Pancreatic damage produced by injecting excess lysine in rats

Intraperitoneal injection of lysine (400 mg/100 g body weight) in rats caused necrosis of pancreatic acinar cells with fat necrosis and a significant increase in serum amylase and lipase. The early morphological changes in the pancreas were investigated. At 3 to 6 h, marked swelling of mitochondria was observed throughout the cytoplasm followed later by dilation of the endoplasmic reticulum and the formation of autophagic vacuoles, indicative of rapid cellular degeneration. These results suggest that transient disturbance of energy formation following mitochondrial swelling resulted in disorders of protein metabolism, with disorganization of the endoplasmic reticulum and pyknosis of the nuclei as later events.     

Plasmodium falciparum signal sequences: simply sequences or special signals?

The malaria parasite, Plasmodium falciparum, synthesises and exports several proteins inducing morphological and biochemical modifications of erythrocytes during the erythrocytic cycle. The protein trafficking machinery of the parasite is similar to that of other eukaryotic cells in several ways. However, some unusual features are also observed. The secretion of various polypeptides was inhibited when P. falciparum-infected erythrocytes were incubated with Brefeldin A. Immunoelectron microscopy studies revealed substantial morphological changes in the endoplasmic reticulum following exposure of parasitised erythrocytes to the drug. Immunofluorescence studies of Brefeldin A-treated parasites suggest that polypeptide sorting to different intracellular destinations begins at the endoplasmic reticulum. The parasite also secretes polypeptides by a Brefeldin A-insensitive route that bypasses the classical endoplasmic reticulum-Golgi complex pathway.     

Changes in the structure of Nippostrongylus brasiliensis intestinal cells during development from the free-living to the parasitic stages

The ultrastructure of Nippostrongylus brasiliensis intestinal cells was examined in free-living, feeding second-stage larvae, infective, nonfeeding third-stage larvae, and parasitic, feeding third-stage larvae. The intestinal cells of second-stage larvae were characterized by a well-developed microvillar border, large numbers of ribosomes, Golgi complexes, rough endoplasmic reticulum, and nuclei with prominent nucleoli. The intestinal cells of infective, third-stage larvae had very few microvilli and the cells were extremely narrow. Few ribosomes, Golgi complexes, and little rough endoplasmic reticulum were present. Nuclei did not contain nucleoli. When worms were introduced into an in vitro culture system, development of intestinal cells began. By 36 hr, microvilli were well differentiated and the cysoplasm contained numerous ribosomes and Golgi complexes, and rough endoplasmic reticulum, mitochondria, and nucleoli were prominent. These morphological changes were related to changes in the physiology of Nippostrongylus brasiliensis which occur during development from a free-living to parasitic form.     

Subcellular calcium and magnesium mobilization in rat liver stimulated in vivo with vasopressin and glucagon

The total Ca2+ content of the endoplasmic reticulum and the total Ca2+ and Mg2+ content of mitochondria were determined by electron probe microanalysis of rat liver rapidly frozen in vivo following brief (5-15 s) stimulation with vasopressin or prolonged (10-12 min) stimulation with vasopressin + glucagon. Brief vasopressin injection into the anterior mesenteric vein released 1.8 +/- 0.3 (S.D.) mmol of Ca2+/kg dry weight, from the rough endoplasmic reticulum (p less than 0.01), reducing Ca2+ content of the endoplasmic reticulum from 4.4 +/- 0.2 (S.E.) (controls) to 2.6 +/- 0.2 mmol of Ca2+/kg dry weight. Following vasopressin injection, endoplasmic reticulum Ca2+ was also significantly (p less than 0.025) lower than that in brief sham injected animals (3.5 +/- 0.2 mmol/kg dry weight). Mitochondrial Ca2+ was between 1.0 and 2.3 (+/-0.2) mmol/kg dry weight of mitochondrion, under all conditions studied, and no significant differences were observed. Both hormonal and brief sham injection into the anterior mesenteric vein increased mitochondrial Mg2+ from 42 (+/-0.8) to 49 (+/-1.8) mmol/kg dry weight (p less than 0.05). Hormonal stimulation of Mg2+ uptake was further confirmed by injection of vasopressin + glucagon into the jugular vein (to avoid any stimulation of the liver by the anterior mesenteric vein injection itself); mitochondrial Mg2+ increased from 43 (+/-0.9) (10-min sham) to 57 (+/-1.3) mmol/kg dry weight, with 10-min vasopressin + glucagon injection (p less than 0.01). These results demonstrate that hormones can release Ca2+ from the endoplasmic reticulum and modulate mitochondrial Mg2+ content in vivo without causing detectable changes in mitochondrial Ca2+.     

