Na+/K+ ATPase α1 and ß3 subunits are localized to the basolateral membrane of trophectoderm cells in human blastocysts.

STUDY QUESTION: Is there a relation between specific Na+/K+ ATPase isoform expression and localization in human blastocysts and the developmental behavior of the embryo? SUMMARY ANSWER: Na+/K+ ATPase α1, ß1 and ß3 are the main isoforms expressed in human blastocysts and no association was found between the expression level of their respective mRNAs and the rate of blastocyst expansion. WHAT IS KNOWN ALREADY: In mouse embryos, Na+/K+ ATPase α1 and ß1 are expressed in the basolateral membrane of trophectoderm (TE) cells and are believed to be involved in blastocoel formation (cavitation). STUDY DESIGN, SIZE, DURATION: A total of 20 surplus embryos from 11 patients who underwent IVF and embryo transfer at a university hospital between 2009 and 2018 were analyzed. PARTICIPANTS/MATERIALS, SETTING, METHODS: After freezing and thawing Day 5 human blastocysts, their developmental behavior was observed for 24 h using time-lapse imaging, and the expression of Na+/K+ ATPase isoforms was examined using quantitative RT-PCR (RT-qPCR). The expressed isoforms were then localized in blastocysts using fluorescent immunostaining. MAIN RESULTS AND THE ROLE OF CHANCE: RT-qPCR results demonstrated the expression of Na+/K+ ATPase α1, ß1 and ß3 isoforms in human blastocysts. Isoforms α1 and ß3 were localized to the basolateral membrane of TE cells, and ß1 was localized between TE cells. A high level of ß3 mRNA expression correlated with easier hatching (P = 0.0261). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The expression of mRNA and the localization of proteins of interest were verified, but we have not been able to perform functional analysis. WIDER IMPLICATIONS OF THE FINDINGS: Of the various Na+/K+ ATPase isoforms, expression levels of the α1, ß1 and ß3 mRNAs were clearly higher than other isoforms in human blastocysts. Since α1 and ß3 were localized to the basolateral membrane via fluorescent immunostaining, we believe that these subunits contribute to the dilation of the blastocoel. The ß1 isoform is localized between TE cells and may be involved in tight junction formation, as previously reported in mouse embryos. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the JSPS KAKENHI (https://www.jsps.go.jp/english/index.html), grant number 17K11215. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors have no conflicts of interest.

Thermal conductivity evidence for a dx2-y2 pairing symmetry in the heavy-fermion CeIrIn5 superconductor.

The phase diagram of the quasi-2D Ce(Ir,Rh)In5 system contains two distinct superconducting domes. By the thermal transport measurements in rotating magnetic fields H, we pinned down the superconducting gap structure of CeIrIn5 in the second dome, located distant from the first dome in proximity to an antiferromagnetic quantum critical point. Clear fourfold oscillation was observed when H is rotated within the ab plane, while no oscillation was observed within the bc plane. In sharp contrast to previous reports, our results are most consistent with dx2-y2 symmetry, implying that the superconductivity in the second phase is also mediated by antiferromagnetic spin fluctuations.

Exotic superconducting properties in the electron-hole-compensated heavy-fermion "Semimetal" URu2Si2.

We show that the charge and thermal transport measurements on ultraclean crystals of URu2Si2 reveal a number of unprecedented superconducting properties. The uniqueness is best highlighted by the peculiar field dependence of thermal conductivity including the first-order transition at Hc2 with a reduction of entropy flow. This is a consequence of multiband superconductivity with compensated electronic structure in the hidden order state of this system. We provide strong evidence for a new type of unconventional superconductivity with two distinct gaps having different nodal topology.

Pathophysiological role of endothelin in ectopic ossification of human spinal ligaments induced by mechanical stress.

