Sarcopenic obesity is associated with a faster decline in renal function in people with type 2 diabetes.

AIM: To evaluate the association between sarcopenic obesity and the decline in estimated GFR in people with type 2 diabetes. METHODS: We enrolled 745 people with type 2 diabetes (mean age 64.6 years, 53.6% men). Body composition was evaluated using dual-energy X-ray absorptiometry. Skeletal muscle index, calculated as appendicular non-fat mass (kg) divided by height squared (m2 ), was used to determine sarcopenia. Sarcopenic obesity was defined as the coexistence of sarcopenia and a ratio of android to gynoid fat mass greater than the median values in each gender. The association of sarcopenic obesity both with the annual rate of decline in estimated GFR and a >30% decline in estimated GFR was evaluated using multivariate linear regression models and Cox proportional hazard models, respectively. RESULTS: Participants with sarcopenic obesity were at an increased risk of a high annual rate of decline in estimated GFR, even after adjustment for the confounding variables (standardized ß = -0.228, P <0.001). Sarcopenic obesity was also significantly associated with risk of a >30% decline in estimated GFR (hazard ratio 4.52, 95% CI 2.16-9.47; P < 0.01) in multivariate model. CONCLUSIONS: Sarcopenic obesity evaluated by dual energy X-ray absorptiometry is associated with a faster decline in renal function in people with type 2 diabetes.

Indications for surgical treatment of funnel chest by chest radiograph.

Forty-seven children with funnel chest (FC) who underwent sternal elevation and 210 normal children were examined to determine the indications for surgical treatment using the vertebral index (VI) and frontosagittal index (FSI). In normal children VI gradually increased and FSI gradually decreased with age. Both indices changed significantly at 3 years of age. Although the VI of FC patients decreased significantly from 33.8 +/- 7.6 (n=40) to 24.4 +/- 3.9 (n=38) postoperatively (P < 0.0001), it was significantly larger than that of normal children over 3 years of age (20.2 +/- 2.2, n=150) (P < 0.0001), and although the FSI of FC patients increased significantly from 22.0 +/- 7.0 (n=40) to 34.5 +/- 6.5 (n=38) postoperatively (P < 0.0001), it was significantly smaller than that of normal children over 3 years of age (41.1 +/- 4.0, n=150) (P < 0.0001). Since many patients had a thin and flat chest despite excellent correction, their postoperative indices were not normal. There was a correlation between VI and FSI in normal children and a high degree of correlation between VI and FSI both before and after operation in FC patients. We conclude that a VI of more than 27 and/or a FSI of less than 29 are indications for surgical treatment based on the mean VI + 3SD and FSI - 3SD of normal children over 3 years of age. These values are almost equal to the mean VI - SD and FSI + SD of patients with physical, cosmetic, and/or psychological disturbances. However, it is not necessary to measure both indices simultaneously. Postoperative VI and FSI did not always reflect the degree of chest-wall depression in FC patients because of their flat chests.

Cancer chemopreventive agents, serratane-type triterpenoids from Picea jezoensis.

Seven serratane-type triterpenoids isolated from the cuticle of Picea jezoensis (Sieb. et Zucc.) Carr. jezoensis (Pinaceae) and the stem bark of Picea jezoensis (Sieb. et Zucc.) Carr. hondoensis (Mayer) Rehder (Pinaceae) were studied their possible inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). All compounds showed strong inhibitory effects on the EBV-EA activation, being stronger than that of oleanolic acid, which exerts on cancer preventive activity in animal carcinogenesis models. Among these compounds, 13alpha, 14alpha-epoxy-3beta-methoxyserratan-21beta-ol and 3beta-methoxy-21alpha-hydroxyserrat-14-en-29-al were investigated for the inhibitory effects in a two-stage mouse skin carcinogenesis test on mouse skin using 7,12-dimethylbenz[a]anthracene as initiator and TPA as promoter. 13alpha,14alpha-Epoxy-3beta-methoxyserratan-21beta-ol was found to exhibit the excellent anti-tumor promoting activity in the in vivo carcinogenesis test.

Structure and stereochemistry of epoxyserratanes from the cuticle of Piceajezoensis var. jezoensis.