Reduction capacity of untreated and of repeatedly isoproterenol treated parotid gland of the mouse

Osmium impregnation was used to show possible differences of reduction capacity of perinuclear space, rough endoplasmic reticulum and the Golgi apparatus of unstimulated mouse parotid gland and in the gland after repeated pharmacological doses of isoproterenol. There were some significant differences between the staining of acinar and duct cells. In all intercalated and striated duct cells the staining is dense in the perinuclear space and in the rough endoplasmic reticulum. Osmiophility was not detected in the Golgi complex of intercalated duct cells. The staining was also lacking in the perinuclear space and endoplasmic reticulum of the acinar cells. The cis face of the Golgi complex and numerous transitional vesicles in the acinar cells showed variability of the reduction capacity of their membrane segments. In chronically treated acinar cells Os black was lacking in the Golgi cisternae, except that the numerous transitional vesicles were heavily stained. These results reveal characteristic differences of reduction capacity of endomembrane compartments in different parotid glandular cells, as well as between untreated and treated acinar cells.     

Pancreatic damage induced by injecting a large dose of arginine

Male Wistar rats were injected intraperitoneally with a large dose of arginine (500 mg/100 g body weight) and were sacrificed 24, 48 and 72 h later. Pancreatic tissue was examined by electron microscopy to study the resulting process of degeneration. Degeneration started with disorganization of the rough endoplasmic reticulum into whorls with a concomitant decrease in the numbers of zymogen granules. The main changes in acinar cells after 24 h were partial distension of the endoplasmic reticulum, whorls of agranular membranes encircling zymogen granules and perinuclear vacuoles. At this time large sequestered areas in the cytoplasm contained disarranged rough endoplasmic reticulum and degraded zymogen granules. The mitochondria showed only slight changes. After 48 h, dissociation and necrosis of acinar cells were noted. Subsequently, the necrotic cells were replaced by interstitial tissue composed of leucocytes and fibroblasts. It was concluded that a large dose of arginine is toxic to the rat pancreas when injected intraperitoneally. The early morphological changes of the acinar cells may be related to metabolic alterations associated with the endoplasmic reticulum. The disorganization of the endoplasmic reticulum and the reduced number of zymogen granules may indicate disturbance of protein synthesis. The focal sequestration and degradation of the cytoplasm seemed to represent changes of the acinar cells associated with removal of damaged organelles.     

蒙药新Ⅱ号对扩张型心肌病大鼠心功能、心肌细胞内质网应激及凋亡的作用

目的：探讨蒙药新Ⅱ号对阿霉素性扩张型心肌病大鼠心功能、心肌病理、心肌细胞内质网应激及凋亡作用。方法：30 只SD 雄性大鼠分为3组（n=10）： 正常对照组、模型组（腹腔注射阿霉素2 mg/kg体重,1次/周,用药4周后观察4周）和蒙药新Ⅱ号组（腹腔注射阿霉素2 mg/kg体重,1次/周,用药4周后,给予蒙药新Ⅱ号30 mg/kg·d灌胃,共4周）。观察大鼠的一般情况,8 周后行高频超声心动图检查,测定各项心功能指标,检查结束后处死大鼠,取心脏,行HE 染色、VG染色及电镜观察。用TUNEL 法检测各组大鼠心肌细胞凋亡, Western blot 检测各组大鼠心肌细胞内质网伴侣蛋白GRP78、GRP94及促凋亡因子CHOP、caspase-3表达。结果：与模型组相比较,①蒙药新Ⅱ号组大鼠心脏左室结构及血流动力学各项指标均有明显改善（P<0.05）。②蒙药新Ⅱ号组大鼠心肌病理积分明显下降（P<0.01）。③蒙药新Ⅱ号组大鼠心肌细胞内质网应激伴侣蛋白GRP78、GRP94、CHOP、caspase-3表达及细胞凋亡明显减少 （P<0.01）。结论：蒙药新Ⅱ号可改善扩张型心肌病大鼠心脏功能,减轻心肌病理变化,缓解内质网应激,减少心肌细胞凋亡。这一实验结果可能是蒙药新Ⅱ号治疗扩张型心肌病作用机制之一。     

