Identification of histidine residues that affect the T/R-state conformations of human hemoglobin using constant pH molecular dynamics simulations.

Human hemoglobin (Hb) is a tetrameric protein consisting of two α and two ß subunits that can adopt a low-affinity T- and high-affinity R-state conformations. Under physiological pH conditions, histidine (His) residues are the main sites for proton binding or release, and their protonation states can affect the T/R-state conformation of Hb. However, it remains unclear which His residues can effectively affect the Hb conformation. Herein, the impact of the 38 His residues of Hb on its T/R-state conformations was evaluated using constant-pH molecular dynamics (CpHMD) simulations at physiological pH while focusing on the His protonation states. Overall, the protonation states of some His residues were found to be correlated with the Hb conformation state. These residues were mainly located in the proximity of the heme (α87 and ß92), and at the α1ß2 and α2ß1 interfaces (α89 and ß97). This correlation may be partly explained by how easily hydrogen bonds can be formed, which depends on the protonation states of the His residues. Taken together, these CpHMD-based findings provide new insights into the identification of titratable His residues α87, α89, ß92, and ß97 that can affect Hb conformational switching under physiological pH conditions.

The anatomy and feeding mechanism of the Japanese giant salamander (Andrias japonicus).

The fully aquatic Japanese giant salamander (Andrias japonicus) is a member of the Cryptobranchidae, and is currently distributed in western Japan, with other members of this group restricted to China and North America. Their feeding behaviour is characterized by a form of suction feeding that includes asymmetric movements of the jaw and hyobranchial apparatus. Previous studies on the North American species, Cryptobranchus alleganiensis, have suggested that this specialized jaw movement is produced by a flexible quadrate-articular joint combined with a loosely connected lower jaw symphysis including two small fibrocartilaginous pads. However, little is known about this feeding behaviour in the Asian species, nor have the three-dimensional asymmetric jaw movements been fully investigated in any member of Cryptobranchidae. In this study, we explore the asymmetric jaw movements in A. japonicus using three methods: (1) dissection of musculoskeletal structures; (2) filming of feeding behaviour to understand in which situations asymmetric feeding is used; (3) analysis of 3D movement of jaws and skull. In the third component, fresh (from frozen) specimens of A. japonicus were manipulated to replicate asymmetric and symmetric jaw movements, with the specimens CT scanned after each step to obtain the 3D morphology of the jaws at different positions. These positions were combined and their Euler angles from resting (closed) jaw position were calculated for asymmetric or symmetric jaw positions. Our filming revealed that asymmetric jaw movements are linked to the position of the prey in relation to the snout, with the jaw closest to the prey opening asymmetrically. Moreover, this action allows the salamander to simultaneously grasp prey in one side of the mouth while ejecting water on the other side, if the first suction attempt fails. The asymmetric jaw movements are performed mainly by rotation of the mandible about its long axis, with very limited lateral jaw movements. During asymmetric and symmetric jaw movements, the posterior ends of the maxilla and quadrate move slightly. The asymmetric jaw movements are permitted by a mobile quadrate-articular joint formed by wide, round cartilages, and by two small fibrocartilage pads within the jaw symphysis that act as cushions during jaw rotation. Some of these soft tissue structures leave traces on the jaws and skull, allowing feeding mode to be reconstructed in fossil taxa. Understanding cryptobranchid asymmetric jaw movement thus requires a comprehensive assessment of not only the symphysial morphology but also that of other cranial and hyobranchial elements.

Boron Clusters Alter the Membrane Permeability of Dicationic Fluorescent DNA-Staining Dyes.

