[Analysis of clinical phenotype and genotype of Chinese children with disorders of sex development].

Objective: To explore the heterogeneity and correlation of clinical phenotypes and genotypes in children with disorders of sex development (DSD). Methods: A retrospective study of 1 235 patients with clinically proposed DSD in 36 pediatric medical institutions across the country from January 2017 to May 2021. After capturing 277 DSD-related candidate genes, second-generation sequencing was performed to analyzed the heterogeneity and correlation combined with clinical phenotypes. Results: Among 1 235 children with clinically proposed DSD, 980 were males and 255 were females of social gender at the time of initial diagnosis with the age ranged from 1 day of age to 17.92 years. A total of 443 children with pathogenic variants were detected through molecular genetic studies, with a positive detection rate of 35.9%. The most common clinical phenotypes were micropenis (455 cases), hypospadias (321 cases), and cryptorchidism (172 cases) and common mutations detected were in SRD5A2 gene (80 cases), AR gene (53 cases) and CYP21A2 gene (44 cases). Among them, the SRD5A2 mutation is the most common in children with simple micropenis and simple hypospadias, while the AMH mutation is the most common in children with simple cryptorchidism. Conclusions: The SRD5A2 mutation is the most common genetic variant in Chinese children with DSD, and micropenis, cryptorchidism, and hypospadias are the most common clinical phenotypes. Molecular diagnosis can provide clues about the biological basis of DSD, and can also guide clinicians to perform specific clinical examinations. Target sequence capture probes and next-generation sequencing technology can provide effective and economical genetic diagnosis for children with DSD.

A dual-label time-resolved fluorescence immunoassay for the simultaneous determination of cystatin C and ß2-microglobulin in urine.

BACKGROUND: Contrast media is widely used in clinical diagnostic and interventional procedures, but may cause damage to the kidney, that is, contrast-induced nephropathy. This study was to establish a dual-label time-resolved fluorescence immunoassay (TRFIA) for the simultaneous determination of renal function markers cystatin-C (Cys-C) and ß2-microglobulin (ß2-MG) for the early diagnosis and follow-up surveillance of contrast-induced nephropathy. METHODS: A sandwich immunoassay was used to detect the concentration of Cys-C, and the competitive immunoassay was used to detect the concentration of ß2-MG in 50 samples of urine. The performance of this dual-label TRFIA was evaluated and compared with commercial assays. RESULTS: The sensitivity for Cys-C detection was 1.26 ng/ml, the average recovery was 99.36%; The sensitivity for ß2-MG detection was 2.13 ng/ml, the average recovery was 100.18%. Bland-Altman analysis showed that the dual-label TRFIA method and the commercial kits had a good agreement, suggesting they can be used interchangeably in clinical urine analysis. CONCLUSION: The present dual-label TRFIA has high sensitivity, specificity and accuracy in clinical sample analysis. This method can be used for the early diagnosis and follow-up surveillance of the contrast-induced nephropathy.

Alternation of neurofilaments in immune-mediated injury of spinal cord motor neurons.

STUDY DESIGN: Observational cross-section study. OBJECTIVE: The objective of our study was to determine if phosphorylation of aggregated neurofilaments (NFs) would occur in autoimmune-mediated motor neuron injury. Our main hypothesis was that autoimmune-mediated damage of spinal cord motor neurons may influence NF phosphorylation and lead to NF aggregation. METHODS: A total of 20 guinea pigs were inoculated with bovine spinal cord anterior horn homogenates (experimental autoimmune gray matter model) and 20 guinea pigs were inoculated with phosphate-buffered saline (control). NF phosphorylation and aggregation were observed by immunohistochemistry and electron microscopic examination. Data were analyzed using Student's t-test with P<0.05 being considered significant. RESULTS: Abnormal phosphorylation and distribution of NF occurred in motor neurons and axons of animals with experimental autoimmune gray matter disease but not in the control animals. CONCLUSION: Aberrant accumulation and phosphorylation of neurofilaments in perikarya of spinal cord motor neurons occur in immune-mediated motor neuron death. As both immunologic response and alteration of neurofilaments are observed in amyotrophic lateral sclerosis (ALS) patients and aberrant neurofilament change harms motor neurons, our present findings suggest that autoimmunity-induced ALS may mediate in part through neurofilament modification.

Starch properties of mutant rice high in resistant starch.