Three new epoxytriterpenes, 14 beta,15 beta-epoxy-21 beta-hydroxyserratan-3-one (1), 13 alpha,14 alpha-epoxy-21 alpha-methoxyserratan-3-one (2), and 13 alpha,14 alpha-epoxy-3 beta-methoxyserratan-21 beta-ol (3), were isolated together with two known triterpenoids, 21 alpha-methoxyserrat-13-en-3-one (4) and 21 beta-hydroxyserrat-14-en-3-one (5), from the cuticle of Picea jezoensis var. jezoensis. The structures of these new compounds were established on the basis of spectral data (NMR, MS) and single-crystal X-ray analyses (1 and 2) and partial synthesis (2 and 3).

Cryoprotective effect of the serine-rich repetitive sequence in silk protein sericin.

The silk proteins, fibroin and sericin, are produced in the silk gland of Bombyx mori, and hydrophilic sericin envelops fibroin with successive sticky layers in the formation of a cocoon. To study the biological functions of sericin, we focused on the serine-rich sericin peptide consisting of 38 amino acids, which is a highly conserved and internally repetitive sequence of a sericin protein. The corresponding gene was chemically synthesized, and the PCR-amplified gene was ligated to oligomerize sericin peptide and fused at the amino terminus to a His-tagged and proteolytic cleavage sequence in an inducible expression vector. When the dimers of sericin peptides were overexpressed in Escherichia coli, the transformants showed a prominent increase in cell viability after freezing in medium. Further, the purified dimeric sericin peptide from E. coli was found to be effective in protecting lactate dehydrogenase from denaturation caused by freeze-thaw. Both of these protective effects against freezing stress in cells and proteins were also observed with sericin hydrolysate. These results indicate that this unique sericin peptide, like sericin, has a high cryoprotective activity and will be valuable as a new biomaterial for industrial use.

Roles of amino acid residues near the chromophore of photoactive yellow protein.

To investigate the roles of amino acid residues around the chromophore in photoactive yellow protein (PYP), new mutants, Y42A, E46A, and T50A were prepared. Their spectroscopic properties were compared with those of wild-type, Y42F, E46Q, T50V, R52Q, and E46Q/T50V, which were previously prepared and specified. The absorption maxima of Y42A, E46A, and T50A were observed at 438, 469, and 454 nm, respectively. The results of pH titration for the chromophore demonstrated that the chromophore of PYP mutant, like the wild-type, was protonated and bleached under acidic conditions. The red-shifts of the absorption maxima in mutants tended toward a pK(a) increase. Mutation at Glu46 induced remarkable shifts in the absorption maxima and pK(a). The extinction coefficients were increased in proportion to the absorption maxima, whereas the oscillator strengths were constant. PYP mutants that conserved Tyr42 were in the pH-dependent equilibrium between two states (yellow and colorless forms). However, Y42A and Y42F were in the pH-independent equilibrium between additional intermediate state(s) at around neutral pH, in which yellow form was dominant in Y42F whereas the other was dominant in Y42A. These findings suggest that Tyr42 acts as the hinge of the protein, and the bulk as well as the hydroxyl group of Tyr42 controls the protein conformation. In all mutants, absorbance at 450 nm was decreased upon flash irradiation and afterwards recovered on a millisecond time scale. However, absorbance at 340--370 nm was increased vice versa, indicating that the long-lived near-UV intermediates are formed from mutants, as in the case of wild-type. The lifetime changes with mutation suggest the regulation of proton movement through a hydrogen-bonding network.

Inhibition of glucocorticoid-induced apoptosis by the expression of antisense gene of mitochondrial ATPase subunit 6(1).

To isolate the apoptosis-linked genes involved in the cell death of thymocytes induced by glucocorticoids, we developed a functional cloning assay. Murine CD4(+)CD8(+) thymic cell line 2-257-20 cells were transfected with cDNA expression libraries obtained from a dexamethasone-resistant cell line. The transfected cells were selected in the presence of dexamethasone, and the plasmids which episomally expanded were then extracted from the surviving cells. One of the rescued cDNAs was found to be an antisense cDNA fragment identical to the mouse mitochondrial ATPase 6 gene. In the stable transfectants with the ATPase 6 antisense gene, the induction of apoptosis by dexamethasone was significantly delayed. Furthermore, the ATP synthesis in these transfectants was also reduced to some extent. ATPase 6 is a subunit of F(o)F(1) ATPase and our results support that ATP synthesis from the mitochondria is necessary for the induction of apoptosis induced by glucocorticoids.

