Human periodontal ligament cell sheets cultured on amniotic membrane substrate.

OBJECTIVE: Periodontal ligament (PDL) cells and their substrates play key roles in periodontal regeneration. However, there has been no report on the use of amniotic membrane (AM) as a substrate for culturing PDL cells. In the current study, we conducted an analysis of PDL cells cultivated on AM to determine the distribution of factors responsible for maintaining the characteristics of PDL. MATERIALS AND METHODS: Amniotic membrane was obtained from women undergoing cesarean sections, whereas PDL tissue was obtained from human maxillary third molars. The harvested PDL cells were maintained in explant culture for three or four passages, following which they were cultured on AM. RESULTS: After 3 weeks of culture, the PDL cells had grown well on AM. Immunofluorescence showed that these cells were capable of proliferating and potentially maintaining their PDL-like properties. In addition, strong cell-cell adhesion structures, namely desmosomes and tight junctions, were shown to be present between cells. Electron microscopy images showed that the cultured PDL cells had differentiated and proliferated on AM with lateral conjugation and adhesion to AM. CONCLUSION: We conclude that AM may represent a suitable substrate for culturing PDL cells and that PDL cells cultured on AM show sheet formation.

Nonunity permeability in metamaterial-based GaInAsP/InP multimode interferometers.

We demonstrated an InP-based optical multimode interferometer (MMI) combined with metamaterials consisting of minute split-ring resonators (SRRs) arrayed on the MMI. The MMI could operate at an optical fiber communication wavelength of 1.5 µm. Magnetic resonance occurred between the SRR metamaterial and light at 1.5 µm, and the relative permeability of the metamaterial increased to 2.4 around this wavelength. This result shows that it is possible to use new materials with nonunity permeability to construct semiconductor-based photonic devices.

The top-down mechanism for body-mass-abundance scaling.

Scaling relationships between mean body masses and abundances of species in multitrophic communities continue to be a subject of intense research and debate. The top-down mechanism explored in this paper explains the frequently observed inverse linear relationship between body mass and abundance (i.e., constant biomass) in terms of a balancing of resource biomasses by behaviorally and evolutionarily adapting foragers, and the evolutionary response of resources to this foraging pressure. The mechanism is tested using an allometric, multitrophic community model with a complex food web structure. It is a statistical model describing the evolutionary and population dynamics of tens to hundreds of species in a uniform way. Particularities of the model are the detailed representation of the evolution and interaction of trophic traits to reproduce topological food web patterns, prey switching behavior modeled after experimental observations, and the evolutionary adaptation of attack rates. Model structure and design are discussed. For model states comparable to natural communities, we find that (1) the body-mass abundance scaling does not depend on the allometric scaling exponent of physiological rates in the form expected from the energetic equivalence rule or other bottom-up theories; (2) the scaling exponent of abundance as a function of body mass is approximately -1, independent of the allometric exponent for physiological rates assumed; (3) removal of top-down control destroys this pattern, and energetic equivalence is recovered. We conclude that the top-down mechanism is active in the model, and that it is a viable alternative to bottom-up mechanisms for controlling body-mass-abundance relations in natural communities.

Endobronchial Topical Amphotericin B Instillation for Pulmonary Chromomycosis After Lung Transplantation: A Case Report.

We report a very rare case of pulmonary chromomycosis caused by Scedosporium prolificans that developed after lung transplantation and was successfully treated with endobronchial topical amphotericin B instillation. The subject was a woman in her 50s with a history of bilateral lobar lung transplantation from living donors for idiopathic pulmonary hypertension. Eight years after the lung transplantation, chest radiography X-ray and computed tomography showed an abnormal shadow in the right lung. Bronchoscopic findings showed obstruction by a fungal component at the laterobasal bronchus B9. She was diagnosed with pulmonary chromomycosis after S. prolificans was detected in the bronchial aspirate. Systemic antifungal treatment with itraconazole was ineffective. Therefore, we administered topical amphotericin B weekly via endobronchial instillation and replaced oral itraconazole with voriconazole. The endobronchial procedure was safe and tolerable. Bronchial obstruction improved after three 3 instillations. We continued topical amphotericin B instillation once every 3 months for 2 years, and the abnormal shadow nearly disappeared. This case report describes infection by S. prolificans, which rarely becomes an etiologic agent in lung transplant patients, and shows that endobronchial topical amphotericin B instillation is a therapeutic option when systemic antifungal treatment is ineffective.

