters function as a high-capacity transporter for p-cresyl sulfate, a uremic toxin.

BACKGROUND: Recent clinical studies have shown that increased serum levels of p-cresyl sulfate (PCS), a uremic toxin, are associated with the progression of chronic kidney disease (CKD) and cardiovascular outcomes. Using rat renal cortical slices, we previously reported that the rat organic anion transporter (OAT) could play a key role in the renal tubular secretion of PCS. However, no information is currently available regarding the transport of PCS via human OAT (hOAT) isoforms, hOAT1 and hOAT3. METHODS: Uptake experiments of PCS were performed using HEK293 cells, which stably express hOAT1 or hOAT3. RESULTS: PCS was taken up by hOAT1/HEK293 and hOAT3/HEK293 cells in a time- and concentration-dependent manner. The apparent K m for the hOAT1-mediated transport of PCS was 128 µM, whereas in hOAT3/HEK293, saturation was not observed at the highest tested PCS concentration of 5 mM. Probenecid, an OAT inhibitor, inhibited PCS transport by hOAT1 and hOAT3. The uptake of p-aminohippurate by hOAT1 and estron-3-sulfate by hOAT3 was decreased with increasing PCS concentration. The apparent 50 % inhibitory concentrations for PCS were 690 and 485 µM for hOAT1 and hOAT3, respectively. CONCLUSION: PCS is a substrate for hOAT1 and hOAT3, and hOAT1 and hOAT3 appear to play a physiological role as a high-capacity PCS transporter. Since hOATs are expressed not only in the kidneys, but also in blood vessels and osteoblasts, etc., these findings are of great significance in terms of elucidating the renal clearance, tissue disposition of PCS and the mechanism of its toxicity in CKD.

Bisphosphonate-related osteonecrosis of the palatal torus.

BACKGROUND: Bisphosphonates are widely used for the treatment of osteoporosis and other bone-degrading disorders; however, bisphosphonate therapy is an important risk factor for osteonecrosis of the jaws. METHODS: We report a rare case of bisphosphonate-related osteonecrosis of the palatal torus. RESULTS: The patient was a 72-year-old woman with osteoporosis who had received 35 mg alendronate sodium hydrate once every week for 6 years. She had a 2-month history of oral pain because of intractable mucositis and ulceration of the palatal torus, with no history of malignant disease, radiation therapy, chemotherapy, steroid use, or recent dentoalveolar surgery. A CT scan showed a bony prominence at the midportion of the hard palate with erosion of its cortex. Her condition was diagnosed as stage 2 bisphosphonate-related osteonecrosis of the jaw caused by trauma, and she was advised to discontinue alendronate. She was prescribed oral antibiotics for 5 days and an oral antibacterial rinse. The mucositis with ulceration healed in approximately 10 weeks, but left a small scar. CONCLUSIONS: Although osteonecrosis of the palatal torus associated with bisphosphonate use is a rare condition, otolaryngologists should consider this condition in the differential diagnosis of intractable ulceration of the hard palate.

Multiparametric MR imaging for differentiating between benign and malignant thyroid nodules: initial experience in 23 patients.

PURPOSE: To evaluate multiparametric MR imaging with combined use of apparent diffusion coefficients (ADCs) and time-signal intensity curves (TICs) for discriminating malignant thyroid nodules from benign ones. MATERIALS AND METHODS: ADCs and TICs of 7 benign and 16 malignant nodules were retrospectively analyzed on lesion-by-lesion (overall ADCs and TICs) and pixel-by-pixel (ADC and TIC mapping) bases. ADCs were determined using b-values of 500 and 1000 s/mm(2) . The TICs were classified into 5 patterns on the basis of increment ratio, peak time, and washout ratio. Stepwise approach based on ADC and TIC criteria was used to discriminate between benign and malignant nodules. RESULTS: Overall ADC discriminated undifferentiated carcinomas from papillary carcinomas at 91% accuracy (≤1.3 × 10(-3) mm(2) /s) and differentiated lymphomas from the other malignant nodules at 100% accuracy (≤0.65 × 10(-3) mm(2) /s). Most malignant thyroid nodules had large (≥45%) areas of rapid-uptake TIC profile with small (≤15%) areas of flat TIC profile, or had small (<45%) areas of rapid-uptake TIC profile with large (≥25%) areas of extremely low or low ADCs (≤1.2 × 10(-3) mm(2) /s). Stepwise approach discriminated malignant nodules from benign ones at 91% accuracy. CONCLUSION: The multiparametric MR imaging helps discriminating malignant thyroid nodules from benign ones.

Low-cost high-speed imaging system for observing vocal fold vibration in voice disorders.

