Role of transforming growth factor-beta in the development of the mouse embryo.

Using immunohistochemical methods, we have investigated the role of transforming growth factor-beta (TGF-beta) in the development of the mouse embryo. For detection of TGF-beta in 11-18-d-old embryos, we have used a polyclonal antibody specific for TGF-beta type 1 and the peroxidase-antiperoxidase technique. Staining of TGF-beta is closely associated with mesenchyme per se or with tissues derived from mesenchyme, such as connective tissue, cartilage, and bone. TGF-beta is conspicuous in tissues derived from neural crest mesenchyme, such as the palate, larynx, facial mesenchyme, nasal sinuses, meninges, and teeth. Staining of all of these tissues is greatest during periods of morphogenesis. In many instances, intense staining is seen in mesenchyme when critical interactions with adjacent epithelium occur, as in the development of hair follicles, teeth, and the submandibular gland. Marked staining is also seen when remodeling of mesenchyme or mesoderm occurs, as during formation of digits from limb buds, formation of the palate, and formation of the heart valves. The presence of TGF-beta is often coupled with pronounced angiogenic activity. The histochemical results are discussed in terms of the known biochemical actions of TGF-beta, especially its ability to control both synthesis and degradation of both structural and adhesion molecules of the extracellular matrix.

Fungal attack on rock: solubilization and altered infrared spectra.

Penicillium simplicissimum, isolated from weathering basalt, produced citric acid when grown in a glucose-mineral salts medium with basalt, granite, granodiorite, rhyolite, andesite, peridotite, dunite, or quartzite. After 7 days' growth as much as 31 percent of the silicon, 11 percent of the aluminum, 64 percent of the iron, and 59 percent of the magnesium in some of the rocks were solubilized, and a number of rocks showed altered infrared absorption in the silicon-oxygen vibration region.

Transformation-associated increase of adhesion, cellular fibronectin, and stress fiber development in a liver epithelial cell line.

Cytoskeletal and adhesion characteristics of DL-ethionine (CAS: 67-21-0)-transformed rat liver epithelial cells (ETC) were compared with those of nontumorigenic, untreated cells of the same cell line both at the same passage level as ETC and at an early low passage level. ETC and high-passage-level cells (HPC) showed increased cell spreading and prominent actin stress fibers compared to low-passage-level cells (LPC). The number of adhesion plaques per unit cell area was higher for ETC than for LPC. At confluence, fibronectin expression was high for ETC, moderate for HPC, and low for LPC. The observed increases in cell spreading and in actin and fibronectin expression appeared to be associated with transformation of this cell line rather than being specific responses to ethionine treatment. This conclusion is suggested by the fact that HPC, which display preneoplastic markers, are similar in many respects to ETC.

Central retinal artery occlusion as the initial sign of aortic valve papillary fibroelastoma.

PURPOSE: We describe a central retinal artery occlusion in a young man who was subsequently found to have an aortic valve papillary fibroelastoma. METHODS: Case report with clinical, echocardiographic, and histopathologic observations. RESULTS: The aortic valve papillary fibroelastoma was successfully treated with tumor resection. CONCLUSION: Appropriate investigations in young patients with central retinal artery occlusion should therefore be conducted, including transthoracic and transesophageal echocardiography, to diagnose and treat this tumor, because failure to diagnose this tumor may result in further embolic events, including stroke and sudden death.

Effect of iron and salt on prodigiosin synthesis in Serratia marcescens.

