Objective:To report a Chinese pedigree with autosomal dominant Waardenburg syndrome, and to identify causative gene mutations.Methods:Clinical data and peripheral blood samples were collected from the proband and her parents. Genomic DNA was extracted, gene mutations were detected through a next-generation skin-targeted sequencing panel, and Sanger sequencing was performed to verify causative mutations.Results:The proband clinically presented with irregular white patches on the abdomen and lower limbs, moderate to severe sensorineural deafness in the right ear, and iris heterochromia in both eyes. The proband′s mother presented with iris heterochromia in both eyes, epicanthus, early canities and thick eyebrows. In the family, both the proband and her mother were diagnosed with Waardenburg syndrome. A causative frameshift mutation c.976-977delinsT (p.Thr327Profs*54) was identified in both the proband and her mother, which caused the AG to TT base substitution at positions 976 - 977 in the coding region of exon 7 of the PAX3 gene, resulted in a frameshift from the amino acid position 327 to 54 in the PAX3 protein (threonine was substituted by proline at amino acid position 327). The proband′s father showed a normal phenotype, and his genetic test results were negative.Conclusion:The novel frameshift mutation c.976-977delinsT (p.Thr327Profs*54) in the PAX3 gene may contribute to the clinical phenotype of the patients with Waardenburg syndrome in the family.

In recent years, mass spectrometry has been widely used to study membrane protein structure and function. However, the application of mass spectrometry to study integral membrane protein is limited because there are many hydrophobic amino acids in the trans-membrane domain of integral membrane protein to cause low sequence coverage detected by LC-MS/MS. Therefore, we used vitamin K epoxide reductase (VKORC1), a human integral membrane protein, as a model to optimize the digestion conditions of chymotrypsin, and developed an in-gel digestion method of chymotrypsin to improve sequence coverage of membrane protein by mass spectrometry. By exploring the effects of calcium concentration, pH value and buffer system on the percentage of sequence coverage, number of total detected and types of unique peptide, and the size of unique peptide, sequence coverage and peptide diversity could be considered under condition of Tris-HCl buffer with 5-10 mmol/L calcium ion concentration and pH value 8.0-8.5. This method could make the sequence coverage of membrane protein to reach more than 80%. It could be widely used in the study of membrane protein structure and function, identification of interaction site between membrane proteins, and identification of binding site between membrane protein and small molecular drug.
Amino Acid Sequence ; Chromatography, Liquid ; Chymotrypsin/metabolism* ; Digestion ; Humans ; Membrane Proteins ; Tandem Mass Spectrometry ; Trypsin ; Vitamin K Epoxide Reductases

Microsatellite is a genetic marker, explored recently. In order to improve related studies on genetics of Anopheles gambiae, simple sequence repeats of the entire mosquito genome with 1～6 bp nucleotide motifs were analyzed.Abundance and distribution of microsatellites across the A.gambiae genome were analyzed and compared between various (exons, introns and intergenic) regions of all the chromosomes. About 2.14% of the mosquito genome was occupied by SSRs. Chromosome X had the maximum density of SSRs. Abundance orA repeats was similar to C repeats. AC was a little more than two times as abundant much as AG. However, AT and CG repeats were rare. For tri- and tetramer repeats, AGC,AAAC and AAAT predominated while ACG, ACT, AGG, CCG, ATGC, CCCG, ACTG, AACT, ACGT, AGAT, CCGG,ACCT and AGCT were rare. For some pentamer repeats, one was completely absent on a certain chromosome, even on several chromosomes. SSRs in exons of all chromosomes were less abundant than in introns and intergenic regions except for mono- and dimer repeats in exons of chromosome 2L. Abundance and distribution of SSRs on the two arms of each chromosome showed much in common.