Qualitative and quantitative observations on the structure of the Schwann cells in myelinated fibres

Various morphological features of the Schwann cells of myelinated fibres in the lizard thoracic spinal roots were studied, and, when possible, quantified using morphometric methods. About 0.8% of the Schwann cells are binucleate and some display clusters of microvilli along the internodes. The percentages of the cytoplasmic area of the Schwann cell occupied by the following cytoplasmic components were determined: mitochondria, Golgi apparatus, granular endoplasmic reticulum, smooth endoplasmic reticulum, multivesicular bodies, dense bodies, autophagic vacuoles, peroxisome-like bodies, lipofuscin granules and lipid droplets. Linear relationships were found between the sectional areas of the mitochondria and granular endoplasmic reticulum of the Schwann cell and both the length of the profile of the Schwann cell plasma membrane and the size of the related axon. The results obtained are compatible both with the hypothesis that the mitochondria and granular endoplasmic reticulum of the Schwann cell are involved in the production and storage of proteins for the plasma membrane of this cell, and with the hypothesis that these organelles are involved in the production and storage of protein metabolites which are subsequently transferred to the related axons.     

Postnatal differentiation of the Golgi apparatus and the dendrites of Purkinje cells of the rat cerebellum

Summary The morphology of postnatal differentiation of the Golgi apparatus, the nucleus, the perikaryon, and the dendrites was studied in Purkinje cells of the rat cerebellum for 30 days after birth using histochemical, histological, and electron microscopic methods.The Golgi apparatus during differentiation undergoes morphological and positional changes. From the 1st to 7th postnatal day, the Golgi apparatus is found in a supranuclear position, and is connected with the axes of differentiating primary dendrites by beam-like processes. From days 8 to 11 this connection disappears, and most of the Golgi apparatus assumes a lateronuclear and infranuclear position. After the 11th or 12th day, the Golgi apparatus is found in perinuclear and peripheral cytoplasmic positions. The formation of granular endoplasmic reticulum occurs in the vicinity of the perinuclear Golgi apparatus. The differentiation of cell and nuclear forms requires approximately 20 days. The morphological changes of differentiation are discussed in relation to the participation of the Golgi apparatus in the differentiation of dendrites and in the formation of the granular endoplasmic reticulum.     

Reduction capacity of untreated and of repeatedly isoproterenol treated parotid gland of the mouse

Summary Osmium impregnation was used to show possible differences of reduction capacity of perinuclear space, rough endoplasmic reticulum and the Golgi apparatus of unstimulated mouse parotid gland and in the gland after repeated pharmacological doses of isoproterenol. There were some significant differences between the staining of acinar and duct cells. In all intercalated and striated duct cells the staining is dense in the perinuclear space and in the rough endoplasmic reticulum. Osmiophility was not detected in the Golgi complex of intercalated duct cells. The staining was also lacking in the perinuclear space and endoplasmic reticulum of the acinar cells. The cis face of the Golgi complex and numerous transitional vesicles in the acinar cells showed variability of the reduction capacity of their membrane segments. In chronically treated acinar cells Os black was lacking in the Golgi cisternae, except that the numerous transitional vesicles were heavily stained.These results reveal characteristic differences of reduction capacity of endomembrane compartments in different parotid glandular cells, as well as between untreated und treated acinar cells.     