As the staple food of over half the world's population, hot cooked rice high in resistant starch (RS) is of particular interest, which will have greater impact in the dietary prevention of diabetes and hyperlipidemia. A mutant rice high in RS in hot cooked rice, described as RS111, was comparatively studied with the wild type and common rice. Despite obviously low RS content in the raw milled rice, the RS content in the hot cooked rice of mutant RS111 was significantly higher than that of the wild type and common rice and, correspondingly, in vitro starch hydrolysis by porcine pancreatic alpha-amylase tends to be incomplete with low hydrolysis extent for the cooked mutant rice high in RS. Obvious differences in physicochemical properties, starch granule morphology, pasting properties, thermal properties, and X-ray diffraction pattern were observed among the mutant RS111, wild type, and common indica rice. The high-RS mutant was characterized by significantly higher apparent amylose content and crude lipid content, higher percentage of oval-shaped granules and bigger oval size, reduced paste viscosity, and low onset temperature, peak temperature, final temperature, enthalpy of gelatinization, and crystallinity.

Synthesis and characterization of three tetranuclear clusters containing a [Mo3OS3Sn]6+ cubane-like core.

Three heterometallic cubane-like clusters, [Mo3(mu 3-O)(mu 3-S)3(SnCl3)(dtp)3(py)3] (dtp = S2P(OC2H5)2-, py = C5H5N) (1), (PPN)[Mo3(mu 3-O)(mu 3-S)3(SnCl3)(dtp)3(mu-OAc)(py)] (OAc = CH3COO-, PPN = (C6H5)3PNP(C6H5)3+) (2), and (Et4N)[Mo3(mu 3-O)(mu 3-S)3(SnCl3)(dtp)2(mu-OAc)2(py)] (3) have been prepared by the reaction of [Mo3(mu 3-O)-(mu-S)3(dtp)4(H2O)] (4), [Mo3(mu 3-O)(mu-S)3(dtp)3(OAc) (py)] (5), and [Mo3(mu 3-O)(mu-S)3(dtp)2(OAc)2 (py)] (6) with SnCl2, respectively. They have been characterized by IR, UV-vis, 31P NMR, 95Mo NMR, and X-ray structure analysis. All of these heterometallic clusters have a [Mo3OS3Sn]6+ core but contain a different arrangement of peripheral ligands. As far as the neutral cluster 1 is concerned, there is no bridging OAc ligand, while only one bridging OAc ligand is observed for cluster 2 and two are for cluster 3. The Mo-Mo distances are about 0.03-0.04 A shorter than those of the starting trimolybdenum clusters. This indicates that the incorporation of SnCl3- fragment into (Mo3) clusters makes the Mo-Mo bonding enhanced. Crystal data for 1: triclinic, space group P-1, a = 10.7423(2) A, b = 14.0357(1) A, c = 16.9346(2) A, alpha = 84.054(1) degrees, beta = 87.095(1) degrees, gamma = 84.517(1) degrees, V = 2525.82(6) A3, Z = 2, R = 0.038 for 5584 reflections (I > 2.0 sigma(I)). Crystal data for 2: triclinic, space group P-1, a = 12.9529(1) A, b = 15.6324(2) A, c = 19.6355(1) A, alpha = 92.083(1) degrees, beta = 97.908(1) degrees, gamma = 110.337(1) degrees, V = 3677.41(6) A3, Z = 2, R = 0.034 for 8665 reflections (I > 2.0 sigma(I)). Crystal data for 3: monoclinic, space group P2(1)/n, a = 14.0852(5) A, b = 15.1324(5) A, c = 23.2691(7) A, beta = 97.371(1) degrees, V = 4918.7(3) A3, Z = 4, R = 0.049 for 4970 reflections (I > 2.0 sigma(I)).

Regulation of the depth of surface liquid in bovine trachea.

The luminal surface of airways is lined by a thin film of airway surface liquid (ASL). Physiological regulation of the depth of ASL has not been reported previously. In this paper, we have used low-temperature scanning electron microscopy of rapidly frozen specimens of bovine tracheal epithelium to demonstrate alterations in the depth of ASL in response to the cholinergic agonist methacholine. We first established that methacholine selectively stimulated airway glands, with maximal secretion at approximately 2 min and a return to baseline within approximately 5 min. A 2-min exposure to methacholine increased the depth of ASL from 23 to 78 microns. Thereafter, depth decreased linearly with time, reaching 32 microns at 30 min. The initial increase in depth was blocked by bumetanide, an inhibitor of active chloride secretion, whereas the slow decline back to baseline was inhibited by amiloride, a blocker of active sodium absorption. We conclude that the methacholine-induced changes in ASL depth reflect transient gland secretion followed by liquid absorption across the surface epithelium.