The effect of blood glucose concentration on labile A1c in diabetic patients.

We investigated the magnitude of labile A1c in total A1c measured rapidly by a chromatographic method, and whether or not there was an effect of blood glucose before and after a meal on labile A1c in 94 type I and 178 type II diabetic subjects. There were strong correlations between serum glucose and labile A1c both in type I (r = 0.76, P less than 0.001) and in type II diabetic subjects (r = 0.72, P less than 0.001). These relationships did not change before and after the meal. As labile A1c increased in proportion to blood glucose, it could be calculated from the blood glucose level in simultaneous blood samples. In type I diabetic subjects, below the 100-mg/dl glucose level labile a was negligible, and above 100 mg/dl about 0.35% labile A1c was increased every 50-mg/dl increment of glucose. In type II diabetic subjects, below a 150-mg/dl glucose level labile A1c was in the normal range (0.58 +/- 0.15%), and above the 150-mg/dl glucose level every 50-mg/dl increment of glucose increased about 0.3% of labile A1c. If this process is used, stable A1c can be calculated easily from total A1c and coincident serum glucose, even though labile A1c is not removed by incubation.

Morphological characteristics of the temporomandibular joint in the pouch young of the Tasmanian devil.

We recently reported the absence of the articular disc, which is a constant structure in mammals, in the temporomandibular joint of the adult Tasmanian devil. However, whether the articular disc disappears with growth of the animal was unknown. The aim of this study was to determine whether a pouch young of the Tasmanian devil has the articular disc. The temporomandibular joint of a fresh carcass of the pouch young, whose crown-rump length was 43 mm, was examined microscopically and by microcomputed tomography. The absence of the articular disc in the pouch young temporomandibular joint was histologically confirmed. It is suggested that the articular disc of the Tasmanian devil is naturally absent.

An autosomal dominant posterior polar cataract locus maps to human chromosome 20p12-q12.

We assigned the locus for a previously reported new type of autosomal dominant posterior polar cataract (CPP3) to 20p12-q12 by a genome-wide two-point linkage analysis with microsatellite markers. CPP3 is characterized by progressive, disc-shaped, posterior subcapsular opacity. The disease was seen in 10 members of a Japanese family and transmitted in an autosomal dominant fashion through four generations. We obtained a maximum lod score (Zmax) of 3.61 with a recombination fraction (theta) of 0.00 for markers D20S917, D20S885 and D20S874. Haplotype analysis gave the disease gene localization at a 15.7-cM interval between D20S851 and D20S96 loci on chromosome 20p12-q12. Since the BFSP1 that encodes the lens-specific beaded filament structural protein 1 (filensin) has been mapped around the CPP3 region, we performed sequence analysis on its entire coding region. However, no base substitution or deletion was detected in the CPP3 patients. The mapping of the CPP3 locus to 20p12-q12 not only expands our understanding of the genetic heterogeneity in autosomal dominant posterior polar cataracts but also is a clue for the positional cloning of the disease gene.

Ultrastructure of retina of manganese-deficient rats.

PURPOSE: To elucidate some biologic functions of manganese in the retina. METHODS: Three-week-old weanling Wistar Kyoto rats were used. Manganese-deficient rats were fed a manganese-deficient solid diet containing 0.23 mg manganese/100 g diet and all other nutrients. Control rats were fed a solid diet with 2.9 mg manganese/100 g diet. The retinas were examined by electron microscopy in the 12th, 18th, and 30th months of experimentation. RESULTS: There was a statistically significant decrease in the plasma manganese levels in manganese-deficient animals compared to controls. In rats fed a manganese-deficient diet for 12 months, photoreceptor cells showed karyopyknosis-like changes of nuclei and a decrease in size and number of outer segments. Rats fed a manganese-deficient diet for 18 months showed a complete loss of photoreceptor cells, and the inner nuclear layer nuclei came in direct contact with the retinal pigment epithelium. Rats with manganese deficiency of 30 months showed invasion by capillaries and processes of Müller-like cells from the sensory retina into the retinal pigment epithelium. In the sensory retina, Müller-like cells proliferated, and neural cells disappeared. CONCLUSIONS: Because manganese is related to Mn superoxide in the mitochondrial matrix and to protein and glycogen metabolism, manganese deficiency may disturb the renewal of photoreceptor outer segment discs, and the decrease in antioxidant action caused by a lower level of Mn superoxide dismutase may accelerate the damage to photoreceptor cells. After neural cell loss, Müller-like cells may proliferate. Manganese appears to be essential for maintaining photoreceptor cells.