Serratia marcescens wild-types ATCC 264 and Nima grew but did not synthesize prodigiosin in a glycerol-alanine medium containing 10 ng of Fe per ml. Wild-type 264 required the addition of 0.2 mug of Fe per ml for maximal growth and prodigiosin synthesis; Nima required 0.5 mug of Fe per ml. Three percent, but not 0.1%, sea salts inhibited prodigiosin synthesis in a complex medium containing up to 10 mug of Fe per ml. NaCl was the inhibitory sea salt component. The inhibition was not specific for NaCl; equimolar concentrations of Na(2)SO(4), KCl, and K(2)SO(4) also inhibited prodigiosin synthesis. Experiments with strains 264 and Nima and with mutant WF which cannot synthesize 4-methoxy-2-2'-bipyrrole-5-carboxyaldehyde (MBC), the bipyrrole moiety of prodigiosin, and with mutant 9-3-3 which cannot synthesize the monopyrrole moiety 2-methyl-3-amylpyrrole (MAP) showed that both MBC synthesis and the reaction condensing MAP and MBC to form prodigiosin were relatively more sensitive to NaCl inhibition than the MAP-synthesizing step. The capacity of whole cells to condense MAP and MBC was present, but inactive, in cells grown in NaCl; removal of the NaCl from non-proliferating salt-grown cells restored the activity. Other evidence suggests the existence of a common precursor to the MAP- and MBC-synthesizing pathways.

Fungal leaching of titanium from rock.

Penicillium simplicissimum solubilized up to 80% of the titanium in granitic rocks but less than 2% of the titanium in basaltic rocks.

Automated electrical impedance technique for rapid enumeration of fecal coliforms in effluents from sewage treatment plants.

Fecal coliforms growing in a selective lactose-based broth medium at 44.5 degrees C generate a change in the electrical impedance of the culture relative to a sterile control when populations reach 10(6) to 10(7) per ml. The ratio of these changes was measured automatically, and the data were processed by computer. A linear relation was found between the log10 of the number of fecal coliforms in an inoculum and the time required for an electrical impedance ratio signal to be detected. Pure culture inocula consisting of 100 fecal coliforms in log phase or stationary phase were detected in 6.5 and 7.7 h, respectively. Standard curves of log10 fecal coliforms in wastewater inocula versus detection time, based on samples collected at a sewage treatment plant over a 4-month period, were found to vary from one another with time. Nevertheless, detection times were rapid and ranged from 5.8 to 7.9 h for 200 fecal coliforms to 8.7 to 11.4 h for 1 fecal coliform. Variations in detection times for a given number of fecal coliforms were also found among sewage treatment plants. A strategy is proposed which takes these variations into account and allows for rapid, automated enumeration of fecal coliforms in wastewater by the electrical impedance ratio technique.

Rapid, single-step most-probable-number method for enumerating fecal coliforms in effluents from sewage treatment plants.

A single-step most-probable-number method for enumerating fecal coliforms in sewage treatment plant effluents is described. The method requires the use of only one lactose-based medium and a single incubation temperature of 44.5 degrees C, and it can be completed in 18 h or less, as compared with up to 72 h for the standard most-probable-number method. The appearance of growth is the sole criterion used for designating positives, which can be determined either by increases in the electrical impedance ratio of inoculated medium, as compared to an uninoculated control using a Bactometer model 32, or by visual examination of inoculated medium for turbidity. In trials with 53 samples of unchlorinated sewage treatment plant effluent, fecal coliform counts by the single-step most-probable-number method, throughout a range of less than 10 to almost 10(7) fecal coliforms per 100 ml of effluent, were in excellent agreement with counts abtained by the standard most-probable-number procedure. Similar agreement was obtained in comparative trials with 31 chlorinated effluent samples from two sewage treatment plants. Overall, 87% of 452 positives were confirmed as containing fecal coliforms. The applicability of the single-step most-probable-number method to automated sewage treatment plant operations is discussed.

Gas-tight flask for the concurrent measurement of gas metabolism and growth in methane-oxidizing bacteria.

A flask is described for use in conjunction with a gas chromatograph and a spectrophotometer to follow methane and oxygen uptake, carbon dioxide production, and cell growth concurrently in methane-oxidizing cultures.

Microbial metabolism and dynamic changes in the electrical conductivity of soil solutions: a method for detecting extraterrestrial life.

