[Alcohol in pregnancy and fetal heart damage]. / Alkohol in der Schwangerschaft und kindliche Herzschäden.

Alcohol abuse and addiction during pregnancy can induce damage of the heart muscle and heart defects in the offspring. Between 1973 and 1991 216 children with fetal alcohol syndrome were diagnosed and examined for congenital heart defects. In 63 children (= 29,1%) heart defects were present, established mainly by heart catheterization and/or echocardiography. Most frequently ventricular septal defects (24/63) and atrial septal defects (23/63) occurred, less frequent were AV-defects (3), PDA (3), Fallots teralogy (5), and other malformations (5). Biopsy during heart surgery in 3 children revealed histological and electron microscopic changes of alcohol-cardiomyopathy. Heart defects are predominantly found in children with higher degree of severity concerning the clinical syndrome, less frequently in alcohol effects (4/38 = 11%). Alcohol in pregnancy, even in the course of so called social drinking, usual drinking and binge drinking has to be considered as an essential and preventable co-factor within the multifactorial etiology of congenital heart defects.

Phenotyping of macrophages with monoclonal antibodies in endomyocardial biopsies as a new approach to diagnosis of myocarditis.

Cryostat sections of endomyocardial biopsies from 53 patients (mean age 41 +/- 5 years, 38 male and 15 female) clinically indicated to suffer from myocarditis were stained using monoclonal antibodies against subpopulations of T-lymphocytes and macrophages and with polyclonal rabbit-anti-human sera marking two calcium-binding proteins expressed by monocytes and macrophages appearing in inflammatory sites only. No inflammatory infiltrate cells were found in 13 cases (25%). Mononuclear cell infiltrates were present in 40 cases (75%). Ten biopsies showed a predominance of macrophages bearing the marker 27E10, characteristic for an early acute inflammation and 18 biopsies contained 25F9 positive macrophages, characteristic for a late stage of inflammation. An intermediate type of inflammation with both macrophage types present was found in 12 patients. Patients with immunohistologically confirmed myocarditis had atrial, ventricular or combined forms of arrhythmias (78%), scars in the vectorcardiogram (100%) and radiological evidence of cardiomegaly (36%). In conclusion, typing and endomyocardial biopsies for macrophage subpopulations is a sensitive new approach to assess the diagnosis of myocarditis.

Degenerative and regenerative processes in the peripheral nerve after disconnection and reanastomosis using the Neodym Yag laser.

The development of neuroma after peripheral nerve surgery is still an unsolved problem. Such lesions are not yet controllable by conventional methods. Reports of results achieved using various types of laser surgery appear contradictory. In this experimental study on rats, the Neodym Yag laser was found to be superior the conventional methods. After severance and reanastomosis of the sciatic nerve using this type of laser, no development of neuroma was observed in any of the animals during the first three months of observation.

[Congenital alcoholic cardiomyopathy in 3 children]. / Konnatale Alkoholkardiomyopathie bei drei Kindern.

Three cases of congenital alcoholic cardiomyopathy and cardiac defects in children are described. The mothers were heavy spirits drinkers. One child had a ventricular septal defect, two had Fallot's tetralogy, all requiring operation. At open-heart surgery left-ventricular myocardial biopsies were obtained in all three children. Histological and electronmicroscopic examination revealed primary toxic and hypotrophic changes, which differed from those seen in alcoholic cardiomyopathy of the adult. The cells, their nuclei and the myofibrils had reduced diameters. The mitochondria were damaged to differing extent. The myofibrils were arranged in parallel and some had contraction bands and ruptures. The sarcoplasmic reticulum was dilated and had vacuoles. The cell surface penetrated into the interstitial tissue. All these changes could have resulted from inhibited embryofetal cell growth and cytotoxic damage by alcohol during pregnancy. It would seem that alcoholic cardiomyopathy occurs only in extreme forms of alcohol abuse during pregnancy. The prognosis is as yet unknown.

[Fine structure of surviving heart muscle cells in ventricular aneurysms]. / Die Feinstruktur überlebender Herzmuskelzellen im Ventrikelaneurysma.