The charge distribution in the rough endoplasmic reticulum/golgi complex transitional area investigated by microinjection of charged tracers

The distribution of microinjected ferritin, ranging in charge from anionic to highly cationic, has been used to indicate differences in surface charge on the rough endoplasmic reticulum and the Golgi complex of intact cells. Highly cationic ferritins (HCF)(HCF1, pI 7.9-9.1; HCF2, pI 8.5-9.4; and HCF3.pI 9.5-10.1) were mostly bound and caused swelling of the rough endoplasmic reticulum. Cationic ferritin (CF) (pI 7.0-8.0) and anionic ferritin (AF) (pI 4.0-4.4) caused no changes in morphology. The distribution of these ferritins in the cytoplasmic space varied with their charge. Significantly more CF was bound to surfaces than was found in the free cytoplasmic space. Conversely, there was significantly more AF in the free cytoplasmic space than close to surfaces. Therefore, the intracellular surfaces are negatively charged. Comparison of the structures in the secretory pathway showed no differences in ferritin binding to transition vesicles, rough endoplasmic reticulum, Golgi saccules or secretory vesicles. The Golgi complex beads are not distinguished by their charge. It is therefore unlikely that charge differences play a role in regulating membrane-membrane interactions in this region of the secretory pathway.     

Subcellular fractionation and morphology of calf aortic smooth muscle cells: studies on whole aorta, aortic explants, and subcultures grown under

A comparative biochemical and morphological study was made of calf aortic smooth muscle cells found in situ and grown in vitro under various conditions. Striking alterations in enzyme contents, physical properties, and morphological appearances of lysosomes, endoplasmic reticulum, plasma membranes and, to a lesser extent, mitochondria were observed upon culturing of calf aortic smooth muscle cells. These changes first appeared in cells growing out of tissue explants. They developed further upon subculturing of the cells and depended greatly on the culture conditions used. The alterations included increases in specific activities of some 5- to 25-fold of four acid hydrolases, an average ninefold increase in 5' -nucleotidase, sevenfold increase in cytochrome oxidase, and fourfold increase in neutral α-glucosidase in subcultured smooth muscle cells compared to aortic cells in situ. Cell fractionation studies showed significant shifts in the equilibrium densities of plasma membranes, microsomes, and lysosomes, but not of mitochondria, in smooth muscle cells growing out from explants and in subcultured cells, compared to cells isolated from intact aortas. Although the cells grown in vitro exhibited typical phenotypic features of smooth muscle cells such as abundant myofilaments and surface vesicles, alterations in the morphological appearance of the endoplasmic reticulum, Golgi apparatus, and, especially, lysosomes were observed. These results demonstrate significant differences in specific cellular characteristics and functions of aortic smooth muscle cells grown in vitro compared to aortic cells in situ.     

On the ultrastructure of follicles and isolated follicular granulosa cells of porcine ovary

Summary The endoplasmic reticulum in granulosa cells of primary, secondary, and small tertiary follicles of the porcine ovary is sparse and largely of the granular type.In granulosa cells of large tertiary follicles the endoplasmic reticulum shows distinct signs of proliferation. Some cells even contain whorls of endoplasmic reticulum membranes, essentially of the agranular variety.Direct continuity between endoplasmic reticulum membranes of the granular and agranular type as well as the continuous increase in agranular membranes suggest that these membranes may originate from the granular membranes.Granulosa cells isolated from large tertiary follicles by microdissection and keptin vitro show essentially the same ultrastructure as granulosa cells of intact large tertiary follicles.Some lipid droplets appear to be localized in cavities of the endoplasmic reticulum. It is suggested that the droplets contain precursor material for steroid hormone synthesis.Finally, the development of the agranular endoplasmic reticulum including the appearance of whorls in some granulosa cells of large tertiary follicles indicates that steroid synthesis may occur in such follicular granulosa cells.Read at the Meeting of the Swedish Society for Pathology in Umeå, September 25, 1965 (Bjersing, 1966).This investigation was supported by grants from the Swedish Medical Research Council (Projects No. 13 X-78-01, 12 X-78-02, and 12 X-78-03).     

