Hepatic effects of Cimicifuga racemosa extract in vivo and in vitro.

Extracts of Cimicifuga racemosa are used frequently for menopausal complaints. Cimicifuga is well tolerated but can occasionally cause liver injury. To assess hepatotoxicity of cimicifuga in more detail, ethanolic C. racemosa extract was administered orally to rats, and liver sections were analyzed by electron microscopy. Tests for cytotoxicity, mitochondrial toxicity and apoptosis/necrosis were performed using HepG2 cells. Mitochondrial toxicity was studied using isolated rat liver mitochondria. Microvesicular steatosis was found in rats treated with > 1,000 mg/kg [DOSAGE ERROR CORRECTED] body weight cimicifuga extract. In vitro, cytotoxicity was apparent at concentrations > or =75 microg/mL, and mitochondrial beta-oxidation was impaired at concentrations > or =10 microg/mL. The mitochondrial membrane potential was decreased at concentrations > or =100 microg/mL, and oxidative phosphorylation was impaired at concenq trations > or =300 microg/mL. The mechanism of cell death was predominantly apoptosis. C. racemosa exerts toxicity in vivo and in vitro, eventually resulting in apoptotic cell death. The results are compatible with idiosyncratic hepatotoxicity as observed in patients treated with cimicifuga extracts.

Defective enamel ultrastructure in diabetic rodents.

We investigated six different types of diabetic rodents. Four expressed a genetic obesity resulting in diabetes. One developed diabetes induced by a diet-dependent obesity, and one with genetic diabetes received anti-diabetic medication. The tooth samples were examined under a scanning electron microscope and with an energy dispersive microanalysis (EDX). The electron micrographs showed severe, varying degrees of damage within the six different diabetic animal types, such as irregular crystallite deposition and prism perforations in genetically obese animals compared to less-disordered prism structures in diet-dependent obesity. Anti-diabetic medication resulted in normal enamel ultrastructure. The EDX analysis revealed a reduction in the amount of calcium and phosphorus in all regions affected by diabetes. Based on these animal studies, we suggest that both juvenile diabetes type I (in infants) and adult diabetes type II (in pregnant mothers, affecting the developing foetus) may affect the normal development of teeth in humans.

Estrous stage determination in rats by means of scanning electron microscopic images of uterine surface epithelium.

With the results presented in this paper we devised an alternative method to precisely date the rat endometrium in relation to the estrous cycle. This is done by the exclusive use of scanning electron microscopy. Owing to its short estrous cycle (4 to 6 days, depending on age), the rat is ideally suited for the examination of cyclic changes occurring in the uterine epithelium. The cycle stage of rats predated by vaginal smear cytology was verified through the measurement of hormones relevant to the estrous cycle, i.e. estradiol-17-beta (E2), progesterone (P), luteinizing hormone (LH), and follicle stimulating hormone (FSH). Based on scanning electron images surface changes could thus be correlated to the cyclic variations of blood levels of sex hormones. The appearance of pseudoglands, the most prominent aspect during the cycle, is correlated with decreasing estrogen and rising progesterone levels. Pseudoglands are formed by apoptosis and necrosis of epithelial cells, and are most numerous during estrus. They had in previous studies been classified as genuine uterine glands.

Alkaline phosphatase distribution in rat endometrial epithelium during early pregnancy: a scanning electron-microscopic study.