Enzyme-synthetic approach to demonstration of phosphorylase activity in the living rabbit cornea.

An enzyme-synthetic method of demonstrating phosphorylase was applied to the living rabbit cornea, and polyglucose particles synthesized from glucose-1-phosphate in vivo were studied electron microscopically. In the corneas in which the medium for phosphorylase was applied from the anterior chamber or the bulbar subconjunctiva, synthesized polyglucose particles were found in the cytoplasmic matrices of the epithelium. When the medium was deposited in the conjunctival sac, a few synthesized polyglucose particles were found in the cytoplasmic matrices of only the superficial layer of the corneal epithelium. These findings suggest that metabolites for glycogen metabolism come mainly from the aqueous humor in the anterior chamber. The polyglucose particles synthesized by the enzyme-synthetic method in vivo resemble native glycogen particles. In addition, these particles were not overproduced because the synthesis of polyglucose is probably regulated in vivo.

Human spleen insoluble fibrinolytic proteinase acting at neutral pH: its partial purification and characterization.

An insoluble fibrinolytic enzyme with a molecular weight of approximately 30,000, was purified from the human spleen. A single protein band possessing fibrinolytic activity was obtained on polyacrylamide gel disk electrophoresis at pH 4.5. The enzyme, tentatively termed spleen fibrinolytic proteinase (SFP), degraded fibrinogen at neutral pH following Michaelis-Menten kinetics. The fibrinogenolytic activity was not inhibited by t-AMCHA, a specific plasmin inhibitor. SFP barely degraded certain synthetic ester or polypeptide substrates for trypsin, chymotrypsin, plasma, Xa, elastase and collagenase. These results indicate a different nature for SFP compared to other enzymes examined. SFP was found to digest no elastin and its fibrinogenolytic activity was strongly inhibited by STI, indicating that it was not an elastase. SFP required neither Zn++ nor CA++ for its fibrinogenolytic activity, indicating that it differed from metal-dependent proteinases such as collagenase. SFP was inhibited by DFP but not by TLCK, suggesting that it contains an active serine residue, but no trypsin type histidine at its active center. These results appear to show that SFP is a unique proteinase in the spleen, which is capable of degrading fibrin and fibrinogen at neutral pH.

Involvement of NK2 receptors rather than NK1 receptors in bronchial hyperresponsiveness induced by allergic reaction in guinea-pigs.