Among a total of 276 upper extremities dissected it was found that 38 (13.7%) of the cases showed variations in the origin and course of the main arterial channels in brachium and antebrachium. In the 138 cadavers the arterial variations appeared bilaterally in 5 bodies (3.62%) and unilaterally in the other 28 bodies (20.29%). The anomalous arteries occurred more often on the right (22, 7.96%) than on the left (16, 5.80%). The patterns of variation encountered were grouped as follows: 1. Anomalous radial artery (1) High originated radial artery in 17 (6.16%) extremities (right 11, left 6). (2) Unusual duplication of the termination of the radial artery in 3 (1.13%)extremities (right 2, left 1). 2. Superficial ulnar artery According to the site of origin above or below the intercondylar line the arteries were divided into two groups: (1) Highly originated superficial ulnar artery in 7 (2.53%)—right 4, left 3. (2) Lowly originated superficial ulnar artery in 5 (1.81%)—right 3, left 2. 3. Superficial median artery and large calibered median artery were found in 6 extremities. The bifurcation of brachial artery into radial and ulnar arteries in 140 sides of male adults was mostly (82.14%) below the intercondylar line with a range from 2.6 cm. to 4.0 cm. The surgical significance of several of the striking variations was discussed.

Among 150 Chinese forearms dissected, an accessory tendon or tendons in the abductor pollieis longus muscle were found in 140 sides (93.3%), while in the rest (6.7%) the anomaly did not exist. In these 140 forearms, one accessory tendon was found in 97 (64.7%)and in the remaining 43 (28.7%) two accessory tendons were present. As regards the extensor pol- licis brevis muscle, one accessory tendon was found in only two cases (1.3%) in the whole series. It was noted that the two tendons of the abductor pollicis longus and extensor pollicis brevis pass through the same osteofibrous canal in 59 (39.4%), through two completely separated osteofibrous canals in 12 (8%), and in 79 out of the 150 forearms dissected. The two osteofibrous canals were partially separated as follows: in 32 (21.3%), the distal half only was separated by a thick fibrous band while in the other 47 (31.3%) only the distal third or even less was separated. The insertions of the main tendon and accessory tendons of the abductor pollicis longus muscle was even more extensive and complicated. Only in 3 cases the insertion could be regarded as normal, in the other 147 (98%), the insertion was either into the base of the 1st metacarpal bone, the abductor pollicis brevis muscle, the capsule of the 1st carpo-metacarpal joint, the greater multangular bone, the opponens pollicis muscle, the fascia over the thenar eminence or the navicular bone. The implications of the variations were discussed.

The arterial supply of the human odontoid process was studied in 48 cadavers of different ages by dissection, clearing, and radiologic methods. The results were as follows:1. The odontoid process obtains its blood supply directly from the anterior and posterior ascending arteries arising from the vertebral artery and indirectly from the horizontal arteries arising from the ascending pharyngeal artery. Around the odontoid process these arteries form a peculiar anastomosis, the upper part of which is called the apical arch.2. There are two groups of nutrient arteries, the basal and the apical nutrient arteries, in the odontoid process. The basal nutrient arteries penetrate into the process at the base through the anterolateral aspect and the central part of the posterior aspect, mostly 1 branch (81.94?3.21％) from either side. The apical nutrient arteries enter it at the apex, and are usually divided into 2 branches (69.44?7.68％). The outer diameters of the basal nutrient arteries in the new horns, the children, and the adults are 0.09～0.15mm, 0.16～0.19mm, 0.24～0.29 mm, and those of the apical nutrient arteries are 0.03～0.06mm, 0.06～0.10mm, and 0.10～0.14 mm respectively.3. The pattern of arterial distribution within the odontoid process varies with the age. In subjects under twelve years the apex of the process has not yet ossified completely, and no anastomosis can be found between the arteries at the base and those at the apex. In adults ossification has been completed. The intraosseous arteries connect with each other and form an anastomosis network, which is most abundant at the base.

The arterial supply of the human sacrum and coccyx was studied in 68 fresh cadavers of different ages by dissection, clearing, casting and radiography.The nutrient arteries of the sacral vertebrae penetrate the body from the dorsal, ventral and lateral surfaces of the body. The central branches are the main nutrient arteries of the body. Their numbers are constant and do not increase with the advancement of age. The peripheral branches are variable and supply only the outer collar of the body. According to the distribution of nutrient artery within the body of the sacrum, they can be classified into three types 1. predominant ventral nutrient artery pattern, 2. predominant dorsal nutrient artery pattern and 3. balanced nutrient artery pattern. It is found that in S_4 and S_5, type 1 occurs more frequently while type 2 occurs usually in S_1 to S_3. Type 1 is relatively common in adults but type 2 is prevailing in fetuses. There is few balanced pattern in different age groups. Both ventral and dorsal nutrient arteries are distributed in the central zone of the body and the lateral nutrient arteries supply that portion near the intervertebral foramen in the adults. The nutrient arteries within body anastomose with each other to form a dense arterial network.The nutrient arteries of the lateral part of the sacrum enter the bone through its ventral, dorsal and medial aspects. Among the dorsal nutrient arteries, there is a main artery that supplies the lateral mass.The arterial supply to the coccyx is scarce. It enters the coccyx mainly through the ventral surface.