Membrane-permeable fluorescent dyes that stain DNA are useful reagents for microscopic imaging, as they can be introduced into living cells to label DNA. However, the number of these dyes, such as Hoechst 33342, is limited. Here, we show that the icosahedral dodecaborate B12Br122-, a superchaotropic carrier that delivers different types of molecules into cells, functions as an excellent carrier for membrane-impermeable fluorescent dyes. Propidium iodide (PI) and 4',6-diamidino-2-phenylindole (DAPI), dicationic membrane-impermeable fluorescent dyes that stain DNA, can permeate cell membranes in the presence of boron clusters. Methyl green (MG), a dicationic dye used in the histological and fluorescent staining of DNA, permeated cell membranes in the presence of boron clusters. In contrast, monocationic membrane-permeable fluorescent dyes, such as acridine orange and pyronin Y, exhibited reduced fluorescence in cells in the presence of boron clusters. Boron clusters do not quench dicationic fluorescent dyes in water in vitro but have quenching effects on monocationic fluorescent dyes. We have demonstrated that the addition of B12Br122- to impermeable dicationic fluorescent DNA-staining dyes, such as DAPI, PI, and MG, which have been widely used for numerous years, imparts membrane permeability to introduce these dyes into living cells.

Fossorial mammals emphasise the forelimb muscle moment arms used for digging: New indices for reconstruction of the digging ability and behaviours in extinct taxa.

Among fossorial mammals, forelimbs are major digging apparatuses for dwelling, sheltering and foraging underground. Forelimb-diggers have independently evolved in many lineages of mammals; thus, the method of digging with forelimbs varies by taxon. Therefore, the reconstruction of digging behaviours in extinct animals leads us to understand the evolutionary process of fossorial adaptation in each lineage. However, no morphological index was found to reconstruct if, or how, extinct taxa dug with forelimbs. In this study, we used the shoulder and elbow muscle moment arms in relation to the out-force lever on the manus as indices of the efficiency of motions. The mechanical advantage of two shoulder motions (medial rotation and retraction) and three elbow motions (extension, flexion and adduction) was measured in 381 extant mammal specimens representing 332 species, 279 genera, 103 families and 24 orders. Assuming that both forelimb-digging and -paddling in water require relatively high-output moment arm efficiency, the studied taxa were categorised into four groups based on the presence or absence of forelimb-digging and -paddling abilities. We found that the efficiencies of all five muscle moment arms in the forelimb-diggers and -paddlers were higher than those of the non-diggers and non-paddlers. Furthermore, among the forelimb-diggers, the taxa that dig compact substrates or frequently burrow tend to emphasise the muscle moment arms compared to the taxa that dig loose substrates or dig less frequently. The comparison among the 53 extant forelimb-diggers revealed that the efficiency marked among the five muscle moment arms reflects the difference in digging strategy: humeral rotation diggers emphasise the shoulder medial rotator and elbow adductor, hook-and-pull diggers emphasise the shoulder retractor and elbow flexor and scratch diggers emphasise the shoulder retractor and elbow extensor. We propose that these indices will be powerful tools for reconstructing the fossorial behaviours of extinct mammals. Applying these indices to extinct taxa, Ceratogaulus, Ernanodon, Metacheiromys and Prozaedyus are capable of more efficient forelimb-digging, and each may have adopted different digging strategies.

Coracoid strength as an indicator of wing-beat propulsion in birds.

Birds generate a propulsive force by flapping their wings. They use this propulsive force for various locomotion styles, such as aerodynamic flight, wing-paddle swimming and wing-assisted incline running. It is therefore important to reveal the origin of flapping ability in the evolution from theropod dinosaurs to birds. However, there are no quantitative indices to reconstruct the flapping abilities of extinct forms based on their skeletal morphology. This study compares the section modulus of the coracoid relative to body mass among various extant birds to test whether the index is correlated with flapping ability. According to a survey of 220 historical bird specimens representing 209 species, 180 genera, 83 families and 30 orders, the section modulus of the coracoid relative to body mass in non-flapping birds was significantly smaller than that of flapping birds. This indicates that coracoid strength in non-flapping birds is deemphasised, whereas in flapping birds the strength is emphasised to withstand the contractile force produced by powerful flapping muscles, such as the m. pectoralis and m. supracoracoideus. Therefore, the section modulus of the coracoid is expected to be a powerful tool to reveal the origin of powered flight in birds.