Ossification of the posterior longitudinal ligament (OPLL) of the spine is characterized by progressive ectopic bone formation in the spinal ligament. To identify the genes related to ossification affected by mechanical stress during OPLL, analyses using cDNA microarray were carried out using cultured human spinal ligament cells that had been subjected to uniaxial cyclic stretching. Samples were obtained from a total of 14 patients: seven cervical or thoracic OPLL patients and seven control patients. Spinal ligament cells derived from tissues of OPLL (OPLL cells) and control (non-OPLL cells) patients were subjected to uniaxial sinusoidal cyclic stretching (0.5 Hz, 20% stretch) for various time periods (0-9 hours). cDNA microarrays revealed that ranges of distribution of both up- and downregulated genes evoked by cyclic stretching were significantly wider in OPLL cells than in non-OPLL cells. Increases in the mRNA expression of endothelin-1 (ET-1) as well as various marker genes related to ossification were also observed. mRNA expression of ET-1 and alkaline phosphatase was increased by mechanical stress in a time-dependent manner, while addition of ET-1 to static cultures of OPLL cells increased mRNA expression of alkaline phosphatase in a dose-dependent manner. During 9 hours of cyclic stretching, ET-1 release increased to about sixfold the amount observed in nonstretched cells. In non-OPLL cells, neither cyclic stretching nor ET-1 induced any increase in alkaline phosphatase expression. These results suggest that mechanical stress promotes the progression of ossification in OPLL cells through autocrine and/or paracrine mechanisms of ET-1.

Effects of in ovo exposure to imazalil and atrazine on sexual differentiation in chick gonads.

We examined the effects of atrazine and imazalil, 2 commonly used pesticides, on sexual differentiation in chickens. Atrazine and imazalil were injected into fertile eggs on d 0. At hatching, sex genotype and phenotype were determined. Gonads were stereomicroscopically and histologically observed. In ovo exposure of atrazine (0.01 to 3 mg/egg) did not influence hatchability, whereas imazalil exposure (2 mg/egg) inhibited hatchability. The sex genotype matched the sex phenotype in controls, atrazine, and imazalil-exposed groups. In control females, the right gonad was regressed at hatching. Regression of the right gonad, however, was inhibited following atrazine and imazalil exposure. In atrazine-exposed female chicks, the left gonads had normal ovary structures, and the remaining right gonads had ovary medulla-like structures. In imazalil-exposed females, some left gonads had an ovary medulla-like structure without the cortex as well as tubules, and the right gonad had testis-like structures. There was no change in male gonads at hatching following atrazine and imazalil exposure. Aromatase activity of the left gonad from female chicks was not changed by any concentration of atrazine exposure. These results suggest that atrazine and imazalil inhibit regression of the right gonad in female chicks, although it is not clear whether the remaining right gonad has aromatase activity. In ovo exposure to atrazine influences sexual differentiation of the ovary by different mechanisms from imazalil, possibly by the induction of aromatase in the right gonad, whereas it is confirmed that imazalil inhibits in vitro aromatase activity in the chick ovary. The results indicated that in ovo exposure to imazalil inhibits sexual differentiation of the ovary by inhibiting aromatase activity.

Magnetic resonance analysis of abnormal diaphragmatic motion in patients with emphysema.

The purpose of this study was to quantitatively evaluate paradoxical diaphragmatic motion using magnetic resonance (MR) imaging. A total of 27 subjects were examined, including 12 normal young adults, six control individuals, and nine patients with emphysema. With subjects in the supine position, 30 sequential sagittal MR images of the entire right lung were obtained during tidal and deep slow breathing. Diaphragmatic movement between sequential images was estimated as the displacement area and the total diaphragmatic movement in a respiratory cycle was calculated. The paradoxical motion of the diaphragm, representing the inverted movement to increase or decrease lung area, since paradoxical movement ratio (Mpr=(total paradoxical diaphragmatic movement/total diaphragmatic movement)x100), was evaluated. In patients with emphysema, paradoxical diaphragmatic motion was observed on MR images during deep breathing. The mean Mpr in emphysematous patients during deep breathing was 10+/-4%, which was significantly higher than 0.5+/-0.2% in young adults (p<0.05), and 1.2+/-0.6% in aged-matched controls (p<0.05). The present results indicate that magnetic resonance images could be used to detect paradoxical diaphragmatic motion in patients with emphysema.

Biomarker distribution after injection into the canine prostate: implications for gene therapy.

OBJECTIVE: To evaluate the distribution of biomarkers after transrectal injection into the canine prostate and to report a method for enhancing the distribution of gene expression. MATERIALS AND METHODS: Carbon black was first used to evaluate the histopathological distribution in canine prostate of single or multiple injections via the transurethral, transperineal and transrectal routes. The distribution of canarypox virus (ALVAC) vector-delivered gene expression was then compared using both fluid-phase injection techniques and delivery in a solid carrier composed of a gelatine sponge matrix. RESULTS: After transurethral administration, carbon black was detected as scattered particles in ducts and acini, mostly in the periphery of the gland. Direct transrectal injection of carbon black resulted in a localized collection at the site of injection, with only a minimal peri-acinar distribution. Transrectal injection of the fluid-phase (virus suspended in diluent) ALVAC vector encoding the beta-galactosidase gene resulted in a similar distribution, with limited gene expression at the site of injection and in the needle track. Delivery of the same number of virus particles in the gelatine sponge matrix resulted in qualitatively greater gene expression. CONCLUSIONS: Direct injection of the canine prostate with biomarkers, including viral vectors, in the fluid-phase results in very localized gene expression, while the distribution was more widespread after delivery in a gelatine sponge matrix.