Surgically-obtained tissue specimens from 41 patients with ventricular aneurysm were studied electron microscopically. The tissue from the resected aneurysms showed substantially varied morphological differences. In some, there were extensive regions of scar containing increased fibrotic material and few cells, in others there were also larger contiguous regions of myocardium with an essentially normal appearance. In the preserved myocardial regions, the cardiac cells showed moderate hypertrophy. There was an increase in contractile substance in parallel with an increase in mitochondria and enlargement of the nucleus with frequent waves and invaginations in the cell membrane. The cells at the marginal regions between fibrous tissue and preserved myocardium were frequently isolated from adjacent cells. In particular, when the isolated cells were completely surrounded by fibrous tissue, clear degeneration was apparent. These cells showed mainly a fibrillolysis with dissolution of the cross-bands and loss of the entire contractile apparatus. In compensation, occasionally there was proliferation of other cell structures, especially the free sarcoplasmatic reticulum. The hypertrophy of the still intact myocardial cells is considered compensatory for the infarct-incurred loss of tissue. The degenerative appearance is mainly attributable to fibrous tissue invasion. The diminished oxygen supply, compromised or abolished impulse conduction, loss of function and passive stretch during systole may be regarded as causes of the degeneration.

Reversibility of long-term effects of GnRH agonist administration on testicular histology and sperm production in the nonhuman primate.

The present investigation evaluates the long-term effects of GnRH agonist treatment on testicular histology, sperm production and the subsequent recovery of these parameters. Four adult rhesus monkeys (M. mulatta) were treated with the GnRH agonist nafarelin (D-Nal(2)6-GnRH), released from i.m.--injected poly-D,L-lactic-co-glycolide microspheres for 20 months. Monthly injection of the GnRH agonist preparation uniformly suppressed serum levels of bioactive LH and testosterone. The size of the testis was reduced to about 30% of pretreatment. Sperm counts were suppressed to azoospermia for a total period of 53 and 77 weeks, respectively, in two monkeys and the other two animals were extremely oligozoospermic. Evaluation of testicular biopsy material after 6, 12 and 20 months of treatment revealed decreased seminiferous tubule diameter, spermatogenic disruption at the level of spermatogonia or spermatocytes, accumulation of lipid droplets and secondary lysosomes in the Sertoli cell cytoplasm, and increased thickness of the tubular wall compared with pretreatment histology. Electron microscopic examination revealed that the increased wall thickness was due to an enlargement of the inner collagen layer. No evidence of fibrosis or calcification could be obtained. Leydig cells were atrophic. Serum hormones, testis size and sperm counts returned to pretreatment values within 5 to 8, 13 to 16, and 18 weeks, respectively, after termination of treatment. Testicular histology, assessed 8 months after cessation of treatment, was indistinguishable from pretreatment. It is concluded that GnRH agonist-containing microspheres are a feasible modality for sustained administration of GnRH agonists and GnRH agonist-induced suppression of pituitary and testicular function is reversible following withdrawal of treatment. Thus, GnRH agonists may have a potential for regulation of male fertility and, presumably, also for treatment of precocious puberty.

Prevention of neuroma formation by Neodym Yag laser--experimental observations.

The influence of Laser radiation on neuroma formation was investigated in rat sciatic nerves. Peripheral nerve trunks were divided by focused Neodym Yag Laser and compared with those transected by microscissors. The nerves were re-exposed at different time intervals up to 240 days after initial operation. True neuroma formation could not be observed after laser transection. Sharp division resulted in widespread amputation neuromas consisting of regenerating axons and connective tissue. Neodym Yag laser radiation by its scattering effect may cause deep coagulation necrosis and thrombosis of vasa nervorum thus sealing the dome of the divided nerve trunk. The formation of amputation neuromas may be suppressed by laser application.

Electron microscopic investigations of the cyclophosphamide-induced lesions of the urinary bladder of the rat and their prevention by mesna.

Fully developed cyclophosphamide-induced cystitis is characterized by nearly complete detachment of the urothelium, severe submucosal edema owing to damage to the microvascular bed and focal muscle necroses. The initial response to the primary attack by the cyclophosphamide metabolites seems to be fragmentation of the luminal membrane. This damages the cellular barrier against the hypertonic urine. Subsequent breaks in the lateral cell membranes of the superficial cells and in all the plasma membranes of the intermediate and basal cells, intercellular and intracellular edema and disintegration of the desmosomes and hemidesmosomes lead to progressive degeneration and detachment of the epithelial cells with exposure and splitting of the basal membrane. The morphological changes of the endothelial cells, which become more pronounced in the later stages of the experiment, the involvement of blood vessels regardless of their diameter and the location-dependent extent of the damage indicate a direct type of damage which is preceded by a mediator-induced increase in permeability, the morphological correlate of which is the formation of gaps in the interendothelial cell connections on the venules. These changes can be effectively prevented by mesna. The only sign of a possible involvement is the increase in the number of specific granules with a presumed lysosomal function in the superficial cells.