The participation of apical membranes of uterine epithelial cells in the process of blastocyst adhesion makes them an interesting object in the study of changes occurring during early pregnancy. In the study of these changes alkaline phosphatase (AIP), a typical brush border enzyme, was chosen for demonstration with the scanning electron microscope (SEM) by means of a backscatter detector. Thus the temporal and spatial pattern of enzyme activity on the uterine luminal surface was made visible with lead salt procedures. AIP activity was shown to be located on apical membranes and microvilli of endometrial epithelial cells with high activity on day 2 of pregnancy decreasing to virtually no activity on day 5. This decrease in overall AIP activity was shown to be asymmetrical with respect to the uterine cavity. It begins on the antimesometrial half of the uterine lining on day 2. A distribution pattern demarcating a presumptive implantation site along the uterine horn was not found. However, on day 5 of pregnancy, a characteristic pattern of surface folds was found, dividing the uterine horn into 'implantation segments'. In addition, SEM investigation revealed a marked variation of AIP activity from one individual cell to the next on day 2 of pregnancy resulting in a mosaic-like pattern. This pattern is lost with the decrease of AIP activity on day 5. Thus heterogeneity of uterine epithelial cells in AIP activity is apparently a feature of nonreceptive epithelium in contrast to the homogeneous epithelium on day 5. It is proposed that epithelial cell homogeneity could be a marker for uterine receptivity.

Ultrastructure as a basis for dating of rat endometrium.

Studies on the endometrial cycle depend upon the precise dating of the cycle stage. In the present paper the generally applied method of vaginal smear was carefully verified through the measurement of the hormones relevant to the endometrial cycle. From rats predated through vaginal smear cytology, the hormone levels of estradiol-17-beta (E2) and progesterone (P), luteinizing hormone (LH), and follicle stimulating hormone (FSH) were measured. The values obtained were then compared to the standardized values of our reference curve. Animals with values that did not fit within the standard deviation of our reference curve were excluded from this investigation. Thus, for the first time exactly dated rat endometrial morphology was studied with electron microscopy. The morphology of the surface epithelium of rat endometrium from all four stages of the cycle is described in detail. In addition a semiquantitative morphometric analysis of the following parameters was performed: cell volume, nuclear volume, the volume density of secretory granules, digestive vacuoles, mitochondria, Golgi apparatus, rough endoplasmic reticulum, and lipid vacuoles as well as the size of lipid vacuoles. With the cellular content of lipid vacuoles and their diameter, it is possible to differentiate between proestrus/estrus and diestrus I/diestrus II, the latter possessing definitely more and larger lipid vacuoles. During estrus the greatest cytoplasmic volume develops. In addition to this, secretory granules are only present during estrus. Finally, diestrus I can be well differentiated from diestrus II, because diestrus I exhibits more digestive vacuoles and during diestrus II a high percentage of free ribosomes is present. On the basis of distinct morphological features, described in this paper, it is now clearly possible to distinguish between the four different cycle stages.

Pseudo-decidualization at the site of implantation in tubal pregnancy.

The tissue from three tubal pregnancies at the site of implantation has been investigated with light and electron microscopy. Large seemingly "decidual" cells were found to be present between the anchoring villi and the tissue of the uterine tubes. These cells are or cytotrophoblastic column origin. Through discontinuities of the syncytiotrophoblast they migrate into the underlying tissue were they grow and synthesize large amounts of glycogen. It is suggested that these cells be named pseudo-decidual cells in order to distinguish them clearly from genuine decidual cells to which they only bear a resemblance on the light microscopical level. The fibrinoid present at the utero-placental interface in ectopic implantation cannot be found in ectopic implantation. From this fact it is deduced that fibrinoid is formed through the action of genuine decidual cells and therefore must be of maternal origin.

The functional morphology of the human endometrium and decidua.