Morphological changes of the terminations of afferent pathways in a neurodegenerative lesion induced by kainic acid

30 days after kainic acid injection into the rat ventrobasal thalamus, lemniscal afferents were labeled using wheat-germ agglutinin conjugated to HRP. They appeared considerably swollen in the area where neuronal post-synaptic targets had been eliminated. Electron microscopic analysis of the lesioned tissue revealed the presence of large profiles containing numerous organelles, particularly smooth endoplasmic reticulum, and giving rise to thin excrescences filled with neurofilaments. Since these morphological features are typical of regenerating "growth cones", we conclude that afferent terminals deprived of their post-synaptic targets undergo morphological changes preparing them for new synapses.     

Effect of gibberellic Acid and actinomycin d on the formation and distribution of rough endoplasmic reticulum in barley aleurone cells

Analysis of structural changes in barley aleurone cells during germination or following incubation of isolated layers in gibberellic acid with or without actinomycin D revealed extensive development of rough endoplasmic reticulum. Following the assembly of stacked rough endoplasmic reticulum, vesiculation occurred mainly in basal regions of the cell, resulting in a polar distribution of rough endoplasmic reticulum vesicles. It is postulated that these vesicles are involved in protein secretion, because smooth vesicles, derived from the rough endoplasmic reticulum, apparently become appressed to the plasma membrane. The increased α-amylase in the ambient medium and in cell homogenates correlated directly with formation and subsequent vesiculation of the rough endoplasmic reticulum. Furthermore, when cells were treated with actinomycin D and gibberellic acid, α-amylase synthesis was inhibited by 45% and secretion by 63%. These cells were characterized cytologically by large areas of disarrayed segments of fragmented rough endoplasmic reticulum, corresponding to a high intracellular level of α-amylase. In addition, small lipid bodies common to the segmented regions of rough endoplasmic reticulum were surrounded by fine fibrous material, short segments of rough endoplasmic reticulum, and free ribosomes, suggesting that actinomycin D had interfered with development and organization of rough endoplasmic reticulum.     

Prolonged osmification as an indicator of the differentiation process in the endomembrane system.

The technique of prolonged osmification was used in the analysis of reducing capacity of perinuclear space, endoplasmic reticulum and cis-Golgi cisternae in different epithelial cells during embryonic differentiation and immediately after the birth. Cells of the mouse gastric and intestinal epithelium and of the exocrine pancreas and mammary gland were analyzed. It was shown that endomembrane compartments exhibit high variability in their capacity to reduce OsO4 into lower valency oxides. Typical staining of cis-Golgi cisternae by osmium black does not occur before the cells achieve the developmental state in which production of specific products starts. The changes in stainability occurring from the perinuclear space and endoplasmic reticulum towards the cis-Golgi cisternae indicate a maturation pathway with no direct correlation to the chemical characteristic of the substances produced in different cell types. In the mammary gland the reduction capacity of endoplasmic reticulum disappeared with the intensive synthesis of lipids. Considering our previous results and those of other authors, the possible reasons for the observed dynamics in reducibility in particular segments of endomembraneous space are discussed.     

Structure-function relationships during preovulatory development of porcine follicles following equine chorionic gonadotropin stimulation.

Porcine follicular maturation begins by recruitment from a continually proliferating pool of small antral follicles; those receiving the appropriate stimulus differentiate rapidly through a series of structural and functional changes. Such ovarian activity can be induced in prepubertal gilts with a single injection of equine chorionic gonadotropin (eCG). Average follicular diameter in eCG treated females increased from approximately 2 mm before stimulation to 3.5 mm by 24 hr after injection, with subsequent growth to ovulatory size (8 or 9 mm) by 96 hr. Both theca and granulosa layers increased in thickness and complexity, and a prominent capillary bed evolved immediately outside the basement membrane separating the two layers. Cytoplasmic organelles associated with increased metabolic activity and steroidogenesis proliferated within the first 24 hr. Progressive changes included increasing amounts of lipid and rough and smooth endoplasmic reticulum, with the latter occurring in vesicular or lamellar forms and as lipid-associated whorls. Bizarre mitochondrial forms also appeared, often associated with lipids. The amount and proportion of rough and smooth endoplasmic reticulum shifted dramatically as follicles matured. By 24 hr, rough endoplasmic reticulum in thecal cells increased from 4.2 to 7% of cell volume, while the amount in granulosa cells increased from less than 3.5% to more than 10%; the quantity remained relatively constant in the theca but declined to prestimulation values in the granulosa layer. Rough endoplasmic reticulum predominated over smooth in the first 24 hr following stimulation but the proportions were then reversed, so that more than 10% of both layers was composed of smooth endoplasmic reticulum by the time ovulation was imminent. Some follicles had or were in the process of ovulating by 96 hr. Their walls were collapsed into prominent folds with the two cell types beginning to mix. Slight undulations and some regions of discontinuity were observed in basement membranes of large unovulated follicles at this time. In specimens collected at 96 hr poststimulation and processed for retention of lipid, lipid-like material was noticeable in the extracellular matrix surrounding cells that contained organelle configurations suggestive of steroidogenesis.     