PURPOSE: The aim of this study was to establish a method to observe vocal fold vibration using a low-cost high-speed laryngeal imaging system. PROCEDURES: We assembled a high-speed imaging system with a consumer digital camera and a rigid laryngeal endoscope. The camera can shoot digital images at a rate of 1,200 frames per second and be purchased for about USD 1,000 in Japan. RESULTS: We examined the normal and pathological vocal folds of 215 subjects with our new system and analyzed the vocal fold vibration in these subjects by playback of a video and kymograph images. CONCLUSIONS: Our high-speed laryngeal imaging system is highly cost-effective and can be a useful tool for examining the vocal folds of patients with voice disorders.

Correlation Between Skin Autofluorescence and Muscle Activities of Lower Limb in Aging Without Disease and Disability.

Skin autofluorescence is a useful index to estimate the accumulation of advanced glycation end-products in human tissues. Elderly persons with higher skin autofluorescence have lower muscle mass, muscle strength and muscle power, however, little is known about the relationship between the skin autofluorescence level and each muscle activity. We measured the values of skin autofluorescence from five places on a lower limb, and the signals of surface electromyogram during isometric contractions from five muscles on that, simultaneously. The waveforms of surface electromyogram were analyzed by Daubechies-4 wavelet transformation. The value of skin autofluorescence was increased in the proximal part of the lower limb compared with the value of the distal part. The principal component of surface electromyogram activity in a time-frequency domain was lower in the proximal part compared with that of the distal part. There was a weak negative correlation between the value of skin autofluorescence on the gluteal region and the value of the mean wavelet coefficient of the surface electromyogram signals within the gluteus maximus muscle. The higher accumulation of advanced glycation end-products on the gluteal region might suggest the lower muscle activity in aging without disease and disability.

Nano-hydroxyapatite improves intestinal absorption of acetazolamide (BCS Class IV drug)-but how?

We earlier reported that coating poorly water-soluble drugs with nano-hydroxyapatite (nano-HAP) improves bioavailability after oral administration. In the present study, we coated BCS Class IV drug acetazolamide (AZ) with nano-HAP (AZ/HAP formulation), and investigated its bioavailability and nano-HAP's role in promoting it. We tested AZ bioavailability after a single oral dose of the AZ/HAP formulation in rats, followed by a series of in vitro, ex vivo and in vivo testing. The binding state of AZ and nano-HAP was analyzed by gel filtration chromatography. AZ permeability was studied using a Caco-2 cell monolayer assay kit, to test for tight junction penetration, then using an Ussing chamber mounted with intestinal epithelium, both with and without Peyer's patch tissue, to examine the role of intracellular transport. Fluorescence-labeled nano-HAP particles were administered orally in rats to investigate their localization in the intestinal tract. The area under the blood concentration time-curve in rats was about 4 times higher in the AZ/HAP formulation group than in the untreated AZ group. Gel filtration analysis showed AZ and nano-HAP were not bound. The Caco-2 study showed equivalent AZ permeability for both groups, but without significant change in transepithelial electrical resistance (TEER), indicating that tight junctions were not penetrated. In the Ussing chamber study, no significant difference in AZ permeability between the two groups was observed for epithelium containing Peyer's patch tissue, but for epithelium without Peyer's patch tissue, at high concentration, significantly higher permeability in the AZ/HAP formulation group was observed. Fluorescent labeling showed nano-HAP particles were present in both intestinal villi and Peyer's patch tissue 30 min after oral administration. Our results suggest that nano-HAP's enhancement of drug permeability from the small intestine occurs not via tight junctions, but intracellularly, via the intestinal villi. Further study to elucidate the mechanism of this permeability enhancement is required.

Introduction of Otolaryngology Outpatient Examination Training Program for junior residents as part of rural regional medical support in Japan.

Background: Nagasaki Prefecture is located in the most western part of Japan, and there are a considerable number of clinics in its many remote islands and rural areas. Thus, the Regional Medical Support Center in Nagasaki Prefecture dispatches doctors to rural hospitals to provide medical support. We introduced an outpatient training program at these rural hospitals for all residents to improve their clinical training in the field of otorhinolaryngology, whereby one otolaryngologist trains one resident. Methods: This otolaryngology outpatient training program is randomly assigned, and conducted for 4-5 days a year, transported by a helicopter in Nagasaki Prefecture, which is a 30-minute one-way trip. We used a case checklist that included the 35 items that should be experienced and are defined as frequent by the Ministry of Health, Labor and Welfare. We also conducted a survey using an anonymous questionnaire. Results: The survey response rate was 100%. Comparing the experience rate of symptoms between the pre-introduction resident and the post-introduction resident who underwent the otolaryngology outpatient training program, the experience rates of common diseases, including vertigo and otolaryngologic symptoms such as nasal bleeding and hoarseness, significantly increased after the program was introduced (p ≤ .001). Notably, the experience rate of headache, cough/sputum, and vertigo was 100%. Conclusion: Our training program provides a suitable medical environment for the resident and secures a doctor who can provide secondary medical service support. Furthermore, the program will improve the level of primary care provided by the residents in remote island and rural area hospitals.