Screening for abdominal aortic aneurysms during a basic medical checkup in residents of a Japanese rural community.

To determine the prevalence of abdominal aortic aneurysms (AAA) and the size distribution of the abdominal aortic diameter in the Japanese population, 1,591 residents of a farming community were screened by ultrasonography during the annual basic medical checkup arranged by the town. The mean diameter of the abdominal aorta was 17.2 +/- 2.2mm and four AAAs were found (0.3%), all in men aged 60 years or older, three of whom were hypertensive. The mean diameter of the common iliac artery was 10.2 +/- 5.1mm and five iliac arterial aneurysms were found (0.3%). One man had both aortic and iliac arterial aneurysms. The iliac arterial aneurysms were also found in men, four of whom were aged 60 years or older and had a smoking habit. We were able to keep the cost of screening low by combining the program with an annual basic medical checkup. These findings indicate that it is important to examine the iliac artery as well as the abdominal aorta because the incidence of aneurysm is similar in both. Screening for AAAs using ultrasonography is accurate enough to detect them before rupture, and can be effectively performed in men aged 60 years or older during their annual basic medical checkup.

Paradoxical motion of the hemidiaphragm in patients with emphysema.

The authors evaluate paradoxical diaphragmatic motion using magnetic resonance (MR) imaging in patients with emphysema. The subjects were 12 healthy volunteers and 10 male patients with moderate to severe air flow obstruction. With subjects in the supine position, 30 sequential sagittal images of the bilateral lungs were obtained during quiet and forced breathing using a 1.5T MR unit with a body coil. The sequence was single shot fast spin echo (SSFSE) with half Fourier transformation. Subtraction images were made from the original images (by subtracting a given image from the preceding image), which visualized the chest wall motion as white or black bands on the edge of the lung fields. The authors evaluated both the original and subtraction images. MR imaging showed abnormal hemidiaphragmatic motion during forced breathing: the ventral portion of the hemidiaphragm moved downward while the dorsal part moved upward like a seesaw in 6 patients. MR images also revealed abnormal ribcage motion; the ventral ribcage moved anteriorly when the hemidiaphragm moved upward in 7 patients. No abnormal motion was observed in healthy volunteers. MR is a noninvasive and useful tool for evaluating the asynchronous respiratory motion in patients with emphysema.

MR in vivo imaging of oxygen suppression effect of soft contact lens on the human cornea.

The O(2) suppression effect of a soft contact lens on the human cornea was measured using dynamic magnetic resonance imaging (MRI) of the anterior chamber transcorneally exposed to O(2). Dynamic T(1)-weighted fast spin echo imaging of anterior chambers (TR = 2 s, TE = 15 ms, 5-mm slice) was performed both before and during oxygen supply to a full goggle placed on the face of volunteers wearing a soft contact lens on one eye and nothing on the other eye as a control. Within 15 min after O(2) administration, significantly lower intensity changes were obtained in the anterior chambers of the eyes with the contact lens than in those of the eyes without one, suggesting that dynamic MRI of the anterior chamber transcorneally exposed to O(2) can be used to evaluate the O(2) suppression effect of a soft contact lens on the cornea.

MR findings of bowlegs in toddlers.

BACKGROUND: Toddlers with severe physiologic tibial bowing are considered to be at risk for the development of Blount's disease. Objective. To correlate MR findings of the knee with the clinical outcome in toddlers with severe physiological tibial bowing. MATERIALS AND METHODS: MR findings were evaluated in 22 affected legs of 14 children with severe tibial bowing (mean age 1.9 years). In 18 affected legs, MR findings were compared with the transition of the tibial metaphyseal-diaphyseal angle (MDA) and tibiofemoral angle (TFA) measured serially between 2 and 3 years of age. RESULTS: MR findings of severe tibial bowing comprised undulation of the posteromedial physis of the tibia (3/22), signal alterations in the medial tibial metaphysis (10/22), T2 prolongation in the posteromedial tibial epiphyseal cartilage (14/22) and signal changes in the medial menisci (18/22). The decrease in the TFA was different in the legs with and without increased signal in the epiphyseal cartilage, and the decrease in the MDA was different in the legs with and without physeal undulation. CONCLUSION: MR imaging findings can predict the retarded resolution of tibial bowing, which may be a risk factor for the development of Blount's disease.