Radiographic and computed tomographic analysis of the position of the tibial tubercle in recurrent dislocation and subluxation of the patella.

Among the factors that influence the patellofemoral alignment and stability, the position of the tibial tubercle was evaluated using plain radiographs and computed tomography (CT). Radiographs of the Q-angle with the knee in full extension and quadriceps muscle relaxed showed that mean Q-angles in the dislocation or subluxation group and control group were 13.8 degrees and 14.3 degrees, respectively. The major reason for this deceptive measurement was a patella positioned laterally in the dislocation or subluxation group. Better estimation was made with the modified Q-angle measurement by using the most concave point of the intercondylar notch as a reference point instead of the center of the patella. By overshadowing CT images of the levels of the patellofemoral joint and the tibial tubercle, it was possible to evaluate the location of the tibial tubercle in relation to the femoral trochlea. Both the angular measurement (lateral deviation angle [LDA]) and the distance measurement (lateral deviation index [LDI]) showed that the tibial tubercle in the dislocation or subluxation group was rotated externally and shifted laterally (LDA, 36.3+/-7.0 degrees and LDI, 30.1+/-5.6) compared with the control group (LDA, 20.2+/-7.1 degrees and LDI, 15.1+/-5.6). Further study is warranted to elucidate the exact mechanism of recurrent patellar dislocation and subluxation.

Localization of insulin receptor-related receptor in the rat kidney.

Insulin receptor-related receptor (IRR), a member of the insulin receptor family, is most abundantly expressed in the kidney. However, its endogenous ligand and physiological roles are still unknown. To elucidate the physiological role of IRR, an orphan receptor, in the kidney, we examined the localization of IRR mRNA and its immunoreactivity in the rat kidney by in situ hybridization and immunohistochemistry, respectively. IRR mRNA was found to be exclusively localized in the cortical collecting duct. The localization of IRR immunoreactivity was consistent with that of IRR mRNA. Furthermore, IRR immunoreactivity was found to be localized on the basolateral plasma membrane of the epithelial cells that were a minor cell subpopulation (20 to 30%) of the duct. The present findings indicated that IRR in the kidney was exclusively localized on the basolateral plasma membrane of type B intercalated cells of the cortical collecting duct.

Clinical significance of thallium-201 and gallium-67 scintigraphy in pulmonary tuberculosis.

One hundred and thirty-nine patients with pulmonary tuberculosis were evaluated by means of gallium-67 and thallium-201 scintigraphy. The disease was clinically active in 83 and inactive in 56. The uptake ratio between the lesion and the contralateral normal lung field was calculated. The ratio determined by 67Ga scintigraphy was expressed as GR, and that determined by 201T1 scintigraphy (early or delayed) as ER or DR. The 201T1 retention index (RI) was calculated using the following equation: RI = DR - ER/ER x 100. The sensitivity, specificity and accuracy of 201T1 scintigraphy with respect to the activity of pulmonary tuberculosis were better than those of 67Ga scintigraphy (the figures for 201T1 scintigraphy were 88.0%, 82.1% and 85.6%, respectively, and those for 67Ga scintigraphy, 83.1%, 60.7% and 74.1%). We found a significant correlation between GR and ER, but there was no significant correlation between RI and ER. When the relationships between ER, C-reactive protein and 1-h erythrocyte sedimentation rate were examined among patients with abnormal uptake, no significant relationships were noted. In the 27 patients who could be followed up, GR and ER decreased with duration of the therapy, indicating a decrease in disease activity which was consistent with clinical findings. The RI was significantly higher in the early stages of therapy than in the later stages (P < 0.01), suggesting a temporary delay in 201T1 washout in the early stages. The washout seemed to be promoted by the effects of the therapy. Overall, it is concluded that 201T1 scintigraphy is more useful and more suitable than 67Ga scintigraphy for the evaluation of disease activity and therapeutic effects in patients with pulmonary tuberculosis.

Mutational analysis of the feedback sites of phenylalanine-sensitive 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase of Escherichia coli.