Serum pepsinogen cut-off values in Helicobacter pylori-infected children.

BACKGROUND: The aim of this study was to evaluate the standard values for gender- and age-stratified serum pepsinogen (sPG) in Helicobacter pylori (H. pylori) non-infected children and to determine the optimal cut-off values of sPG for predicting H. pylori-infected gastritis in children. METHODS: A prospective study for determination of sPG levels was performed in children with epigastric pain who underwent esophagogastroduodenoscopy over the past 16 years. After excluding subjects diagnosed with inflammatory bowel diseases, eosinophilic gastrointestinal disorders, or immunoglobulin A vasculitis, the diagnosis of H. pylori infection was defined by positive tissue culture or concordant-positive results for histology and the rapid urease test. RESULTS: A total of 405 subjects were diagnosed as being H. pylori-infected (79) or non-infected (326). In the H. pylori non-infected group, there were no significant differences in sPG levels among age groups; males had higher sPG I and sPG II levels than females. In the H. pylori-infected group, sPG I and sPG II levels were significantly higher and the sPG I/II ratio was lower than those in the non-infected group. In receiver operating characteristics analyses in diagnosing H. pylori infection, the areas under the curves for sPG I, sPG II and sPG I/II ratio were 0.896, 0.980, and 0.946, respectively. The optimal cut-off value of sPG II of ≥9.0 ng/mL was considered positive for H. pylori infection (sensitivity: 92.4%, specificity: 93.9%). CONCLUSIONS: The optimal cut-off value of sPG II of ≥9.0 ng/mL may be a good predictor of H. pylori-infected gastritis in children.

Feeding behaviour and functional morphology of the neck in the long-snouted aquatic fossil reptile Champsosaurus (Reptilia: Diapsida) in comparison with the modern crocodilian Gavialis gangeticus.

The extinct freshwater choristoderan reptiles Champsosaurus and Simoedosaurus are characterised by large body size and an elongated snout. They have often been considered as eco-analogues of crocodilians based on superficial similarities. The slender-snouted Champsosaurus has been described as a 'gavial-like reptile', which implies it feeds underwater with a lateral swipe of the head and neck, as in the living slender-snouted crocodilians such as Gavialis gangeticus. In contrast, the short-snouted Simoedosaurus is often compared with short-snouted living crocodilians and is considered to take single prey items. However, the neck mobility and flexibility needed for feeding movements are poorly understood even in extant crocodilians. This study explores the relationship between cervical morphology and neck flexion, focusing particularly on lateral and dorsal movements in G. gangeticus by comparison with shorter-snouted crocodilians. The paper also describes a method to estimate the maximum angle of neck dorsiflexion in choristoderes based on the cervical morphology of extant crocodilian species. Three indices were used in this study, of which Index 3 is newly proposed, to compare cervical morphology and intervertebral joint flexibility: (1) Enclosed zygapophyseal angles (EZA) as an index of dorsoventral/ bilateral flexibility, (2) moment arm (M) of dorsiflexor muscles as an Index of resistance against ventroflexion and (3) the orientations of zygapophysial facets for a maximum angle of dorsiflexion. These Indices were validated using µCT scanning of fresh specimens of G. gangeticus and Caiman latirostris in lateral and dorsal flexion. A unique mechanism of lateral flexion was identified in G. gangeticus that uses a combination of the following features: (1) lateral flexion mainly restricted to the anterior cervical vertebrae (v2/v3: high EZA, with more horizontal zygapophyses) and (2) high degree of dorsiflexion at the v3/v4 and v4/v5 joints with potential for dorsal flexibility through the middle-posterior neck, which is used in inertial feeding. In contrast, Champsosaurus and Simoedosaurus possess relatively short cervical vertebrae, as in short-snouted crocodilians. The middle-posterior cervical vertebrae of Champsosaurus are specialised for lateral flexion (high EZA), and there is only limited capacity for dorsiflexion throughout the neck. Like G. gangeticus, therefore, Champsosaurus may have used its slender snout to grab fish from shoals using lateral sweeping motions of the head and neck, but the movement is through the neck, not the craniocervical joint. However, inertial feeding is less likely to have occurred in this genus, and the aligned palatal dentition may have aided the lingual transport of prey into the mouth. Simoedosaurus, on the other hand, appears to have been less specialised, with a neck that combined lateral and dorsolateral flexion, a move that could have been effective in catching both terrestrial and aquatic prey. Where these two choristoderan genera occurred in the same place, they may have divided their niche by prey types.