Effects of hydrostatic pressure on permeability of airway epithelium.

The presence of blood proteins and excess liquid in the airway lumen during airway inflammation may be secondary to extravasation and elevation of subepithelial hydrostatic pressure. This study examines how hydrostatic pressures of 5-20 cm H2O affect hydraulic conductivity and macromolecular permeability of primary cultures of bovine tracheal epithelium. Hydraulic conductivity was not altered by transepithelial pressure gradients of up to 20 cm H2O directed from the mucosal to serosal side of the tissue (m-s). By contrast, a 20-cm H2O s-m pressure resulted in a marked increase in hydraulic conductivity with the critical pressure lying between 10 and 20 cm H2O. Electrical conductance (i.e., permeability to ions) was not altered by m-s pressure gradients, or by a 5-cm H2O s-m gradient, but was increased by s-m pressures > or = 10 cm. Fluxes (s-m and m-s) of fluorescein and fluorescent dextrans (70 and 2000 kDa) were not altered by pressures of up to 20 cm H2O m-s. By contrast s-m pressure gradients of 20 cm H2O dramatically increased the s-m fluxes of these probes. The increases in flux were completely reversible. The results indicate that s-m pressure gradients greatly increase the hydraulic conductivity of airway epithelium by creating pores with an effective diameter greater than 54 nm.

Regulation of depth and composition of airway surface liquid.

The depth and composition of human airway surface liquid (ASL) may depend on secretion from airway glands, ion transport across the surface epithelium, goblet cell discharge, transepithelial gradients in hydrostatic pressure, and surface tension. Published values for the frequency of airway glands and for the secretory rates of individual glands suggest that total gland secretion in human trachea can amount to approximately 60 microL x cm(-2) x h(-1). Volume absorption directly measured across cultures of surface epithelium from human trachea is approximately 5 microL x cm(-2) x h(-1). These flows should alter the depth of ASL at +10 and -1 microm x min(-1). We have looked for changes in ASL depth of this magnitude using low-temperature scanning electron microscopy (LT-SEM) of rapidly frozen specimens of bovine trachea. Stimulation of gland secretion with methacholine led to an initial increase in depth of approximately 25 microm x min(-1) followed by a decline at approximately 1.5 microm x min(-1). Whereas the initial increase in depth was probably due to transient gland secretion, the later decline reflected active absorption of liquid across the surface epithelium. Finally, we present preliminary data showing that LT-SEM can be combined with X-ray microanalysis to determine the elemental composition of ASL.

[Clinical and experimental studies on qumotang for bleeding after medical abortion in 214 cases].

OBJECTIVE: To relieve the side-effect of heavy vaginal bleeding after medical abortion. METHODS: Two hundred and fourteen clinical cases were enrolled, 111 of them were treated with Qumotang (QMT) and 103 cases were the control group. RESULTS: The average bleeding durations is 8.7 +/- 6.4 days and 13.6 +/- 7.1 days respectively. The percentages of the cases with bleeding amount less than normal menstruation in QMT group and the control were 18.44% and 30.63% respectively, the difference was significant (P < 0.05). CONCLUSIONS: QMT increased the intrauterine pressure and contraction frequency of rabbit uterus in vivo and it is the effective herbal recipe for bleeding after medical abortion. QMT presented its efficacy of removing blood stasis and stopping bleeding, increasing the contraction of uterine muscles and vessels.

Ultrastructure of tracheal surface liquid: low-temperature scanning electron microscopy.