The functional morphology of the human endometrium and decidua has been investigated with particular attention to all aspects of implantation and contraception. The postovulatory triad (of glandular epithelium), comprising subnuclear glycogen, giant mitochondria, and the NCS seems to be involved in both implantation and contraception and has been studied in detail. Initial stages of glycogen formation were found to be related to straight, uncoiled forms of polysomes which, by acting as acceptor molecules for glucose start glycogen synthesis. Discharge of glycogen from the glandular epithelium is achieved by a type of apocrine secretion. Computer-aided reconstruction of giant mitochondria, based on ultra-thin serial sections, revealed that giant mitochondria of the secretory phase of the cycle are not comprised of a single complex mitochondrion, but of numerous single mitochondria. Towards the end of the cycle, these mitochondria either reduce their size to normal by means of budding of protrusions or they are degraded in autophagic vacuoles. The NCS was studied for the first time from the initial stages of formation to the final stages of intracellular degradation. Based on ultra-thin serial sections the NCS was reconstructed with manual and computer-aided reconstruction. It was found to consist of seven sets of tubules, each containing three rows of coiled tubules which are wound in one-and-a-half turns from the site of origin at the nuclear membrane to the karyoplasmic pole. The individual tubules are interconnected by means of lacunae or connecting tubules. The formation of the NCS is suppressed by progesterone IUDs, but not by copper IUDs, and it is not formed in anovulatory cycles or under low-dose gestagen therapy (Minipill). The NCS is believed to participate in regulatory mechanisms of the glandular epithelium. Thus, the contraceptive action of the progesteroneIUD may in part be due to the suppression of the NCS and not only due to decidualization of the endometrial stroma. From the middle to late secretory phase K cells (endometrial granulocytes) and predecidual cells are formed. Their development and ultimate fate was studied during the normal physiological cycle, under a progesterone IUD, and also during pregnancy. K cells do not develop from stromal fibrocytes, as has been postulated in the past, but from lymphocyte-like cells which are present in the endometrium after ovulation. These cells multiply through mitotic division within the endometrial stroma. Under a progesterone IUD and during pregnancy they discharge their secretory granules and reduce their size drastically by giving off glycogen-containing cytoplasmic patches.(ABSTRACT TRUNCATED AT 400 WORDS)

Liver architecture.

The development of liver parenchyma starts from entodermal cells which grow out from the gut into the mesenchyma of the septum transversum. In the definitive organ this close association of epithelial cells (hepatocytes) and mesenchyma-derived nonparenchymal cells is maintained. The liver, and with it each hepatocyte, acts in two directions: the vascular poles of the hepatocytes serve in an ingestive sense, while at their biliary poles secretory functions are exerted. Hepatic microvascularization comprises two afferent vessels (arterial and portal terminal branches), the sinusoids and the terminal hepatic venule. Sinusoidal cells surround the capillaries but also have highly specialized functions with regard to filtration, phagocytosis, fat storage and defense. The autonomic innervation plays an important role in the regulation of metabolic functions. Above the cellular level the proper architecture of the liver parenchyma has been the object of controversial discussions for centuries. The concept of the liver lobule, the portal unit, the liver acinus and other structures are presented and discussed. Finally, the liver parenchyma is described as an irregular interdigitating system of regions related to the terminal blood vessels.

Microarchitecture of the human endometrium by scanning electron microscopy: menstrual desquamation and remodeling.

Twenty-two hysterectomy specimens were collected over a period of two decades in order to investigate the morphological sequence of menstrual desquamation and its consecutive remodeling of the endometrium. The technique used was described earlier by H. Ludwig and H. Metzger (1976). Scanning electron microscopy is the only way to illustrate and describe the microarchitecture of the endometrial surface. At the beginning of the menstrual bleeding, glandular stumps surviving the desquamation of the layer functionalis stick out from the debris at the top of the basal layer. Fibrin mesh formation, the liberation of lysosomes, and the emigration of white blood cells and macrophages, both being already present in the midluteal endometrial stroma, can be observed. They are interrelated with the clearance of the menstrual wound. Coincidentally with the process of desquamation the re-epithelization starts and takes the first four to six days of the normal cycle. The events are illustrated by selecting specimens of uteri from women with comparable data but from different days of the normal cycle. Surprisingly transitory excess formation of epithelial outgrows (micropolyps) are observed. They disappear later in the cycle, some might persist and form micropolyps, which will be the source of occasional intermenstrual bleeding--so far the polyps are vascularized. The endometrial surface is covered de novo by a lining surface epithelium at the sixth day. Ciliogenesis occurs within this epithelium. Other ciliated cells emanate from the glandular epithelium. In early stages of menstrual regeneration the growth pattern of the epithelial monolayer forming the lining surface in spiral traces according to their origination from the circle-structures of the endometrial glands. Before the incoming menstrual break-down small crevices, clefts or defects appear within the lining surface endometrium, a few white blood cells, enriched in the stroma around the vessels, might even reach the surface. The apical membranes of several non-ciliated cells exhibit rounded leaks, others show ruptures. It is the tissue break-down around the superficial endometrial vessels, what creates the onset of menstrual blood flow. In the very early preparations of the bleeding endometrium those opened capillary vessels could be identified.