Normal in-plane respiratory motion of the bilateral hemidiaphragms evaluated by sequentially subtracted fast magnetic resonance images.

The purpose of this study was to demonstrate hemidiaphragmatic motion using magnetic resonance imaging. The subjects were 13 healthy, nonsmoking volunteers. The sequence consisted of a fast gradient-recalled echo for coronal images and a single-shot fast spin echo with half Fourier transformation for sagittal images. The coronal and sagittal acquisition times were 0.83 msec and 0.58 msec per image, respectively. Thirty sequential images were obtained during quiet, deep respiration. Two series of subtraction images were made from these original sequential images (a given image was subtracted from the preceding or the next image in the original series). Hemidiaphragmatic motion was visualized as white or black bands on subtraction images. In the coronal plane through the left ventricle, there was a difference in motion between the right and left hemidiaphragms in 10 subjects during tidal respiration and in 12 subjects during deep respiration. On the sagittal images, the dorsal part of the hemidiaphragm moved more rapidly during quiet, deep respiration, especially on the left side.

Variability of platyspondylic lethal chondrodysplasia: another case report.

We report the radiological and histological findings of another case of platyspondylic lethal chondrodysplasia. The patient was a girl, who died of respiratory failure at 18 days of age. The radiological changes comprised moderate platyspondyly with ovoid-shaped vertebral bodies, broad and short ilia, rhizomelic shortening and mild bowing of the long bones (particularly of the humeri), relatively long short tubular bones, and retarded epiphyseal ossification and ragged metaphyses, which were most similar to those of a mild variant of this entity, the Luton type. However, the histological findings of cartilage, including hypercellularity of the reserve zone with round resting chondrocytes, relatively normal column formation of the proliferative and hypertrophic zones, and incorporation of hypertrophic cartilage with a columnar arrangement into metaphyseal bony trabeculae, resemble those of a severe variant of this entity, the Torrance type. Our observation provides an insight into the phenotypic variabilities of platyspondylic lethal chondrodysplasia.

Dynamic MRI of transcorneal dispersion of oxygen into the anterior chamber of human eye.

To measure the transcorneal dispersion of oxygen into the anterior chamber, dynamic T1-weighted fast-spin-echo MRI (TR=2 seconds, TE=15 msec, 5-mm slice) of the human eye was performed both before and during oxygen supply to a full goggle placed on the face. During the course of the imaging, a significant increase in the signals in the anterior chamber occurred. This indicated that transcorneal dispersion of oxygen into the anterior chamber can be evaluated by this procedure, suggesting that this method may be useful for diagnosing dysfunction of the cornea or aqueous flow.

[Changes in acetylcholine level and its related enzyme activities in rat brain following focal ischemia].

At 1 to 4 weeks following middle cerebral artery occlusion in Wistar rats, levels of acetylcholine, which is neuro-transmitter, were measured. In the hippocampus, which plays an important role in memory, levels of acetylcholine in the ischemic group could not be detected the significant difference compared with those in the sham-operated group. But in the anterior cortex and the caudate-putamen, levels of acetylcholine in the ischemic group were recognized the significant difference compared with those in the sham operated group. Moreover activities of choline acetyltransferase, which is the synthetic enzyme of acetylcholine, and activities of acetylcholinesterase, which is the degradative enzyme of acetylcholine, were measured in the anterior cortex which was recognized decrease of the acetylcholine level and the hippocampus which could not be detected the difference of the acetylcholine level. Activities of both enzymes in the hippocampus could not be recognized the significant difference between the ischemic group and the normal group. But in the anterior cortex activities of both enzymes in the ischemic group were significantly decreased compared with those in the normal group. These results suggest that these decreases in the ischemic group were due to damage of injection of cholinergic neuron from Meynert nucleus (basal nucleus of Meynert) to anterior cortex.

Diffusion-weighted imaging of the human optic nerve: a new approach to evaluate optic neuritis in multiple sclerosis.