Absent X-linked inhibitor of apoptosis protein expression in T cell blasts and causal mutations including non-coding deletion.

BACKGROUND: X-linked inhibitor of apoptosis protein (XIAP) deficiency is one of inborn errors of immunity characterized by recurrent hemophagocytic lymphohistiocytosis and refractory inflammatory bowel disease (IBD), mimicking Crohn's disease. The aim of this study is to make an accurate diagnosis of XIAP deficiency based on genetic and XIAP expression studies and to investigate endoscopic findings shared by patients with this disease. METHODS: Four male patients with recurrent hemophagocytic lymphohistiocytosis and long-term refractory IBD were studied for the diagnosis of XIAP deficiency. Endoscopic findings of the four patients were also studied in parallel. RESULTS: These four patients were diagnosed with XIAP deficiency based on the absent XIAP expression in cultured T-cell blasts. Sequence analysis of the responsible gene, XIAP, demonstrated two novel nonsense mutations of p.Gln114X and p.Glu25X, and a previously reported nonsense mutation of p.Arg381X. Although no mutations in the coding region were detected in the fourth patient, further studies demonstrated a novel 2,199 bp deletion encompassing non-coding exon 1, presumably affecting transcription and stability of XIAP mRNA. All of the patients eventually underwent hematopoietic stem cell transplantation, leading to a complete or partial remission of IBD. These four patients shared an endoscopic finding of multiple wide and longitudinal ulcers with straight and non-raised edge in the colon. CONCLUSIONS: X-linked inhibitor of apoptosis protein expression in T-cell blasts could facilitate the diagnosis of this disease, especially with causal mutations in non-coding regions.

Bio-inspired asymmetric aldehyde arylations catalyzed by rhodium-cyclodextrin self-inclusion complexes.

Transition-metal catalysts are powerful tools for carbon-carbon bond-forming reactions that are difficult to achieve using native enzymes. Enzymes that exhibit inherent selectivities and reactivities through host-guest interactions have inspired widespread interest in incorporating enzymatic behavior into transition-metal catalytic systems that highly efficiently produce enantiopure compounds. Nevertheless, bio-inspired transition-metal catalysts that are highly enantioselective and reactive have rarely been reported. In this study, we applied γ-cyclodextrin-imidazolium salts to the rhodium-catalyzed asymmetric arylations of aldehydes. The method exhibits wide substrate scope and the corresponding arylcarbinols are obtained in excellent yields under optimized conditions, with enantiomeric excesses of up to 96% observed. Kinetic and competition experiments revealed that self-inclusion of the Rh complex contributes to the high enantioselectivity and reactivity achieved by this catalytic system. Thus, this bio-inspired self-inclusion strategy is promising for the development of highly enantioselective and reactive transition-metal catalysts for asymmetric carbon-carbon bond formation.

Cyclodextrin-Based [c2]Daisy Chain Rotaxane Insulating Two Diarylacetylene Cores.

A [c2]daisy chain rotaxane with two diarylacetylene cores was efficiently synthesized in 53 % yield by capping a C2 -symmetric pseudo[2]rotaxane composed of two diarylacetylene-substituted permethylated α-cyclodextrins (PM α-CDs) with aniline stoppers. The maximum absorption wavelength of the [c2]daisy chain rotaxane remained almost unchanged in various solvents, unlike that of the stoppered monomer, indicating that the two independent diarylacetylene cores were insulated from the external environment by the PM α-CDs. Furthermore, the [c2]daisy chain rotaxane exhibited fluorescence emission derived from both diarylacetylene monomers and the excimer, which implies that the [c2]daisy chain structure can undergo contraction and extension. This is the first demonstration of a system in which excimer formation between two π-conjugated molecules within an isolated space can be controlled by the unique motion of a [c2]daisy chain rotaxane.