Clinical evaluation of [18F]pitavastatin for quantitative analysis of hepatobiliary transporter activity.

It is widely accepted that uptake and efflux transporters on clearance organs play crucial roles in drug disposition. Although in vitro transporter assay system can identify the intrinsic properties of the target transporters, it is not so easy to precisely predict in vivo pharmacokinetic parameters from in vitro data. Positron emission tomography (PET) imaging is a useful tool to directly assess the activity of drug transporters in humans. We recently developed a practical synthetic method for fluorine-18-labeled pitavastatin ([18F]PTV) as a PET probe for quantitative evaluation of hepatobiliary transport. In the present study, we conducted clinical PET imaging with [18F]PTV and compared the pharmacokinetic properties of the probe for healthy subjects with or without rifampicin pretreatment. Rifampicin pretreatment significantly suppressed the hepatic maximum concentration and biliary excretion of the probe to 52% and 34% of the control values, respectively. Rifampicin treatment markedly decreased hepatic uptake clearance (21% of the control), and moderately canalicular efflux clearance with regard to hepatic concentration (52% of the control). These results demonstrate that [18F]PTV is a useful probe for clinical investigation of the activities of hepatobiliary uptake/efflux transporters in humans.

Effect of pneumococcal conjugate vaccine on prevalence of otitis media with effusion among children in Vietnam.

PURPOSE: Otitis media with effusion (OME) is common in young children and is associated with Streptococcus pneumoniae infection. We aimed to determine the impact of pneumococcal conjugate vaccine (PCV) introduction on the prevalence of OME and OME associated with vaccine-type (VT) or non-VT. METHODS: Population-based cross-sectional surveys were conducted in pre- (2016) and post-PCV periods (2017, 2018, and 2019) at selected communes in Nha Trang, Vietnam. For each survey, we randomly selected 60 children aged 4-11 months and 60 aged 14-23 months from each commune. Nasopharyngeal sample collection and tympanic membrane examination by digital otoscope were performed. S. pneumoniae was detected and serotyped by lytA qPCR and microarray. Odds ratios (OR) and 95% confidence intervals (CIs) were calculated using Firth's logistic regression, stratified by age group. RESULTS: Over the four surveys, 2089 children had a bilateral ear examination. Compared to pre-PCV, the prevalence of OME reduced in 2018 (OR 0.51, 95 %CI 0.28-0.93) and in 2019 (OR 0.53, 95 %CI 0.29-0.97) among the <12-month-olds, but no significant reduction among the 12-23-month-olds. The prevalence of OME associated with VT pneumococcus decreased in 2018 and 2019 (2018: OR 0.14, 95 %CI 0.03-0.55; 2019: OR 0.20, 95 %CI 0.05-0.69 in the <12-months-olds, 2018: OR 0.05, 95 %CI 0.00-0.44, 2019: OR 0.41, 95 %CI 0.10-1.61 in the 12-23-months-olds). The prevalence of OME associated with non-VT pneumococcus increased in the 12-23-month-olds in 2017 (OR 3.09, 95 %CI 1.47-7.45) and returned to the pre-PCV level of prevalence in 2018 and 2019 (OR 0.94, 95 %CI 0.40-2.43 and 1.40, 95 %CI 0.63-3.49). CONCLUSION: PCV10 introduction was associated with a reduction of OME prevalence in infants but not in older children.

Histidine146 of human serum albumin plays a prominent role at the interface of subdomains IA and IIA in allosteric ligand binding.

