Human micro-insemination by injection of single or multiple sperm: ultrastructure

The process of micro-insemination by single or muhiple spermtransfer into the perivitelline space (PVS) or by direct sperminjection into oocytes was examined by transmission electronmicroscopy. Spermatozoa from normal and oligozoospermic menwere injected into oocytes, obtained from consenting IVF patients,mostly by zona-puncture using micromanipulators. Spermatozoawere washed by the Percoll or Ficoll methods and capacitatedusing Whittingham's T₆ or modified Tyrode's medium or incubatedin strontium medium before injection. The women were stimulatedby three IVF methods and oocytes were recovered by laparoscopyor ultrasonography. Sixty-one oocytes were cultured in T₆ orHam's F-10 media (3–24 h) and were subjected to micromanipulation.Four oocytes were also studied after zona-drilling. Normal 2-pronuclearova were developed after single-sperm transfer satisfying allmorphological criteria of fertilization. Both monospermic andpolyspermic fertilization resulted after multiple sperm transfer,indicating that a vitelline block to polyspermy may exist inhumans. The majority of oocytes examined were unfertilized.Spermatozoa with intact or reacted acrosomes and those undergoingthe acrosome reaction were found in the PVS and in the ooplasm.Abnormal spermatozoa were also seen in these locations. Quantitatlonof acrosomal status in 16 oocytes after multiple-sperm transfer,revealed that 24% of spermatozoa were acrosome reacted or reactingin the PVS following Ficoll entrapment, while 76% of spermatozoawere intact (33% of these abnormal). Sperm transfer seemed tobe the least invasive, while direct sperm injection was comparativelydestructive to oocytes. Drilling with acid made larger breachesin the zona when compared with mechanical perforation and spermatozoaoccasionally escaped through breaches. Three 2-pronuclear ovaobtained after multiple sperm transfer have resulted in twopregnancies, in cases of severe oligozoo spermia, during thecourse of this study.     

Cushing's Syndrome Secondary to Olfactory Neuroblastoma

A case of olfactory neuroblastoma in a 36-year-old woman who presented with florid Cushing's syndrome is reported. A nasal polyp, which proved to be an olfactory neuroblastoma, was resected. The procedure was followed by complete remission from the endocrinologic abnormalities. Postoperatively, the patient was well for 5 years until recurrence of both Cushing's syndrome and the nasal polyp was noted. Following combined transnasal-transcranial resection of the tumor, which extended into the anterior cranial fossa, the patient again experienced complete remission of Cushing's syndrome. Immunohistochemistry showed the tumor to be positive for neuron-specific enolase, synaptophysin, chromogran-in, adrenocorticotropic hormone, (J-endorphin, and S-100 protein. Electron microscopy revealed neuritic processes containing microtubules and neurosecretory granules. This is the first reported case of Cushing's syndrome secondary to olfactory neuroblastoma.     

Scanning electron microscopic study of the human auditory ossicles

The human mallei, incudes and stapedes from 34 cadavers were examined using scanning electron microscope (SEM) to compare the bone surface type among different regions of auditory ossicles for males and females. On the malleus of both males and females, almost all of the surfaces showed a smooth fibrous appearance, characteristic of resting surface. Limited bone-forming or resorbing surfaces were identified on the malleus. As compared with the malleus, the percentage area of the resorbing surface and the vascular canal openings were higher on the incus and stapes, especially on the long process (Crus longum) of the incus and the neck of the stapes for both males and females. The percentage area occupied by the resorbing surface of the long process of the incus and the neck of the stapes correlated with that of the vascular canal openings. We consider that the malleus maintained the stable condition, while the long process of the incus and the neck of the stapes demonstrated marked bone resorption. We suppose that the bone erosion may be related to the vascularization in these regions. Though the percentage area of the resorbing surface and the vascular canal openings had the tendency to be high in females, we did not find any significant differences between the males and females. There was no significant correlation between the age and the area of resorbing surface or vascular canal openings.     

Wg3h细胞系转染PSV_2-neo质粒前后生长特性及表面超微结构改变的研究

本文对转染PSV_2-neo质粒后的Wg3h细胞系(Wg3h-neo)在长期传代中的生长特性和表面超微结构与母系Wg3h细胞进行了比较研究。结果发现:转染后Wg3h细胞的DNA合成及生长速度明显高于母系Wg3h细胞,生长饱和密度增大。在软琼脂培养中不形成集落,接种在裸鼠不长肿瘤。在扫描电镜下,细胞表面的微绒毛较母系Wg3h细胞丰富。Southern印迹杂交实验证明PSV_2-neo质粒已整合到宿主细胞的基因组中。转染后Wg3h细胞的生长特性和表面超微结构均发生了某些转化特征。     

Biological effects of static magnetic fields on the microcirculatory blood flowin vivo: a preliminary report

There have been few studies of the effect of static magnetic fields on microcirculatory haemodynamics in vivo. The rat skinfold transparent chamber technique was used, which provides an excellent means of observing and quantifying direct in vivo microvascular haemodynamic responses to static magnetic fields up to 8 T. An intravital videomicroscope was used to measure the changes in blood flow before and after exposure to a magnetic field for 20 min in a horizontal type superconducting magnet with a bore 100 mm in diameter and 700 mm long. After exposure, microcirculatory blood flow showed an initial increase for about 5 min followed by a gradual decrease and a return to the control value. It is hypothesised that these changes represent rebound hyperaemia following reduced blood flow during exposure.     