The addition of 0.5% glucose solutions to 12 different air-dried soils always resulted in increased electrical conductivity and water-soluble Ca and Mg in the soil solutions. The kinetics and magnitude of these changes for at least two and usually all three of these parameters over a 14-day period were clearly distinguishable from the changes in heat-sterilized controls or unsterilized controls without added glucose. In general, maximal values were achieved more rapidly under aerobic than anaerobic incubation. Some soils (less than half) also showed significant increases in water-soluble Na or K when compared with the controls. The 12 different soils studied represented four general soil groups: I, leached acid upland soils; II, saline alkaline soils; III, nonsaline neutral soils; and IV, high organic soils. Viable counts ranged from 10(4) to 10(7) per cm(3) of air-dried soil. Glucose metabolism by the indigenous soil microbiota was always accompanied by a significant decrease in the pH of soil solutions, but not necessarily by an increase in the viable count. The feasibility of using electrical conductivity and water-soluble Ca and Mg measurements to detect metabolic activity, either alone or in conjunction with other life detection techniques, is discussed.

Characterization of rat liver cells transformed in culture by DL-ethionine.

A rat liver-derived epithelial cell line transformed with DL-ethionine and the corresponding control cell line were characterized according to morphological and cytochemical criteria to establish their origin from liver epithelium and to identify cellular changes due to transformation by DL-ethionine. The presence of intermediate junctions confirms the epithelial nature; glycogen accumulation and glucose-6-phosphatase activity confirm the hepatic origin of the cells. Persistent alterations resulting from ethionine transformation were variations in cell shape and size, focal multilayered growth, an increase in the nucleolar:nuclear ratio, and a reduction in the number of cells displaying a primary cilium. Hyperplasia of the inner nuclear membrane, elongation and branching of mitochondria, and a reduction in the length and frequency of cell junctions were also characteristic of the transformed cells.

Video endoscopy for laser photoresection in tracheobronchial pathology: some considerations after 9 years experience with 2105 treatments.

Between 1984 and 1993 we performed 2105 laser treatments in 1210 patients: 52% of treatments were done for malignant pathology, 45% for benign tracheal stenoses and 3% were in a miscellaneous group. The procedure was carried out with a rigid bronchoscope under general anaesthesia. In patients with malignant tumors, it is a good palliative treatment-safe, well tolerated and with immediate results; it can be repeated as many times as needed with and is well accepted by the patient. In patients without tumors, this method avoids emergency tracheotomies. The long term results are now under evaluation.

Effects of retinoid deficiency on the development of the heart and vascular system of the quail embryo.

The regulatory role of retinoids in growth and differentiation has been examined in vitro and in vivo by light and scanning electron microscopy using retinoid-deficient and control quail embryos between the 5-15 somite stage, as well as 2- and 2.5-day-old embryos. Fertile, retinoid-deficient eggs were obtained from flocks of quail maintained on a retinoid- and carotenoid-deficient diet, supplemented only with small amounts of retinoic acid methyl ester as described by Thompson et al. 1969. As described previously, retinoid deprivation during embryonal development causes abnormalities in organs of epithelial and mesenchymal origin, most dramatically preventing the formation of the extraembryonal circulatory system in the avian embryo. Our in vivo studies show that the basis for the latter defect is the failure of the primitive heart tubes to open at their posterior end, thus preventing the formation of omphalomesenteric veins normally connecting the embryonal with the extraembryonal circulatory system. Early manifestation of the retinoid-deficient defect may result also in formation of a cardia bifida, late manifestation in development of a single dilated ventricle. In contrast, the extraembryonal vascular system of blood islands is well developed. Heart function as shown by the rate of heart beat is reduced in deficient embryos. Our in vitro studies demonstrate similar defects in the development of the circulatory system by culture of normal 24-h embryos on retinoid-deficient agar medium; conversely, normal development is observed upon culture of retinoid-deficient embryos on retinoid-containing agar medium.

Colocalization of TGF-beta 1 and collagen I and III, fibronectin and glycosaminoglycans during lung branching morphogenesis.