Effect of Orento, a Traditional Japanese Medicine, on IL-6, IL-8 Secretion, Type 1 Collagen Production and Alkaline Phosphatase Secretion in the Human Osteosarcoma Cell Line Saos-2.

Background: Orento, a traditional Japanese medicine, is known as Kampo medicine in Japan. We investigated the possible efficacy of Kampo medicine for periodontal disease. In this study, we examined the in vitro effects of orento on the proliferation of the inflammatory cytokines interleukin (IL)-6 and IL-8, the production of type 1 collagen, and the secretion of alkaline phosphatase (ALP) in the human osteosarcoma cell line Saos-2 (Saos-2 cells). Methods: The proliferation of Saos-2 cells was assessed by MTT assay. IL-6 and IL-8 levels, type 1 collagen production and ALP secretion were evaluated using enzyme-linked immunosorbent assay and ALP assays. Saos-2 cells were treated with or without 0.1, 1, 10, 100 and 1000 µg/mL of orento for 24 h. Results: Orento (10 µg/mL) significantly induced the proliferation of Saos-2 cells. At this concentration, orento suppressed IL-6 and IL-8 and enhanced type 1 collagen production and ALP secretion. Conclusions: These results indicate that orento controls the IL-6 and IL-8 secretion and cellular metabolism of osteoblasts, resulting in the secretion of early bone-related biomarkers.

Comprehensive Targeted Sequencing Identifies Monogenic Disorders in Patients With Early-onset Refractory Diarrhea.

OBJECTIVES: Causes of early-onset refractory diarrhea include exudative diarrhea associated with very early-onset inflammatory bowel diseases, osmotic or secretory diarrhea, and protein-losing enteropathy. Monogenic disorders are included in these diseases, yet a comprehensive genetic analysis has not been fully established. METHODS: We established targeted gene panels covering all responsible genes for early-onset diarrhea. In total, 108 patients from 15 institutions were enrolled in this study. We collected clinical data from all patients. Seventy-three patients with exudative diarrhea, 4 with osmotic or secretory diarrhea and 8 with protein-losing enteropathy were subjected to genetic analysis. RESULTS: A total of 15 out of the 108 enrolled patients (13.9%) were identified as monogenic. We identified 1 patient with RELA, 2 with TNFAIP3, 1 with CTLA4, 1 with SLCO2A1, 4 with XIAP, 3 with IL10RA, 1 with HPS1, 1 with FOXP3, and 1 with CYBB gene mutations. We also identified 1 patient with NFKB2 and 1 with TERT mutations from the gene panel for primary immunodeficiency syndromes. The patient with refractory diarrhea caused by heterozygous truncated RelA protein expression is the first case identified worldwide, and functional analysis revealed that the mutation affected nuclear factor kappa B signaling. Genotypes were significantly associated with the clinical and pathological findings in each patient. CONCLUSIONS: We identified variable monogenic diseases in the patients and found that genes responsible for primary immunodeficiency diseases were frequently involved in molecular pathogenesis. Comprehensive genetic analysis was useful for accurate molecular diagnosis, understanding of underlying pathogenesis, and selecting the optimal treatment for patients with early-onset refractory diarrhea.An infographic for this article is available at: http://links.lww.com/MPG/B853.

The range of atlanto-occipital joint motion in cetaceans reflects their feeding behavior.