In fifteen human fetuses spcimens the peritoneal stomata were studied with SEM and TEM, and measured by image processing system. In order to prove that the peritoneal stomata are the passageway of absorbed matter from the peritoneal cavity, animal experiments were made. There are two types of the mesothelial cells on diaphragmatic peritoneum, i. e. the cuboidal cells and the flattened cells. The peritoneal stomata, which arranged in clusters or strips, were only found between the cuboidal cells. The shape and size of the stomata were often irregular. The average area of the stoma on the muscular portion is 10.43?1.61?m~2, while on the tendinous portion is 7.93?1.67?m~2. The connective tissue underlies below the stomata, under which no basement membrane was found. Many lymphatic capillaries were observed in the connective tissue, which may promote absorption of matter from the peritoneal cavity. In animal experiments, some particles of trypan blue were absorbed through the stomata of rabbit diaphragmatic peritoneum. The authors consider that the stomata, are first observed in human, are important pathway for draining matter from the peritoneal cavity.

The arterial supply of human cervical vertebral bodies (C_3-C_7) was studied in 74 fresh cadavers of different ages by dissection and translucent preparation.1. The cervical vertebral bodies mainly obtain their blood supply from the vertebral artery, but the lower two (C_6-C_7) also receive their blood supply from the branches of the inferior thyroid, the deep cervical, the costocervical trunk, the highest intercostal and the subclavian artery. These arteries form a ladder-like anastomosis on the anterolateral surface of the vertebral bodies, and a rectangular or hexagonal anastomosis on the dorsal surface.2. The nutrient arteries enter the vertebral body from the anterolateral and dorsal aspects. They can be divided into the central branches which reach the center of the body and the peripheral ones which lie on the peripheral part of the body. Each of the central branches appears as a straight, unbranching stem. Their centrifugal terminals at the center of the body are arborized and extend to the upper, lower, left and right part of the body to supply the central core of the vertebral body which corresponds to the area of the nucleus pulposus. The peripheral branches, short and early branched, supply the peripheral part of the vertebral body which corresponds to the area of the annulus fibrous.3. The number of the central branches on the anterolateral aspect varies between 0-3 and that on the dorsal aspect is 0-2. The number of the peripheral branches on the anterolateral aspect is 2-13 and that on the dorsal aspect is 0-6. These branches anastomose with each other within the body of the vertebra. The end artery only appears in the developing cartilaginous regions of the body.

By transmission electron microscopy and freeze etching technique 15 human fetuses were utilized to study the ultrastructure of the mesothelial cells on the parietal peritoneum. The mesothelial cells of the diaphragmatic peritoneum contained numerous vesicles which were frequently communicated with the free surface, the basement membrane, intercellular space and the peritoneal stomata. Some of the vesicles seemed to fuse each other and form vacuoles. Vacuoles also occurred close to, or communicated with the basement membrane and cell free surface. Sometimes they appeared as secretory particles. The microvilli contained vesicles opened to the free surface. The mesothelial cells on the pelvic wall displayed abundant endoplasmic reticulum and Golgi apparatus, but scanty vesicles. So, the mesothelial cells on parietal peritoneum of human fetuses might be classified into two types, i. e. the vesiclecontaining cells on the diaphragmatic peritoneum and the ER-containing cells on the peritoneum of the pelvic wall. The vesicle-containing cells seemed to uptake material from the peritoneal cavity. Abundant endoplasmic reticulum and Golgi apparatus reflected a high synthetic activity, hence the ER-containing cells might be possibly related to the production of peritoneal fluid.