1. In this study, the role of neuropeptides in antigen-induced bronchoconstriction and bronchial responsiveness in guinea-pigs was evaluated by use of phosphoramidon, the inhibitor of neutral endopeptidases (NEP), the NK1 receptor antagonist, FK888, and the dual NK1/NK2 receptor antagonist, FK224. The role of endogenous tachykinins in bronchial hyperresponsiveness induced by inhaled capsaicin was also observed with FK888 and FK224. 2. Allergic bronchoconstriction and bronchial responsiveness was evoked by inhalation of ovalbumin (OA), and increasing doses of methacholine were inhaled at 5-min intervals for 30 min after OA challenge in passively sensitized and artificially ventilated guinea-pigs. Animals were treated with a 30 s inhalation of phosphoramidon (10(-3)M) or saline 10 min before the OA challenge. FK888 (1.0 or 10 mg kg-1) or FK224 (1.0 or 10 mg kg-1) was administered intravenously 5 min before the OA challenge. 3. Treatment with phosphoramidon did not alter the increase in the lateral pressure at the tracheal tube (Pao) caused by OA inhalation or the increase in bronchial response to methacholine following the allergic reaction. Pretreatment with FK224 did not inhibit the increase in Pao after antigen provocation but did significantly inhibit antigen-induced bronchial hyperresponsiveness in a dose-dependent manner, while FK888 did not affect either allergic bronchoconstriction or post-allergic bronchial hyperresponsiveness. 4. Histamine, 25, 50, 100 or 200 micrograms ml-1 was inhaled for 20 s at 5-min intervals in non-sensitized guinea-pigs which were pretreated with inhalation of subthreshold dose of capsaicin (10(-7) M). FK888 or FK224, each at a dose of 0.1 or 1.0 mg kg-1, or vehicle was given to guinea-pigs intravenously 3 min before inhalation of capsaicin. The capsaicin inhalation significantly potentiated bronchial responsiveness to histamine, compared with control. The capsaicin-induced bronchial hyperresponsiveness was completely blocked by FK224 in a dose-dependent manner but not by FK888. 5. These results suggest that NK2 receptors rather than NK1 receptors may play an important role in bronchial hyperresponsiveness induced by antigen challenge as well as capsaicin while tachykinins do not play a primary role in the acute bronchospasm elicited by antigen challenge in passively sensitized guinea-pigs.

A partial involvement of histamine in ultrasonically nebulized distilled water-induced bronchoconstriction in guinea-pigs.

1. Inhalation of ultrasonically nebulized distilled water (UNDW) can induce bronchoconstriction only in asthmatics, but mechanisms of the response are not well known. We recently reported a guinea-pig model of UNDW-induced bronchoconstriction (UNDW-IB) in which UNDW induces bronchoconstriction when UNDW is inhaled 20 min after a challenge with aerosolized ovalbumin (OA) in passively sensitized, anaesthetized and artificially ventilated guinea-pigs. 2. To elucidate the role of histamine in the UNDW-IB, we examined the effects of antihistamines, diphenhydramine hydrochloride (DH) and chlorpheniramine maleate (CP), and measured histamine content in bronchoalveolar lavage fluid (BALF) in the animal model. 3. DH in doses of 0.1, 1.0 and 10 mg kg(-1) and CP in doses of 0.01, 0.1 and 1.0 mg kg(-1) administered intravenously 15 min after the OA challenge partially reduced the UNDW-IB at 1 and 2 min after the UNDW inhalation in a dose-dependent manner. Histamine content in BALF recovered 10 min after the UNDW inhalation following the OA provocation was significantly increased compared with that after saline inhalation and before the UNDW inhalation following the OA challenge. 4. Intravenous atropine in a dose of 0.5 mg kg(-1) or inhaled disodium cromoglycate in concentrations of 1 and 10 mg ml(-1) did not alter the UNDW-IB. 5. These results suggest that histamine is involved in part in the UNDW-IB in our animal model.

Analysis of Galpha protein recognition profiles of angiotensin II receptors using chimeric Galpha proteins.

Receptors with a heptahelical structure initiate signal transduction by interacting with specific Galpha proteins. The aim of this study was to analyze the ability of type 1 (AT1) and type 2 (AT2) angiotensin receptors to recognize the receptor coupling regions of Galpha proteins using our previously described technique (Ikezu, T., Okamoto, T., Komatsuzaki, K., Matsui, T., Martyn, J.A.J., Nishimoto, I., 1996. Negative transactivation of cAMP response element by familial Alzheimer's mutants of APP. EMBO J. 15, 2468-2475; Komatsuzaki, K., Murayama, Y., Giambarella, U., Ogata, E., Seino, S., Nishimoto, I., 1996. A novel system that reports the G-proteins linked to a given receptor: a study of the type 3 somatostatin receptor. FEBS Lett. 406, 165-170). Chimeric Galphas protein constructs, whose receptor binding regions contained sequences from the four major families of Galpha proteins (Galphaq, Galphai, Galpha12, Galphas), were cotransfected with AT1 or AT2 receptors in COS cells, then stimulated with angiotensin II (Ang II). Changes in cellular cAMP were assayed on cell lysates by enzyme immunoassay. In the case of the Galphaq family, cotransfection of AT1 with Galpha11/Galphas, Galpha14/Galphas, Galpha16/Galphas, elicited significant increases in cAMP after agonist stimulation. Confirmatory results were found using an independent [35S]GTPgammaS binding assay. Further examination using chimeric G proteins for Galpha12 proteins and Galphai family proteins provided evidence that the AT1 receptor can recognize sequences from Galpha12, Galphai1/i2, Galphaz, Galphao, while both receptors interacted with Galphai3. These results provide a Galpha protein recognition database for both AT1 and AT2 receptors, which may be important for understanding the full spectrum of cellular responses mediated by the hormone Ang II.