A layer of liquid lines the airways in the lung. Previous microscopic studies have suggested that it is in two phases, with a mucous gel lying above a periciliary sol. However, shrinkage artifacts due to chemical fixation, dehydration, and drying have prevented reliable estimates of the depth of these layers. To avoid such problems, we have studied the surface liquid of bovine trachea by low-temperature scanning electron microscopy (LTSEM). A polished copper probe cooled to liquid nitrogen temperature was applied to the mucosal surface of sheets of excised tracheal epithelium to effect rapid freezing of surface liquid. Tissue sheets were then mounted in an LTSEM (AMRay 1000A with Biochamber) which maintains samples at -180 degrees C with a Joule-Thompson refrigerator built into the stage. Tissues were fractured at right angles to the epithelial surface, coated with gold, and viewed, all at 10(-5) to 10(-6) torr without transfer through air. The sample was stable under the electron beam at accelerating voltages up to 20 kV. Epithelial features (nuclei, cilia, microvilli, mucous granules) were well preserved. The mucosal surface of the cells was covered with material on the order of 8 microns in depth. The mucous gel and periciliary sol could be seen as distinct layers and could be distinguished by the size and pattern of ice crystal voids generated by radiant-etching of the fractured surface of the sample.

Design of the oxygen and substrate pathways. VI. structural basis of intracellular substrate supply to mitochondria in muscle cells.

This paper quantifies the structures involved in the transport and oxidation of carbohydrates and fatty acids within the muscle cell. The structural capacity is measured on whole-body random samples of the musculature of dogs and pygmy goats and compared with maximal rates of oxygen consumption and substrate oxidation. Comparing dogs and goats of the same body size provided a 1.55-fold difference in the maximal rate of oxidation when related to muscle mass. As in previous studies, we found that the volume of mitochondria was approximately proportional to aerobic capacity. The maximal glucose flux from intracellular stores to mitochondria is 1.6 times greater in the dog than in the goat; we find that the amount of glycogen stored in the muscle cells is 4.2 times as great in the dog, but part of the intracellular glycogen pool is used for anaerobic rather than for oxidative metabolism. The maximal fatty acid flux from intracellular stores to mitochondria is 1.5 times larger in the dog, and the amount of lipid stored is 2.3 times as great in the dog. Every lipid droplet is in direct contact with the outer membrane of a mitochondrion and the contact surface area is 3.6 times greater in the dog than in the goat. Additional measurements are needed to investigate the role of structural limitation at this step. The amount of substrates stored intracellularly in the muscle cells of the dog is about twice as much as would match the differences in the maximal rates of utilization. This allows the endurance-specialized dogs to run for longer periods at higher rates of oxidation.

[Effect of er-xian decoction and its disassembled prescription on enzyme activities and their gene expression of antioxidant enzymes in aging rat].

In order to study the mechanism of aging, this experiment adopted molecular biological techniques to observe the effect of Er-Xian decoction (EXD)and its disassembled prescription on the enzyme activities and their gene expression of superoxide dismutase (SOD)and catalase (CAT). The results showed that in old rat, the amount of lipid peroxide (LPO) increased significantly; the enzyme activities such as SOD, CAT decreased significantly; and the lowering of these activities had a direct bearing on the gene expression level of their enzyme protein. After treatment by EXD and its disassembled prescription in aging rat, the amounts of LPO reduced significantly, the activities of SOD, CAT enhanced significantly. Moreover, it was observed that the increasing of these enzyme activities was related to the enhancing of their gene expression. It was suggested that EXD had the effect of anti-aging in that it could raise the activities of antioxidant enzymes and reduce the product of free radicals by means of regulating and enhancing the gene expression levels of their enzyme protein.

Lung lesions in experimental hydrostatic pulmonary edema: an electron microscopic and morphometric study.

Distinct barrier lesions and an apical-basal distribution of alveolar edema fluid in either moderate or high elevated pressure edema lungs have been found in previous studies. In the present study, quantitative measurements were obtained by using electron microscopy and morphometry of extravascular lung water and barrier lesions, on the relations between interstitial and alveolar edema fluid as well as between extravascular lung water and barrier lesions. The study further addressed the question of whether 6% bovine serum albumin (BSA) perfusion could induce lung ultrastructure alterations. It was found that interstitial fluid distribution is similar to that of alveolar edema fluid. Epithelial blebs are also distributed with an apical-basal gradient, and are always submerged in alveolar edema fluid. Perfusion with 6% bovine serum albumin does not induce any lung ultrastructure alterations. The results indicate that endothelium and epithelium play a different role in controlling fluid movement between capillary and extravascular spaces and thus in preventing the formation of interstitial and alveolar edema. Because the interaction of cells and tissue must be taken into account, simple physiological models of pulmonary fluid exchange may not be adequate to explain pulmonary edema formation.