The cetaceans display a wide variety of lifestyles, especially with regard to their feeding behavior. However, the evolutionary process of the feeding behavior in cetaceans is still poorly understood, in part because reconstructing the feeding behavior of extinct taxa remains difficult. In cetaceans, cranium mobility relative to the trunk largely depends on the range of motion permitted by the atlanto-occipital joint, given the lack of flexibility of the cervical series. In this study, we examined 56 extant cetacean skeletal specimens from 30 species in 25 genera and nine families in order to investigate the relationships between anatomical traits and feeding behavior. Our results suggest that the range of dorso-ventral motion allowed by the atlanto-occipital joint (ROM) depends on prey habitat and the feeding technique of cetaceans. Cetaceans feeding on benthic/demersal prey had a relatively large ROM compared with those feeding on pelagic prey. In addition, ROM was largest in raptorial feeders, intermediate in suction feeders, and smallest in ram-filter feeders. Among raptorial feeders, ROM tended to be larger in taxa that facultatively tear off the prey's flesh compared with taxa that swallow their prey whole. Therefore, we conclude that ROM is a powerful tool to reliably reconstruct the feeding behavior of extinct cetacean taxa.

Physical constraints on sand crab burrows: Mechanical properties of wet sand explain the size and spatial distributions of burrows on beaches.

The diameter and vertical depth of sand crab tunnels in sandy beaches are usually restricted to a few centimeters scale and several tens of centimeters, respectively. We designed a study to determine what physical factors restrict tunnel diameter and predict the maximum attainable tunnel diameter and depth. We collected field data on the size and spatial distributions of ghost crab (Ocypode spp.) burrows on two sandy beaches (Kawage Beach in Tsu, Mie Prefecture, Japan and Sakieda Beach in Ishigaki, Okinawa Prefecture, Japan), where O. ceratophthalma dominants the ghost crab fauna. We measured burrow depths and distance from shoreline in concert with water content of sandy beaches. To explain our observed distributions of crab burrows in the field, we performed experiments in a lab microcosm, comprising a horizontal tunnel through wet sand. We measured the static stability of tunnel structures in relation to water content and two strengths computed from loading force exerted on the sand overlying the tunnels. By comparing field and experimental data, we found that crabs construct their burrows in appropriately wet zones (wet enough to provide sufficient cohesion of the sand grains in tunnel walls to prevent collapse) and that tunnel diameters and depths are sufficiently small to prevent deformation and collapse of their tunnels.

The sandwich structure of keratinous layers controls the form and growth orientation of chicken rhinotheca.

The upper beak bone of birds is known to be overlain by the rhinotheca, which is composed of the horny sheath of keratinous layers. However, the details of the structure and growth pattern of the rhinotheca are yet to be understood. In this study, the microstructure of the rhinotheca from chicken specimens of different growth stages (ranging from 1 to ~ 80 days old) was analyzed using a combination of thin section and scanning electron microscopy observations, and small-angle X-ray scattering analysis. We found that the rhinotheca comprises three different layers - outer, intermediate, and inner layers - throughout its growth. The outer layer arises from the proximal portion of the beak bone and covers the dorsal surface of the rhinotheca, whereas the intermediate and inner layers originate in the distal portion of the beak bone and underlie the outer layer. This tri-layered structure of the rhinotheca was also observed in wild bird specimens (grey wagtail, king quail, and brown dipper). On the median plane, micro-layers making up the outer and inner layers are bedded nearly parallel to the rostral bone at the base. However, more distally positioned micro-layers of the outer layer are more anteverted distally. The micro-layers of the intermediate layer are bedded nearly perpendicular to those of the outer and inner layers on the median plane. The growth of micro-layers in the intermediate layer adds thickness to the rhinotheca, which causes the difference in profile between the beak bone and the rhinotheca in the distal portion of the beak. Moreover, the entire intermediate layer grows distally as new proximal micro-layers form. The outer layer is dragged distally by the intermediate layer as a result of its distal growth, for the three layers are closely packed to each other at their boundaries. Furthermore, the occurrence of the intermediate and inner layers in the distal portion of the rostral bone may be because the distal end of the beak is frequently used and worn, and the rhinotheca therefore needs to be replaced more frequently at the distal end. The rhinotheca structure described here will be an important and useful factor in the reconstruction of the beaks of birds in extinct taxa.