Fatty acids are endogenous ligands of human serum albumin (HSA) that induce conformational changes and participate in allosteric ligand binding to HSA. In a previous study, we showed that, when myristate (MYR) is present, the binding of [(14) C]ketoprofen (KP) to subdomain IA of HSA was increased, indicating that, when MYR binds to HSA, a new binding site in formed in that region. Meanwhile, an N-B transition has been reported to increase the binding of ligands at alkaline pH when the status of albumin is the B-conformer. Six histidine single mutants of HSA, H9A, H39A, H67A, H105A, H128A and H146A were produced and photolabeled with [(14) C]KP at pH 6.5, 7.4 and 8.2 and the role of each histidine in causing the N-B transition induced allosteric ligand binding was examined. Cyanogen bromide cleavage of the photolabeled native HSA showed that subdomain IA was the site of the allosteric binding of KP at pH 8.2. From the photolabeling results, H146 was found to play a prominent role whilst H128 played little or no role in the allosteric binding. However, the remaining 4 mutants did not show a clear photolabeling pattern that was similar to either native HSA or H146A and, as a result, no firm conclusions can be made. An additional histidine mutant, H146I, was produced to confirm the results for H146A. A similar experiment using H146I showed that a benzene ring-like structure at position 146 is required for the allosteric ligand binding to occur.

Cavernous hemangioma of the accessory parotid gland.

We report a rare case of a hemangioma arising from the accessory parotid gland. The patient, a 45-year-old woman, complained of a right midcheek mass. Magnetic resonance imaging showed a well-defined mass located in the right buccal space, anterior to the masseter muscle, and adjacent to the Stensen duct. The mass had high T2-weighted signal intensity and showed strong patchy enhancement with gadolinium. This mass was surrounded by a common capsule with the accessory parotid gland. These findings indicated a hemangioma originating from the accessory parotid gland. The mass was completely removed by an intraoral approach without postoperative facial palsy, skin deformity, and difficulty in secreting saliva. Histologic examination of the tumor revealed multiple, thin-walled, and dilated blood vessels, confirming the diagnosis of a cavernous hemangioma. Magnetic resonance imaging was extremely useful in diagnosing the mass as a hemangioma before surgery, clarifying relationships between the mass and adjacent structures and determining the surgical approach to the mass.

Prevalence and characteristics of children with otitis media with effusion in Vietnam.

PURPOSE: Otitis media with effusion (OME) commonly occurs and persists in young children. It can cause hearing impairment and damage to the tympanic membrane without treatment. We aimed to determine the prevalence and association of Streptococcus pneumoniae in the nasopharynx of healthy children before the introduction of a pneumococcal conjugate vaccine. METHODS: In October 2016, nasopharyngeal swabs collection and otoscope examinations by an otolaryngologist were conducted in children aged less than 24 months in Nha Trang, Vietnam. OME was diagnosed as the presence of middle ear fluid using a digital otoscope equipped with a pneumatic otoscope. Quantitative PCR targeting pneumococci-specific lytA (the major autolysis gene) and bacterial culture were performed to detect S. pneumoniae. The point prevalence of OME in the study area was estimated. The association between OME and S. pneumoniae in the nasopharynx was evaluated using a multivariable logistic regression model. RESULTS: Among the 274 children who underwent bilateral ear examinations and nasopharyngeal swab collections, 47 had OME (17.2%, 95% confidence interval [CI] 12.9-22.1%) and 96 were colonized with S. pneumoniae (35.0%, 29.4-41.0%). OME and nasopharyngeal S. pneumoniae carriage were positively associated in children aged less than 12  months (adjusted odds ratio [aOR] 3.83, 1.40-10.51). Day-care attendance and living in a rural area were independently associated with OME (aOR 5.87, 2.31-14.91, and aOR 3.77, 1.58-8.99, respectively). CONCLUSIONS: The nasopharyngeal pneumococcal carriage was associated with OME among children aged  <12 months. A further study after introducing a pneumococcal conjugate vaccine (PCV) is required to better understand the effect of PCV and S. pneumoniae carriage on OME in young children.

Immunohistochemical Expression of Choline Acetyltransferase and Catecholamine-Synthesizing Enzymes in Head-and-Neck and Thoracoabdominal Paragangliomas and Pheochromocytomas.

Paragangliomas (PGLs) are neural-crest-derived, non-epithelial neuroendocrine tumors distributed along the parasympathetic and sympathetic nerves. Head-and-neck PGLs (HNPGLs) have been recognized as nonchromaffin, nonfunctional, parasympathetic tumors. By contrast, thoracoabdominal paragangliomas and pheochromocytomas (PPGLs) are chromaffin, functional, sympathetic tumors. Although HNPGLs and PPGLs have the same histological structure, the zellballen pattern, composed of chief and sustentacular cells surrounded by abundant capillaries, the pathobiological differences between these types of PGLs remain unclarified. To determine the phenotypic features of these PGLs, we performed an immunohistochemical study using specific antibodies against choline acetyltransferase (ChAT), an enzyme involved in acetylcholine synthesis, and enzymes for the catecholamine-synthesis, tyrosine hydroxylase (TH), and dopamine beta-hydroxylase (DBH), in 34 HNPGLs from 31 patients, 12 thoracoabdominal PGLs from 12 patients, and 26 pheochromocytomas from 22 patients. The expression of ChAT, TH, and DBH was 100%, 23%, and 10% in the HNPGLs; 12%, 100%, and 100% in the pheochromocytomas; and 25%, 67%, and 100% in the thoracoabdominal PGLs, respectively. These results designate HNPGLs as acetylcholine-producing parasympathetic tumors, in contrast to PPGLs being catecholamine-producing tumors. The other most frequently used neuroendocrine markers are synaptophysin and chromogranin A expressed 100% and 80%, respectively, and synaptophysin was superior to chromogranin A in HNPGLs. This is the first report of HNPGLs being acetylcholine-producing tumors. Immunohistochemistry of ChAT could be greatly useful for pathologic diagnosis of HNPGL. Whether measurement of acetylcholine levels in the blood or urine could be a tumor marker of HNPGLs should be investigated soon.