猪生殖与呼吸综合征病毒感染Marc-145细胞的病毒形态发生和细胞超微结构观察

以猪生殖与呼吸综合征病毒四川分离株PRRSV-SC1株感染体外培养的Marc-145细胞为模型,通过透射电镜对PRRSV的病毒形态发生学和宿主细胞超微结构的动态变化规律进行研究。结果显示,病毒粒子呈球形,有囊膜,大小约45-65nm,内含直径约25-30nm的核衣壳。病毒感染细胞后以细胞内吞方式进入细胞,在胞浆内复制,装配好的病毒以出芽或细胞外分泌释放到细胞外。感染细胞超微结构变化主要表现为:细胞胞浆空泡增多,内质网扩张,线粒体增生、嵴肿胀、脱落,最后空泡化,细胞表面的微绒毛脱落,出现典型的细胞凋亡特征,并观察到凋亡小体,最后整个细胞裂解、破碎。     

Vesicularization of the endoplasmic reticulum is a fast response to plasma membrane injury

The endoplasmic reticulum of most cell types mainly consists of an extensive network of narrow sheets and tubules. It is well known that an excessive increase of the cytosolic Ca²⁺ concentration induces a slow but extensive swelling of the endoplasmic reticulum into a vesicular morphology. We observed that a similar extensive transition to a vesicular morphology may also occur independently of a change of cytosolic Ca²⁺ and that the change may occur at a time scale of seconds. Exposure of various types of cultured cells to saponin selectively permeabilized the plasma membrane and resulted in a rapid swelling of the endoplasmic reticulum even before a loss of permeability barrier was detectable with a low-molecular mass dye. The structural alteration was reversible provided the exposure to saponin was not too long. Mechanical damage of the plasma membrane resulted in a large-scale transition of the endoplasmic reticulum from a tubular to a vesicular morphology within seconds, also in Ca²⁺-depleted cells. The rapid onset of the phenomenon suggests that it could perform a physiological function. Various mechanisms are discussed whereby endoplasmic reticulum vesicularization could assist in protection against cytosolic Ca²⁺ overload in cellular stress situations like plasma membrane injury.     

Functional and morphological changes induced by tunicamycin in dividing and confluent endothelial cells.

Cultured bovine aortic endothelial cells treated with tunicamycin, an inhibitor of glycoprotein synthesis, developed a concentration-dependent inhibition of N-acetylglucosamine-1-phosphate transferase activity, and this inhibition was correlated with a substantial decrease in [3H]mannose incorporation by the cells. Endothelial cells were very sensitive to tunicamycin, and changes in their morphology occurred as a result of the inhibition of glycoprotein synthesis. The cells became elongated, the surface irregular, roughened, and granular, and there was an increase in the interstitial space between the cells. Electron dense material was accumulated within and dilated the rough endoplasmic reticulum, and the distribution of the glycoproteins laminin and fibronectin throughout the endothelial cell monolayer was modified. These morphological changes coincided with functional impairment with the permeability of endothelial cell monolayers to both 125I-albumin and [3H]inulin being increased by treatment with tunicamycin (10(-6) M) for 24 h. These results indicate that the synthesis of glycoproteins is crucial for cell-cell adhesion and the functional properties of the endothelial lining of blood vessels.