Cardiovascular and cerebrovascular effects of isoflurane-induced hypotension in the baboon.

Mean arterial pressure (MAP) decreased by 10, 20, 40, and 50% of its baseline value in six anesthetized baboons with the administration of increasing concentrations of isoflurane. Mean arterial pressure, right atrial pressure, pulmonary artery pressures, and cardiac output were measured at each decrement in arterial pressure, and after the withdrawal of the isoflurane. At the same times, cerebral blood flow, the cerebral metabolic rate for oxygen, and the cerebral metabolic rate for glucose were determined. Cerebrovascular reactivity was assessed before, during, and after the administration of isoflurane, and morphologic evidence of ischemic cell change was sought at the end of the investigation. Isoflurane produced dose-related decreases in systemic vascular resistance and MAP; heart rate, cardiac output, and stroke volume did not change significantly. Although cerebral metabolism was depressed in a dose-related manner, the response of the cerebral blood flow (CBF) was biphasic. At low isoflurane concentrations, CBF decreased significantly and then increased to reach baseline values at the highest concentration. Cerebrovascular reactivity was intact at baseline and at the 20% decrement in MAP; it was impaired at the 50% decrease in MAP and when studied 100 min after the withdrawal of isoflurane. There was no morphologic evidence of ischemic cell damage in any animal.

Binding and degradation of proteoglycans by cultured arterial smooth muscle cells. II. Binding sites of proteoglycans on the cell surface.

Exogenous proteoglycans stained for electron microscopy with colloidal gold and/or cuprolinic blue bind to the surface of cultured arterial smooth muscle cells at two different sites. (I) About 20% of the proteoglycans adsorbed to the cells from the culture medium interact as monomeric and multimeric proteoglycans with smooth or coated membrane areas. (II) The bulk of exogenous proteoglycans exhibits high affinity binding to cell membrane-associated 10 nm fibrils containing or being closely associated with fibronectin and to collagen. It is suggested that the self association of proteoglycans and their binding to the cell membrane and to cell surface-associated fibronectin and collagen are important for maintaining an appropriate micro-environment for the cultured cells.

Characterization of the cytolytic activity of a cloned antigen-specific T suppressor cell derived from a tolerant CBA/J mouse.

The antigen-specific T suppressor cell clone HF1 isolated from a CBA/J mouse made tolerant by low doses of bovine serum albumin has suppressive and cytolytic activity. The analysis of the latter gave the following results. Natural killer (NK)-sensitive YAC-1 (H-2a) and RBL-5 (H-2b) target cells are lysed whereas other NK targets, like EL4 (H-2b) or the human K562 cell line are resistant. Cytolytic activity is not antibody mediated. Its inhibition by sugar phosphate or monoclonal antibodies against LFA-1 antigens is such that HF1 can neither by typed as T killer nor as NK cells. It seems to represent a distinct T lymphocyte type.

The correlation between tissue differentiation and production of mammary tumor virus (MTV) in transplanted murine mammary tumors. Electron microscopic observations.

The spontaneous mammary tumors of the NMRI mouse are well developed microcystic adenocarcinomas. Serial isologous transplantation of the tumors results in nearly complete dedifferentiation to a solid tumor, in which only electron-microscopically rudimentary acinus-like microlumina can be observed. The adenocarcinomas produce A and B particles in abundance, with the A particles appearing intracellularly in the adluminal cytoplasmic regions of the epithelial cells in association with typical cellular structures and the B particles being restricted to closed extracellular compartments such as vacuoles or acini alone. The loss of alveolar organization in the solid tumors is followed by an almost complete reduction in mature B particles, while A particles are still regularly observed and appear to be less reduced in number. This suggests that the production of extracellular B particles is dependent upon the secretory activity of the tumor cells and that in nonsecreting cells it is predominantly a late step in virus release that is inhibited, not the synthesis of intracellular precursors.

Kidney tubule cell injury of male rats after estrogen treatment.