The apparent diffusion coefficient (ADC) in the optic nerve was measured from diffusion-weighted magnetic resonance imaging using an intravoxel incoherent motion (IVIM) sequence. The subjects were seven normal volunteers and eight patients with multiple sclerosis (MS) with a total of four optic nerves with acute neuritis and nine nerves with chronic neuritis. The mean ADC (4.18 +/- 1.13 x 10(-3) mm2/s, n = 9) in the optic nerves with chronic neuritis was significantly higher than that in normal volunteers (1.56 +/- 0.675 x 10(-3) mm2/s, n = 14) and that in the nerves with acute neuritis (0.94 +/- 0.43 x 10(-3) mm2/s n = 4) (P < 0.001). The ADC is useful in assessing MS foci in the optic nerves.

In vitro cellular aging stimulates interleukin-1 beta production in stretched human periodontal-ligament-derived cells.

Although the severity of periodontal disease is known to be affected by host age, the pathological role of aging in periodontal disease, and especially that attributable to trauma from occlusion, has not been well-characterized. Interleukin (IL)-1 beta is a key mediator involved in periodontal diseases, a potent stimulator of bone resorption. Furthermore, it is produced by human periodontal ligament (PDL) cells in response to mechanical stress. To investigate the age-related changes in the biosynthetic capacity of IL-1 beta in PDL cells, we examined the effects of in vitro cellular aging with mechanical stress on IL-1 beta protein and gene expression by human PDL cells. Human PDL cells (young = 5th or 6th passage; old = 18-20th passage) were cultured on flexible-bottomed culture plates, and the cells were deformed at 6 cycles per min at 2 steps of tension force for 1 to 5 days. We found a two-fold increase in IL-1 beta production by old PDL cells subjected to mechanical tension compared with that by young PDL cells, although the constitutive levels of IL-1 beta were similar in both the young and old PDL cells. This increase was tension-dependent. IL- 1 beta mRNA was also detected in both cell types under basal conditions, and its expression was further enhanced by application of mechanical tension by use of reverse-transcription-polymerase chain-reaction (RT-PCR) and in situ hybridization methods. The increase in signal rate was higher in the old cells than in the young cells. IL-1 beta-converting enzyme mRNA remained unchanged. It is possible that a large amount of IL- 1 beta produced by PDL cells from an aged host in response to mechanical force may be positively related to the acceleration of alveolar bone resorption.

Cephalometric analysis of masseter muscle and dentoskeletal morphology in dentate and edentulous humans.

A study was done to clarify the attachment position of the superficial masseter muscle and its relationship with craniofacial morphology in dentate and edentulous subjects. Data were obtained from lateral cephalometric radiographs of a total of 39 cadavers in which the superficial masseter muscle had been defined with colloidal barium. Principal component analysis provided evidence that the lower masseter muscle width was associated with the size of the ramus, and related to the position of the anterior margin of the muscle. Linear discriminant function data suggested that ramus width, coronoid height and the distance between the anterior margin of the masseter muscle and the mandibular notch contributed most to the observed difference between dentate and edentulous subjects. The derived function correctly assigned 100% of dentate and 95% of edentulous subjects. The results of this study indicated that a reduction of masticatory function was associated with the position of the anterior border of the masseter muscle insertion and also with differences in ramus dimension, the most significant of which were differences in the coronoid process and gonial angle. In general, age was not a significant determinant of variation in superficial masseter muscle dimensions or orientation.

Effect of tension-force on plasminogen activator activity from human periodontal ligament cells.

The plasminogen activator (PA)-plasmin proteolytic system has recently received considerable attention because of its participation in a wide variety of biological activities and in pathological conditions involving tissue destruction. Excessive mechanical stress such as occlusal trauma is associated with alveolar bone loss in severe periodontitis. Therefore, mechanical stress may involve degradation of the extracellular matrix by occlusal trauma through activation of the PA-plasmin proteolytic system. We examined the effects of mechanical stress on PA activity, gene expressions of tissue type (t) PA, urokinase type (u) PA and PA inhibitor-1 (PAI-1) in human PDL cells. Human PDL cells were cultured on flexible-bottomed culture plates and placed on a Flexercell Strain Unit. The cells were flexed at 6 cycles (5 s strain, 5 s relaxation) at 9% and 18% elongation for 5 d. Application of tension-force induced significantly higher PA activity in stressed PDL cells than in non-stressed controls, and did so in a time- and magnitude-dependent manner (p < 0.001, ANOVA). Western-blot analysis revealed that the high level of activity was due to tPA and not uPA. Gene expression of tPA mRNA in stressed PDL cells, as examined by RT-PCR, increased on d 5. These findings suggest that tPA may be involved in periodontal metabolism in response to mechanical stress.