In Escherichia coli, aroF, aroG, and aroH encode 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase isozymes that are feedback inhibited by tyrosine, phenylalanine, and tryptophan, respectively. In vitro chemical mutagenesis of the cloned aroG gene was used to identify residues and regions of the polypeptide essential for phenylalanine feedback inhibition.

Expression of insulin receptor-related receptor in the rat brain examined by in situ hybridization and immunohistochemistry.

Insulin receptor-related receptor (IRR) is a member of the insulin receptor family. However, its endogenous ligand and physiological roles are unknown. To elucidate the physiological roles of IRR, an orphan receptor, in the brain, we examined its expression at mRNA and protein levels in the brain by in situ hybridization and immunohistochemistry, respectively. The expression of IRR mRNA in the brain was highly restricted to the forebrain including the nucleus of the diagonal band, medial septal nucleus, ventral pallidum, accumbens nucleus and caudate putamen, and the brainstem including the prepositus hypoglossal nucleus, medial vestibular nucleus, gigantocell reticular nucleus, paragigantocellular nucleus and ventral cochlear nucleus. Most IRR mRNA-positive cells in the forebrain but not in the brainstem were cholinergic neurons. However, most IRR mRNA in the forebrain and brainstem was coexpressed with that of trkA, a high-affinity receptor for nerve growth factor. IRR-immunoreactive cell bodies were also detected in the forebrain and brainstem. The pattern of IRR immunoreactivity was similar to that of IRR mRNA. Its restricted pattern indicates that IRR plays unique roles in the brain, in contrast to insulin and insulin-like growth factor-I receptors, other members of the insulin receptor family, which are widely expressed in the brain.

Distribution of cellular repopulation and collagen synthesis in a canine anterior cruciate ligament autograft.

Whether the central core of an anterior cruciate ligament autograft reconstruction is nutritionally compromised at a time when revascularization is known to be complete has not been determined by methods that detect matrix synthesis. In a canine model of anterior cruciate ligament reconstruction with patellar tendon autograft, the adequacy of the supply of metabolites for cellular matrix synthesis was determined by autoradiographic analysis. Total collagen synthesis and cellularity were also quantified. Total collagen synthesis was found to be significantly elevated (p = 0.014 by analysis of variance) in the ligament reconstructions as compared with normal anterior cruciate ligaments or patellar tendons but cellularity was not (p = 0.13 by analysis of variance). Autoradiography demonstrated even distribution of [3H]proline incorporation throughout the graft and normal tissue. When revascularization was complete there was an adequate supply of metabolites for cellular synthesis of protein macromolecules within all regions of the ligament reconstruction. At 3 months after reconstruction, the grafts were found to be actively remodeling their collagen matrix. Since the long-term function of an anterior cruciate ligament autograft is dependent on viable fibroblasts to maintain the collagen matrix the canine anterior cruciate ligament reconstruction contains living cells that are able to remodel the matrix under appropriate conditions.

Non-destructive, ultrasonic evaluation of meat quality in live Japanese Black steers from coloured images produced by a new ultrasound scanner.

An improved colour scanning scope was used for evaluating meat quality (marbling) of live Japanese Black steers. This equipment consisted of a small size ultrasonic probe (2 MHZ) and LCD display. Seventeen fattened Japanese Black cattle were scanned at the region of the 7th rib about one week before slaughter. A picture of the cross-sectional area of the back was obtained immediately after applying the probe and contained 15 colours representing different signal strengths. The time for each scan was 2 seconds. The picture signals were fed into a computer for rapid estimation of fat percentage of the M. longissimus thoracis. After slaughter, the fat content and chemical characteristics were determined on the M. longissimus thoracis obtained from the same rib section. The range of fat content was 7.0 to 23.7% (average 18.47%). A high correlation coefficient (r = 0.90; r.s.d. = 2.01%) was obtained between actual fat percentage of the M. longissimus thoracis and colour-scanning scope SR200 estimates based on the percentage of the weak blue dot(1) in the echo. Estimates of the subcutaneous fat thickness and the cross-sectional area of M. longissimus thoracis from the scans were in good agreement with the actual carcass measurements (r = 0.69; r.s.d=0.52 cm and r = 0.81; r.s.d. = 4.26 cm(2), respectively). These results show that the new colour scanning scope is a useful instrument for estimating meat quality (marbling) in live cattle.