The possible in vivo role of TGF-beta 1 in regulating various proteins of the extracellular matrix, including fibronectin, collagen I and III, and glycosaminoglycans, was examined by immunohistochemical methods during critical stages of lung morphogenesis in the 11- to 18-day-old mouse embryo. Sections of Bouin-fixed, paraffin-embedded whole embryos were exposed to polyclonal antibodies specific to synthetic peptides present in the precursor part of TGF-beta 1 (pro-TGF-beta 1), in the processed TGF-beta 1 (antibody CC), collagen I and III, fibronectin, followed by the PAP or ABC technique to visualize the location of the antibody. GAG were stained with Alcian Blue 8GX. Our results indicate colocalization of TGF-beta 1 expression and that of matrix proteins in the developing lung when branching morphogenesis (cleft formation) and tissue stabilization occur. The presence of TGF-beta 1 at the epithelial-mesenchymal interfaces of stalks and clefts at a time when matrix proteins can first be visualized in these areas, suggests a direct participation of the growth factor in the development of the basic architecture of the lung.

Localization of transforming growth factor-beta 1 in mitochondria of murine heart and liver.

Using both electron microscopic immunohistochemistry and cell fractionation techniques, we show that transforming growth factor-beta 1 (TGF-beta 1) is found in mitochondria of rat and mouse cardiac myocytes and rat hepatocytes. Four different polyclonal antibodies, raised against various epitopes encompassing the mature portion of the TGF-beta 1 molecule as well as the pro-region of its precursor, were used for the electron microscopy studies. The localization of TGF-beta 1 in mitochondria was confirmed by detection of the native peptide in mitochondria isolated from rat heart and liver; the majority of native TGF-beta 1 found in liver homogenates was recovered in highly pure mitochondrial fractions. The functional role of TGF-beta in the mitochondrion is unknown at present.

Transplacental mutagenicity of cisplatin: H-ras codon 12 and 13 mutations in skin tumors of SENCAR mice.

Cisplatin is an anticancer agent sometimes used in pregnant women. It is also a potent initiator of skin tumors in mice when administered transplacentally. For characterization of the transplacental mutagenicity of cisplatin, tumors initiated in fetal skin by cisplatin or 7,12-dimethylbenz[a]anthracene (DMBA) and promoted by postnatal 12-O-tetradecanoyl-phorbol-13-acetate (TPA) were analyzed for H-ras mutations by 'cold' single-strand conformation polymorphism analysis and direct sequencing. The expected high incidence of exon II codon 61 mutations (20/20) was found in transplacental DMBA-initiated tumors, with no exon I change. By contrast, 6/10 cisplatin tumors had seven mutations in codons 12 or 13 of exon I, all at GpG dinucleotides. Four of these were unique codon 13 GGC --> GTC changes, significantly different from the DMBA group and from historical TPA-only controls. The activation of codons 12 and 13 by cisplatin is in accord with the known in vitro preference of cisplatin for GpG sites for intrastrand cross-linking adduct formation. These results provide the first evidence that cisplatin can act transplacentally to cause specific mutations in fetal skin that are not seen in skin tumors caused by treatment of adult skin with this agent. This is evidence for unique molecular fetal carcinogenic pathways and underscores concern about human fetal risk due to maternal cisplatin treatment.

Transforming growth factor-beta 1 specifically localizes in elastin during synovial inflammation: an immunoelectron microscopic study.

We report here that extracellular TGF-beta 1 is associated exclusively with microfibrils of elastin which are present in the extracellular matrix of the inflamed articular joint of the rat. Inflammation was initiated by bacterial cell walls localized in the synovium following intraperitoneal injection of the bacterial components. This synovitis is associated with both destruction of connective tissue components and matrix deposition. The growth factor was localized by using a polyclonal antibody raised to a synthetic peptide corresponding to amino terminal 30 amino acids of TGF-beta 1 in conjunction with a gold-labeled secondary antibody. The results suggest a close association of TGF-beta 1 with proteoglycans which are known to be a major component of the microfibrils in elastin. Proteoglycan-mediated binding and concentration of TGF-beta 1 in specific areas of the extracellular matrix may constitute a mechanism whereby the growth factor could be targeted to specific sites of action.