Scanning microscopy of the developing vagina in postnatal gerbils.

Scanning electron microscopy of postnatally developing gerbil vagina (birth to maturity) shows that longitudinal folds form prior to transverse folds; the process of fold formation is initiated on the dorsal wall and proceeds ventrally. From days 1 to 7 postnatally, the vaginal epithelium is composed of either flat or bulging cells, depending on the vaginal region. The luminal cell surface is covered with uniform stubby microvilli and solitary cilia. Between days 9 and 20, the flat cells with distinct cell boundaries spread toward more proximal areas, leading to the formation of mixed patches of cells with flat or rounded apices. Individual elongated microvilli or tufts of forked microvilli may sprout from their surfaces. Solitary cilia gradually disappear. The transition from immature to mature vaginal epithelium starts around day 20, when individual cells recess below the level of neighboring cells. This process spreads throughout the vagina during the following days, reflecting local changes in the subsurface layers of the epithelium preparatory to exfoliation. Around day 40 the actual exfoliation of the luminal cell layer starts. By this time the surface characteristics of many of the desquamating cells have changed. In addition to microvilli, microridges are being formed. The process of exfoliation is finished by about day 60. The newly appearing cell layers now transform into typical cornified cells of the cycling vaginal epithelium.

The vaginal surface epithelium during postnatal development in gerbils.

The postnatal development of the gerbil vagina has been investigated and special attention has been paid to the luminal surface epithelium structures. Different types of microvilli and solitary cilia are discussed.

[Scanning electron microscopy of the IUD tail]. / Examen des fils de D.I.U. au microscope electronique a balayage.

PIP: The surface ultrastructure of over 40 IUDs including Multiload, Copper 7, Copper-T, Nova-T, FDI, Lippes Loop, and Dalkon Shields, in place for periods ranging from less than 1 month to over 9 years was studies with the scanning electron microscope to assess the influence of various factors on the etiology of pelvic inflammatory disease (PID). Strings of IUDs in place for less than 1 month showed few changes. 3 sections of strings cooresponding to intravaginal, intracervical, and intrauterine location were easily distinguishable after 3 months of use and were comparable in all types of IUDs regardless of type of string. IUDs with relatively smooth strings became contaminated less rapidly the 1st few months of use, but all strings eventually began to acquire coatings which served as substrates for adhesion of exfoliated epithelial cells, blood cells, uterine and cervical secretions, and other matter. The deposits were usually invaded by bacteria. There appeared to be a correlation between the length of the intravaginal string and the quantity of vaginal epithelial cells, but not of bacteria. The surfaces of intravaginal segments appeared identical regardless of whether the wearer suffered from PID. After 3-6 months of use, the intracervical segments contained large deposits with a globular structure. The deposits, unlike those in the other 2 sections, were very hard and appeared to contain little water. The intracervical sections appeared to contain age rings resulting apparently from variations in the composition of materials deposited, although they were largely composed of cellular debris, mucus, and occasionally trapped bacterial residues. No calcium was seen under X-ray, but copper was usually observed in IUD strings in place at least 9 months. The thickness of the intracervical deposits depended more on such factors as the quantity and composition of cervical and vaginal secretions than on the duration of use of the IUD. Bacteria were found in most intracervical string sections, but no relationship was established between presence of bacteria and PID. The intrauterine string segment was usually more homogeneous and contained more water than either of the other sections. Bacteria were sometimes observed, particularly in devices in place for more than 2 years, although they were not necessarily associated with signs of inflammation. Coatings of material seemed to form more rapidly on strings with 2 strands than on those with just 1 strand, and on strings with irregular surfaces or knots. The thicker the coating of materials on the intracervical and intravaginal segments, the greater the likelihood of bacteria on the intrauterine segments. Since the thickness of the coating increased with time, it does not seem advisable to leave an IUD in place for longer than about 18 months.^ieng

Scanning electron microscopy of the IUD tail.