抗角蛋白抗体进入活细胞的共聚焦显微镜观察

目的：观察抗角蛋白抗体能否进入活细胞。方法：以鼠单克隆抗体（mAb)IgG作用于培养中的人Tca8113细胞，以黑素瘤细胞和抗HBsAg抗体作用的Tca细胞作为阴性对照。细胞固定后与FITC标记的羊抗鼠IgG结合，用荧光显微镜及共聚焦显微镜，观察细胞的荧光着色。结果：抗角蛋白mAb作用的Tca8133细胞胞浆呈亮绿色，着色较均匀，细胞核未见着色。两种对照均未见着色。结论：抗角蛋白mAb可进入活细胞，并结合于胞浆成分。     

免疫电镜技术运用过程中电子染色方法的选用

目的探究免疫电镜不同染色方法对免疫组化阳性实验结果影响的关系。方法组织切片经常规免疫组化（雌激素受体GPR30）并行DAB-硫酸镍铵显色后进行电镜切片，然后分为双氧铀-柠檬酸铅双染、双氧铀单染与未染色3组，以便对不同电子染色结果进行比较。结果GPR30免疫阳性产物位于细胞核外的膜性结构上，在铀-铅双染组显示很高的电子密度，但是背景染色也很深，在铀单染组的反差比较好，而未染色组的反差更好。结论免疫电镜技术中针对不同的免疫阳性反应选用不同的电子染色方法，有利于阳性结果的判断与鉴别。     

A study of the prevalence of rotavirus infection in children with gastroenteritis admitted to an infectious diseases hospital.

In a 12 month survey of infants and children with gastroenteritis admitted to Fairfield Hospital, Melbourne, rotavirus was found in approximately 42% of patients. This virus was detected more often during the winter months, particularly in children aged between 12 months and 3 years. Detection of rotavirus by electron microscopy was found to be more sensitive than by counterimmunoelectrophoresis. Routine bacterial and viral studies revealed that bacterial pathogens and common enteric viruses were associated with relatively few cases of gastroenteritis. There is little doubt that rotavirus is the most important aetiological agent of acute gastroenteritis in yvirus is the most important aetiological agent of acute gastroenteritis in young children in Melbourne.     

Fine structure and mineral components of fibrous stonelike masses obtained from the human mesenteries

We investigated the fine structure and mineral components of 29 stonelike masses obtained from the mesenteries of four adult cadavers, using optical microscopy, backscattered electron imaging, scanning electron microscopy, energy-dispersive X-ray microanalysis, and X-ray diffraction. Although the overall appearance of the stonelike masses measuring about 5–20mm in diameter and 0.06–3.1g in dry weight was roughly grouped into smooth bulb- and uneven bulk-shaped types, all the calcified masses basically consisted of core and mantle regions. The smooth bulb-shaped masses had a broad mantle with many concentric rings, whereas the uneven bulk-shaped masses contained a large core. In their core regions, spherulitic and short bundle-shaped deposits composed of needle-shaped apatite crystals were mainly found among loose collagen fibers. Their mantle regions, on the other hand, showed the concentric structures of dense collagen fibers in the intra- and/or extrafibrous calcification with fine sandy grain-shaped deposits. The mineral elements were mainly Ca and P, and the major crystals were hydroxyapatite. Hexahedral whitlockite containing Mg was a minor component. The fiber-rich mantle regions showed lower calcification and lesser crystallization than the fiber-poor core region. When necrotic or some tumor adipose tissues and necrotic lymphoid tissues that might have been caused by some digestive diseases are recognized as foreign matter, their tissues occasionally will be calcified and grow into stonelike masses. These stonelike masses tend to occur more often in women than in men.     

Small epithelial cells in human liver cirrhosis exhibit features of hepatic stem-like cells: immunohistochemical,electron microscopic and immunoelectron microscopic findings

AIMS: To investigate whether cells with features similar to those of the oval cells of rodents and the small epithelial cells (SEC) recently described in certain human liver diseases, i.e. hepatic progenitor cells, also occur in human liver cirrhosis. METHODS AND RESULTS: Surgical specimens from 35 cases of hepatitis B virus-positive cirrhosis (30 cases containing hepatocellular carcinoma) were investigated by immunohistochemical staining for cytokeratin 7 and albumin. Electron microscopic investigations, and immunoelectron microscopic investigations using the same antibodies and a double-labelling technique were performed in 15 and seven cases, respectively. SEC were observed in proliferated bile ductules, at the margins of regenerating nodules and in the fibrous septa in all cases of cirrhosis. The SEC were morphologically similar to the SEC described previously, and to the oval cells seen in experimental hepatocarcinogenesis. They were characterized by their small size, oval shape, scanty electron-dense or electron-lucent cytoplasm, a high nucleo-cytoplasmic ratio, tonofilaments and intercellular junctions. Immunoelectron microscopy revealed that the SEC co-expressed cytokeratin 7 and albumin. Both relatively undifferentiated SEC and SEC with morphological and immunophenotypical signs of differentiation towards biliary epithelial cells and hepatocytes were found. CONCLUSIONS: SEC that exhibit morphological and immunophenotypical features of the SEC seen in certain other liver diseases are found in cirrhosis. These findings further support the hypothesis that a bipotent hepatic stem cell that may give rise to biliary epithelial cells and hepatocytes exists in the human liver.