Ultrastructural changes in the tubular epithelium of the rat kidney following a large dose of estrogen (300 micrograms per week for 20 weeks) were studied by means of electron microscopy. The ultrastructural changes in estrogen-treated rats were confined to the proximal tubule. The changes consisted of intracytoplasmic vacuoles and dilatation of the intracellular space between adjacent tubule cells. The cisternae of the endoplasmic reticulum exhibited some degree of vesiculation characterized by club-like formations. In addition, large masses of collagen could be observed within the peritubular capillaries. In contrast to the controls the tubular cells of estrogen-treated rats exhibited large areas or granules with dense inclusions and membranous, filamentous material in a process of being transformed into an autolysosome and residual body. The results are discussed in the light of those of other authors.

Ultrastructural study of cholestasis induced by longterm treatment with estradiol valerate. I. Tight junctional analysis and tracer experiments.

Over a period of 20 weeks estradiol valerate (1.5 mg/kg body weight/week) was administered subcutaneously to male Wistar rats from which the livers were examined at four week intervals employing a freeze-fracture technique and colloidal lanthanum tracer studies. In connection with intrahepatic cholestasis, distinct alterations in the tight junctions were observed, consisting of disorganization, rarification and proliferation. Disruption of the tight junctions was not seen and colloidal lanthanum did not penetrate into the bile canalicular lumen. Holding the view that the term "leakiness" of tight junctions should be defined with reference to the tracer employed, we conclude that in the liver one tight junctional strand is sufficient to prevent the escape of larger bile constituents such as bile acids and that a back diffusion of bile acids over the tight junctional barrier does not play a role in the pathogenesis of the estrogen-induced cholestasis. Interruptions of tight junctions, as described by other authors, are interpreted as a secondary mechanical effect. On the other hand, we consider an increased permeability of the tight junctions to water and small solute molecules as probable; possibly this increased permeability is brought about by alterations in the microfilaments. A model for the pathogenesis of the estrogen-induced intrahepatic cholestasis is proposed.

Cytochalasin B affects the gap and tight junctions of mouse hepatocytes in vivo.

This study investigates the effect of cytochalasin B at a dosage of 0.2 mg per mouse per day for a period of 7 days in an in vivo experiment on mouse liver. Using thin-sectioning and freeze-fracture technique both quantitative and qualitative analysis was made of membranes and cell contacts (gap and tight junctions). Significant alterations of both membranes and junctions were observed. The intercellular space showed vacuolar dilatation in some cases and there were vacuoles observed within the cytoplasm. The microvillar bile canaliculi were dilated. However, no colloidal tracer was observed within the lumen following lanthanum perfusion. With the aid of the freeze-fracture method it was possible to demonstrate that the strands of the tight junctions were highly disorganized. In some cases reduction and in other cases proliferation of tight junctions was observed. Large, proliferative plaques of tight junctions were found both in contact with the tight junctions of the bile canaliculus and ending freely on the plasmalemma. The gap junctions appeared enlarged as well. Their average size increased from 0.42 micron 2 to 0.90 micron 2 (p less than 0.005). The enlargement was also accompanied by an increase in the proportion of the plasma membrane occupied by the junctions: 3.42% in control animals, 10.25% in the livers of mice treated with cytochalasin B. Frequently evaginated and internalized gap junctions were seen in the experimental group. In view of the fact that cytochalasin B, in addition to other effects, also has an effect on the microfilament system of the cell, it may be supposed that microfilaments play a role in maintenance of the orderly structure or in the formation of tight and gap junctions. This remains hypothetical, however, and additional studies are necessary in order to conclusively clarify this issue.

Effect of silybin on hepatic cell membranes after damage by polycyclic aromatic hydrocarbons (PAH).

Young female NMRI mice were given a single intraperitoneal injection of a PAH mixture containing 3 micrometers benzo(a)pyrene in a suspension of 0.5 ml tricapryline. 3 and 4 days later a number of the mice received injections of 100 mg/kg body weight of Silybin in the tail vein. Thin sections of the animals' livers were studied under the electron microscope using the freeze fracture technique. Following intraperitoneal injection of the PAH suspension, and without treatment with Silybin, marked dilatation of the intercellular space and frequent occurrence of intracytoplasmic vacuoles was observed in addition to pronounced alterations in the hepatocytic nuclei and mitochondria. Following the administration of Silybin these alterations were no longer observed, though 50% of the cell membranes exhibited a marked band-like proliferation of the tight junctions. This observation may be interpreted as supporting the postulated membrane-stabilizing effects of Silybin.