PIP: The surface ultrastructure of the tails of various IUDs (Multiload, Copper 7, Copper T-Device, Nova-T, FDI, Lippes Loop, and Dalkon Shield) was studied with the scanning electron microscope. The parts of the IUD tail exposed to the vaginal, cervical, and uterine milieu showed strikingly different surface characteristics. The surface ultrastructure of these 3 parts is influenced by the duration of use, nature of microbiological contaminants, pregnancy with the IUD in situ, and pelvic inflammatory disease (PID). Duration of use and PID alter the surface ultrastructure most drastically. In comparing tails of the different IUDs, the IUDs with relatively smooth tails became less rapidly contaminated during the 1st few months of use. The surface ultrastructure of the IUD tail was formed through a coat of material which consists of cellular debris, dead bacteria, mucus, etc. The thickness of the coat was found to be largely dependent on the duration of IUD use. The thicker the coat, the more likely bacteria were found on the intrauterine part of the tail. The possible significance of these findings with respect to the development of PID is discussed.^ieng

Morphologic alterations in the epithelium of the human oviduct induced by a low dosis gestagen.

Morphological alterations in the epithelium of the human oviduct related to continuous administration of low dosage lynestrenol were studied in 10 women by electron microscopy. The effect of lynestrenol on the morphology of the tubal epithelium was shown to be time dependent. First the discharge of secretory products from the non-ciliated cells was inhibited. Secondary to this the synthetic activities of the cells were drastically decreased. This inactivation of the tubal epithelium reached its maximum after 6 months. At this time the epithelial cells, particularly the non-ciliated cells were almost devoid of organelles and had an electron transparent cytoplasm. In addition to the action of lynestrenol on the secretory and synthetic activities of the cells a strong ciliogenetic effect could be observed after about 3 months of lynestrenol administration. The ratio of ciliated to non-ciliated cells changed from 1:1 at the beginning of the treatment to 2.9:1 after 6 months. This ratio was then maintained and could still be found in patients who had used lynestrenol for 25 and 39 months. In contrast to the persistent effect of lynestrenol on the ciliary apparatus of the cells the synthetic activities and the secretory mechanism were partially restored with long standing use. The importance of the observed changes and their possible participation in the contraceptive action of lynestrenol are discussed.

[The low dosis gestagen therapy (author's transl)]. / Die Low-Dosis-Gestagen-Therapie

In young women aged 22-34 years the hormone profile under continuous therapy with 0.5 mg lynestrenol/day have been investigated. Estradiol, progesterone and FSH values have been determined in plasma by the RIA or protein-binding method, whereas LH excretion was daily measured in the urine starting on the 5th day of cycle. The low-dose-gestagen therapy shows all possible hormonal patterns ranging from the ovulatory to anovulatory one. The estradiol-17 B values in half of the patients were higher than in the cycle preceding treatment. In the first few treatment cycles all women showed markedly decreased progesterone values. However, this defect seems to recover and return to normal in later cycles. In order to be able to further explain the alterations occurring during the luteal phase under the treatment with 0.5 mg/day of lynestrenol, ovarian tissues wedge resection of women undergoing gynecological surgery has been examined macroscopically and microscopically. At the same time the hormonal profile has been determined in plasma. These investigations revealed that the morphological alterations in the ovary proceed in an unchanged way under the mentioned treatment. Histologically it could be demonstrated that ovulation is delayed by several days. The histological findings of tubal tissue (from patients who underwent sterilization) showed that the secretional activity of the tubal epithelium is considerably decreased and ciliogenesis increased. In our clinical study neither tablet nor patient failures occurred. With the exception of cycle- and tempo-disturbances no severe adverse reactions were reported. Breakthrough bleedings and spotting as well as amenorrhea in most cases manifested already in the first treatment cycles. It is important that the daily tablet is taken at midday because the maximum effect on the cervical mucus of the mini-pill therapy is most pronounced 4-6 hours after tablet intake. Especially in young patients it is recommended to get written reports on tablet intake. The strict following of these directions mainly guarantees a successful mini-pill therapy.