Void structure stability in wet granular matter and its application to crab burrows and cometary pits.

Cohesive granular matter can support stable void structures, which can universally be found in various scenes from everyday lives to space. To quantitatively characterize the stability and strength of a void structure in cohesive granular matter, we perform a simple tunnel-compression experiment with wet granular matter. In the experiment, a horizontal tunnel in a wet granular layer is vertically compressed with a slow compression rate. The experimental result suggests that the tunnel deformation can be classified into the following three types: (i) shrink, (ii) shrink with collapse, and (iii) subsidence by collapse. Using the experimental result, we estimate the stable limit of various void structures in a cohesive granular layer from crab burrows on a sandy beach to the pits observed on cometary surfaces.

How does the curvature of the upper beak bone reflect the overlying rhinotheca morphology?

The beak has independently been evolved accompanied by the edentulism in many tetrapod linages, including extant Testudinata and Aves, and its form and function have been greatly diversified. The beak is formed by beak bones and the overlying keratinous cover, although their profiles are different from each other. Therefore, it is difficult to reliably reconstruct the entire profile of the beak in extinct taxa, whose keratinous tissues are rarely preserved. For elucidation of the morphological relationship between beak bone and overlying keratinous cover, we compared the curvature distribution of the culminal profiles of the upper beak bone and the overlying keratinous cover (rhinotheca) with each other using CT-scan, in 66 extant testudinatan and avian specimens (Aves: 33 genera, 24 families; Testudinata: 12 genera seven families). In both, rhinotheca and beak bone, the curvature of the profile was nearly constant rostral to a certain point, which was defined as the transition point, and the transition points of the rhinotheca and beak bone were close to each other. The profiles of the rhinotheca and beak bone rostral to their transition point were different in curvature and length. However, the ratio between the curvatures of rhinotheca and the beak bone strongly correlated with the arc angle of the rostral culminal profiles of the beak bone. The upper beak profile in extinct taxa is expected to be reconstructed more reliably using the abovementioned relationship between the beak bone and the rhinotheca.

Fitting unanchored puzzle pieces in the skeleton: appropriate 3D scapular positions for the quadrupedal support in tetrapods.

Deducing the scapular positions of extinct tetrapod skeletons remains difficult, because the scapulae and rib cage are connected with each other not directly by skeletal joint, but by thoracic muscles. In extant non-testudine quadrupedal tetrapods, the top positions of the scapulae/suprascapulae occur at the anterior portion of the rib cage, above the vertebral column and near the median plane. The adequacy of this position was tested using three-dimensional mechanical models of Felis, Rattus and Chamaeleo that assumed stances on a forelimb on a single side and the hindlimbs. The net moment about the acetabulum generated by the gravity force and the contractive forces of the anti-gravity thoracic muscles, and the resistance of the rib to vertical compression between the downward gravity and upward lifting force from the anti-gravity thoracic muscle depend on the scapular position. The scapular position common among quadrupeds corresponds to the place at which the roll and yaw moments of the uplifted portion of the body are negligible, where the pitch moment is large enough to lift the body, and above the ribs having high strength against vertical compression. These relationships between scapular position and rib cage morphology should allow reliable reconstruction of limb postures of extinct taxa.

Synthesis and biological evaluation of steroidal derivatives bearing a small ring as vitamin D receptor agonists.

A novel series of 3-ketolithocholic acid derivatives as well as estrone derivatives bearing a small ring for the conformational fixation of the side chain were synthesized by using a catalytic [2+2] cycloaddition and a ring-contraction rearrangement. The steroidal derivatives were evaluated for transcriptional activation of vitamin D receptor by luciferase reporter assays. Among them, two estrone derivatives showed a higher efficacy of the transactivation of vitamin D receptor than 3-ketolithocholic acid, and the small ring moieties were found to be important for the efficacy.