The Diagnostic Dilemma of GATA3 Immunohistochemistry in Pheochromocytoma and Paraganglioma.

Although GATA3 has been recognized as a useful marker for mammary and urothelial carcinomas, there is large variation in GATA3 expression detected in pheochromocytoma (PC) and paraganglioma (PGL), from 90% to less than 5%. For GATA3 to be a useful diagnostic marker for PCCs/PGLs, the reasons for such discrepancy must be elucidated. Thus, we compared different immunohistochemistry protocols. Three protocols for GATA3 immunohistochemistry, including the use of an automated slide stainer or manual staining with an autoclave and EDTA buffer vs citric acid buffer, were compared. Whole sections of paraffin-embedded tumors, including 30 PCCs, 37 PGLs including 15 head and neck PGLs, 5 retroperitoneal PGLs, 17 urinary bladder PGLs, and 14 neuroblastoma group tumors, were examined and compared with mammary and urothelial carcinoma sections as positive controls. Using the automated slide stainer (Benchmark ULTRA; Ventana Medical Systems) with both buffers, mammary and urothelial carcinomas demonstrated strong GATA3 positivity; however, PCCs/PGLs showed negative or weak heterogeneous staining. Manual staining with an autoclave for antigen retrieval resulted in increased GATA3 immunoreactivity in all head and neck PGLs, all retroperitoneal PGLs, 88% of urinary PGLs, 17% of PCCs, and all neuroblastomas, except for ganglion cells. The normal adrenal medulla stained weakly and heterogeneously. In conclusions, immunohistochemistry for GATA3 in PCCs/PGLs requires stronger antigen retrieval than that in mammary and urinary carcinomas. This finding is especially important to consider if GATA3 is applied for the differential diagnosis of PGLs in unusual sites as supplemental data to the expression of catecholamine-synthesizing enzymes.

Characteristics of Lower Limb Muscle Activity in Elderly Persons After Ergometric Exercise.

This study examined the characteristics of lower limb muscle activity in elderly persons after ergometric pedaling exercise for 1 month. To determine the effect of the exercise, surface electromyography (SEMG) of lower limb muscles was subjected to Daubechies-4 wavelet transformation, and mean wavelet coefficients were compared with the pre-exercise coefficients and the post-exercise coefficients in each wavelet level. The characteristics of muscle activity after pedaling exercise were also compared between the elderly subjects and young subjects. For the elderly subjects, the mean wavelet coefficients were significantly decreased in the tibialis anterior and the gastrocnemius medialis at wavelet levels of 3, 4, and 5 (125-62.5, 62.5-31.25, and 31.25-15.625 Hz, respectively), by pedaling exercise. However, the mean power of wavelet levels of 2 and 3 (250-125 and 125-62.5 Hz) within the rectus femoris and the biceps femoris were significantly increased in the young subjects. The effect of pedaling exercise is different from the effects of heavy-resistance training. It was suggested that the muscle coordination, motor unit (MU) firing frequency, and firing fiber type of lower limb muscles are changed with the different characteristics between elderly and young persons by pedaling exercise for 1 month.

Paraganglioma of the carotid body and intrapericardium.

A 65-year-old Japanese woman with an intrapericardial tumor and neck tumor was admitted to our hospital. Intrapericardial tumor had not been resected because of massive bleeding from the hypervascular tumor and its invasion into the pericardium, ascending aorta, and pulmonary artery. The neck tumor had been successfully resected, and paraganglioma was pathologically diagnosed. Abnormal accumulation in the intrapericardial tumor was seen with 123I-metaiodobenzylguanidine scintigraphy. Moreover, gene mutation of succinate dehydrogenase type D was found. Finally, paraganglioma of the carotid body and intrapericardium was diagnosed. .