Adrenal angiotensin II type 1 and type 2 receptors in Cushing's and Conn's syndromes.

Cushing's and Conn's syndromes are well recognised endocrine diseases, but the pathobiology of the tumors causing these disorders is unclear. In this study we examined AT1 and AT2 gene expression in adrenal adenomas of Cushing's and Conn's syndromes. AT1 and AT2 receptor mRNA, as well as alternatively spliced AT1 transcripts, were detected by RT-PCR using adjacent adrenal cortex tissue as controls. Whereas no consistent differences in AT1 mRNA were seen compared to control adrenal cortex, AT2 mRNA levels were significantly decreased in the adenomas of Cushing's and Conn's syndromes. No changes in alternative splicing of AT1 mRNA were observed in the adrenal tumors. The fact that no consistent changes were seen in AT1 mRNA or its splicing, whereas AT2 mRNA were reduced in both forms of hormone producing adrenal tumor suggests that the AT2 receptor, rather than the AT1 subtype, may be correlated with adrenal tumorigenesis.

Cortisol secretion in relation to body fat distribution in obese premenopausal women.

Urinary cortisol output and serum cortisol concentrations were measured in the steady state, under "field" conditions, and during standardized inhibitory and stimulatory tests in premenopausal, obese women, and were analyzed in relation to adipose tissue distribution. Urinary cortisol output was increased under field conditions in women with an elevated waist to hip circumference ratio (WHR) and, in particular, in women with a large abdominal sagittal diameter, indicating visceral fat accumulation. However, dexamethasone inhibition of cortisol secretion was normal. Stimulation with corticotropin analogue and with physical (cold-pressor test) or mental (color-word or mathematic) stress tests also showed elevated responses of serum cortisol, but not of prolactin or growth hormone concentrations. It is suggested that women with visceral fat accumulation have elevated cortisol secretion due to an increased sensitivity along the hypothalamic-pituitary-adrenal axis, and that this may be causing their abnormal fat depot distribution.

Effects of angiotensin inhibitors on renal injury and angiotensin receptor expression in early hypertensive nephrosclerosis.

Angiotensin converting enzyme inhibitors (ACEI) are known to inhibit the progression of established renal failure. The aim of this study was to compare the efficacy of an ACEI and an AT1 receptor antagonist (AT1R-Ant) in preventing the development of renal disease, at an early stage of hypertensive nephrosclerosis. SHRSP/Izm rats (n = 61) were treated from 10 wk until 22 wk with the ACEI delapril (40 mg/kg/d) or the AT1R-Ant candesartan cilexetil (1 mg/kg/d). Proteinuria, and structural/ultrastructural changes were assessed at 14 and 22 wk. Treatment with either agent resulted in reductions in blood pressure and cardiovascular hypertrophy. Neither proteinuria nor major renal histological changes were evident at 14 wk. At 22 wk, however, proteinuria accompanied by nephrosclerotic changes was seen in the untreated SHRSP/Izm. Treatment with either ACEI or AT1R-Ant resulted in similar reductions in proteinuria (untreated, 32.2 +/- 7.4; delapril-treated, 5.5 +/- 1.2; candesartan-treated, 3.9 +/- 0.3 mg/100 g/d). Prominent sclerosis of small-to-medium sized renal arteries was seen in the untreated SHRSP/Izm at 22 wk, but was similarly attenuated by the ACEI and AT1R-Ant. The glomerular ultrastructure was comparable between the two groups. No significant changes in renal AT1a or AT1b receptor subtype mRNA expression were seen throughout the course of the study. In contrast, a decrease in AT2 receptor mRNA was seen in the drug-treated groups at 14 wk but not at 22 wk. These results suggest that both ACEI and AT1R-Ant have similar efficacy in attenuating the onset of renal injury in early hypertensive nephrosclerosis, and that treatment with either agent is associated with a transient decrease in AT2 receptor mRNA expression.