Expression of insulin receptor-related receptor mRNA in the rat brain is highly restricted to forebrain cholinergic neurons.

Insulin receptor-related receptor (IRR) is a member of the insulin receptor family. However, its endogenous ligand and the physiological roles of IRR are unknown. To elucidate the physiological roles of IRR in the brain, we examined the expression of its mRNA in the rat brain by in situ hybridization. In contrast to the widespread expression of insulin receptor and insulin-like growth factor-1 receptor mRNAs in the brain, the expression of IRR mRNA was highly restricted to the forebrain cholinergic neurons. All the forebrain cholinergic neurons expressed IRR mRNA. The present findings indicate that IRR has a selective role in the brain for forebrain cholinergic function.

Insulin receptor-related receptor messenger ribonucleic acid in the stomach is focally expressed in the enterochromaffin-like cells.

The insulin receptor-related receptor (IRR) is a member of the insulin receptor family. In contrast to the widespread expression of insulin receptor and insulin-like growth factor-I receptor messenger RNA (mRNA), the expression of IRR mRNA is highly restricted to the kidney and stomach. IRR mRNA in the kidney is focally expressed in the renal distal tubule cells. However, the cellular localization of IRR mRNA in the stomach remains to be elucidated. Here, we examined the cellular localization of IRR mRNA in the rat stomach by in situ hybridization. IRR mRNA in the stomach was abundantly localized in the basal third of the oxyntic glands of the fundic stomach. IRR mRNA in the stomach was colocalized with mRNA for histidine decarboxylase, a marker for the enterochromaffin-like (ECL) cells, indicating that the expression was restricted to ECL cells. ECL cells actively produce and store histamine, which is an important physiological stimulant of acid secretion from the parietal cells. The preferential localization of IRR mRNA in ECL cells suggests that the IRR plays an important role in the function of these cells.

Phosphorylation of an inhibitory subunit of cGMP phosphodiesterase in Rana catesbeiana rod photoreceptors. I. Characterization of the phosphorylation.

Interaction between the inhibitory subunit (P gamma) and catalytic subunits of cGMP phosphodiesterase is essential for the regulation of cGMP phosphodiesterase in vertebrate rod photoreceptors. P gamma phosphorylation in vitro has been studied using a kinase which is extracted from amphibian rod outer segments. Various chromatographies of the kinase preparation using ionic exchange, gel filtration, and heparin-Sepharose columns indicate that a kinase with M(r) 70,000 is responsible for the P gamma phosphorylation. The kinase does not require any of the known activators for protein kinases but is inhibited by cGMP in a concentration-dependent manner. Together with analysis by laser-desorption mass spectrometry, measurement of 32P radioactivity in phosphorylated P gamma indicates that P gamma extracted with GTP-bound transducin alpha subunit is not phosphorylated and that a phosphate is incorporated into more than 80% of the P gamma by the kinase. Phosphoamino acid analysis, sequencing of phosphorylated peptides derived from phosphorylated P gamma, and phosphorylation of synthetic peptides indicate threonine 22 in P gamma is phosphorylated by the kinase. Phosphorylated P gamma has a higher inhibitory activity for active cGMP phosphodiesterase than non-phosphorylated P gamma. These data suggest that threonine 22 in P gamma is phosphorylated by a specific kinase and that the P gamma phosphorylation governs the interaction between P gamma and catalytic subunits of cGMP phosphodiesterase in vertebrate rod photoreceptors.

Application of in vitro bromodeoxyuridine labeling of oral cancers.

We used immunohistochemical methods to investigate whether in vitro labeling with bromodeoxyuridine (BrdU) of oral cancer tissues is useful in pathological diagnosis. BrdU-positive cells were found around the basal region of the epithelium in normal oral mucosa, in a papilloma specimen and in tissue with moderate dysplasia. The labeling rates (percentage of BrdU-positive cells, LR) for these cases were 1.2%, 6.9% and 7.5%, respectively. In well-differentiated carcinoma, more layers of BrdU positive cells were observed from the basal layer toward the surface. The LR in this region was 13.9-17.0%. In vitro BrdU labeling of oral tumors may be useful in pathological diagnosis, since the LR is lowest in normal tissues, higher in benign tumors, and highest in malignant tumor tissues.