PIP: Hormone levels and uterine and ovarian cytology were studied in women treated with .5 mg lynestrenol/day. Plasma estradiol, progesterone, and follicle stimulating hormone were determined, and urinary luteinizing hormone excretion was measured daily starting on the 5th day of the cycle. All possible hormonal patterns, from ovulatory to anovulatory, were observed during treatment. In half of the patients estradiol-17beta levels were higher than in the cycle preceding treatment. Progesterone levels decreased markedly in the 1st few treatment cycles, but then returned to normal in later cycles. Ovarian morphology was not altered by treatment, but ovulation was delayed by several days. It is recommended that the ''minipill'' be taken at midday, since its effect on cervical mucus is most pronounced 4-6 hours later.

[Experiences with the modern intrauterine devices Cu-7 and Cu-T (author's transl)]. / Erfahrungen mit modernen "Intrauterine Contraceptive devices" Cu-7 und Cu-T

PIP: A study comparing the Copper 7 (CU7) and Copper-T (CuT) IUDs is presented. Cu7s were inserted in 300 women and CuT devices in 100. The pregnancy rate was 1.4 for the Cu7 and 0 for the CuT. The continuation rate for the Cu7 was 70% and 86% for the CuT. The higher discontinuation rate for the Cu7 was due to a higher rate of expulsion among nulliparae, of which there were none in the CuT group, and a higher incidence of removals due to personal reasons, e.g. desire to have children. Cramps and bleeding irregularities were the most frequent side effects, occurring in 10%-40% of the material. Electron microscopic studies showed that the strength of the copper IUD did not decrease with the length of use. Copper is the most effective IUD metal. Copper IUDs have been found to cause increased fibrinolytic and leukocytic reactions. Insertions are best performed on the 3rd or 4th days after menstruation. The greater the surface area of an IUD, the lower the expulsion rate and the higher effectiveness and rate of side effects it has. The exact mechanisms of IUD effectiveness are not known, but it has been shown that the presence of an IUD affects nearly all phases of ovulation, nidation, and fertilization.^ieng

Studies on the liver of Xenopus laevis. I. The ultrastructure of the parenchymal cell.

The liver of Xenopus laevis was examined with electron microscopy. Its structure was found to be markedly different from that of mammals, particularly regarding the morphology of the hepatocytes to be classified as typical. It was established that the main function of such a cell is the storage of glycogen, and further that it possesses only scant organelles and other inclusions. Since this type of cell was found most frequently in the liver of untreated animals, it was designated as normal cell. The fact appears noteworthy that in the normal liver of Xenopus laevis an abundance of cell types occur which are otherwise found to be proliferated under experimental conditions, e.g. cells with pronouncedly augmented RER, enlarged Golgi complexes, increased lipid inclusions etc. This high number of divergent hepatocytes and the fact that all intermediate stages between the individual extremes are present and not to be accounted for by the position of the cell within the liver was interpreted as being the expression of a cyclic passage of the various stages of activity. It is of special interest that augmented degradation of glycogen in the liver cell takes place only during vitellogenesis. Acute and chronic hunger, as well as adaptation to cold, hardly affect the morphology of the normal cell, especially as far as the glycogen is concerned. The possible causes for this are discussed.