Bifemelane in the treatment of central retinal artery or vein obstruction.

Patients with retinal artery or vein obstruction received 150 mg of bifemelane hydrochloride daily for 1 to 11 months and other treatment (urokinase, steroid hormone, hyperbaric oxygen, or laser photocoagulation) or the latter without bifemelane. After treatment with bifemelane, improvements in visual acuity were seen in 12 of 17 eyes with central retinal artery obstruction, in three of four eyes with central retinal vein obstruction, and in five of nine eyes with branch retinal vein obstruction. Among the patients not receiving bifemelane, improvements were seen in two of six patients with central retinal artery disease, in 14 of 27 eyes with central retinal vein obstruction, and in 13 of 28 eyes with branch retinal vein obstruction. It is concluded that bifemelane can be used to improve visual acuity in many patients with central retinal artery obstruction.

Genetically distinct autosomal dominant posterior polar cataract in a four-generation Japanese family.

PURPOSE: To describe the clinical findings of a form of posterior polar cataract in a large Japanese family and to determine whether the posterior polar cataract is causally related to other autosomal dominant cataracts with known genes, chromosomal locations, or both. METHODS: Systemic and ocular histories were obtained and comprehensive ophthalmic examinations were performed in 15 of 37 members of the Japanese family. The posterior polar cataract was transmitted in an autosomal dominant manner through four generations. Although there is some variation in the degree of opacification, the posterior polar cataract in this family is characterized by progressive disk-shaped posterior subcapsular opacities. Genetic linkage analysis was performed with 41 polymorphic microsatellite markers located in chromosomal regions known for linkage to cataracts. Genomic DNA extracted from the 15 individuals was amplified by polymerase chain reaction, the genotype at the marker loci was determined in each family member, and the lod score was calculated at each locus. RESULTS: Significant linkage of the posterior polar cataract was ruled out from the following 10 loci or chromosomal regions: 16q22 and 1p36, to which two forms of autosomal dominant posterior polar cataract have been assigned: 1q21-q25, 2q33-q35, 13cen, 17p13, 17q11-q12, 17q24, 21q22, and 22q, which are the regions responsible for other autosomal dominant congenital cataracts. CONCLUSIONS: This study confirms the genetic heterogeneity of autosomal dominant posterior polar cataracts and demonstrates that the posterior polar cataract in this Japanese family is phenotypically and genetically distinct from previously mapped cataracts.

A family with optic atrophy and congenital hearing loss.

A 37-year-old woman had optic atrophy in both eyes and low-tone hearing disturbance of both ears noted after 34 years of age. Her visual acuity was 0.5 in the right eye and 0.6 in the left. The visual fields of both eyes showed slight progressive concentric narrowing. Hearing loss was gradually progressive. Her 13-year-old daughter also had optic atrophy in both eyes and low-tone hearing loss in both ears after 11 years of age. Her visual acuity was 0.8 in the right eye and 1.0 in the left. Her visual fields showed slight concentric narrowing. She had enlarged blind spots in both eyes. The mother and her daughter had deuteranomaly. Family history showed that the father, one brother and three sisters of the mother had congenital hearing loss. No other cause for the optic nerve atrophy and hearing disturbance could be found except heredity.

Conradi-Hünermann syndrome with ocular anomalies.

We report a Japanese girl with the Conradi-Hünermann form of chondrodysplasia punctata and anterior segment malformations characteristic of Axenfeld-Rieger syndrome. The patient also had cataracts and unilateral optic atrophy. A possible role for homeobox-containing genes in the etiology of this type of chondrodysplasia punctata is suggested as an explanation for the coincidence of these two syndromes.