Biomedical subjects
Publications and source records attributed to M J Hendrix.
Lymphatic vascular endothelium both on tissue section and in culture exhibits positivity for Factor VIII-associated antigen although staining is generally less intense and more spotty than in comparable blood vascular endothelium. Lymphatic endothelium also exhibits Weibel-Palade bodies. Neither marker, therefore, reliably distinguishes blood vascular endothelium from lymphatic endothelium.
Subcutaneous tumours were induced in castrated golden Syrian hamsters by 7,12 dimethylbenz[a]anthracene (DMBA), an agent known to produce papillomas and carcinomas. The morphological characteristics of the cellular and extracellular constituents of the chemically-induced tumours were indicative of melanoma. Tumours were induced by three injections of DMBA into the jugular vein over a 3 month period. Dermal tumour development within the dorsal integument and groin region ultimately projected into the epidermis and occurred during the 3 month period subsequent to the last DMBA injection. Suspect melanoma tumours were excised and processed for light microscopic (LM) and transmission electron microscopic (TEM) studies. Histochemical staining methods facilitated the characterization of the differentiated tumour components in this hamster melanoma model. The model presented could allow observations from initial melanoma transformation events through advanced stages of metastasis within a window of 7 months.
The variants of the B16 murine melanoma cell line were assayed for their invasive characteristics in the membrane invasion culture system (MICS) and concomitantly tested for their ability to form lung metastases in vivo. Specifically, the B16F1 (low metastatic variant) and the B16F10 (high metastatic variant) murine melanoma cell lines were examined for their ability to invade human amniotic basement membranes (BMs) in vitro and simultaneously examined for lung colony formation in vivo. The B16F1 and B16F10 cell lines both demonstrated similar invasion profiles over 72 h with the total percent invasion through the BMs for both cell lines not exceeding 5.0%. In vivo observations reconfirmed the significant difference in the metastatic capabilities of the 2 variants. These data suggest that tumor cells with differing metastatic propensities can invade an amniotic BM at similar rates, but their survival and metastatic lesion forming capabilities in vivo may vary considerably.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
A 6.5-week-old human embryo with an approximate crown-rump length of 13.2 mm was obtained from a tubal pregnancy. Two hours before surgical removal, the embryo was imaged with real time ultrasound and was noted to have rhythmic cardiac motion. Subsequent to surgical removal, the embryo was dissected free from the placenta and prepared for routine scanning electron microscopic (SEM) studies. Progressive stages of dissection with microsurgical instruments followed by SEM photography elucidated the three-dimensional aspects of embryonic development of many structures, including the lens placode, tongue bud, Rathke's pouch, atrial and ventricular foramina, primitive intestinal loop and undifferentiated external genitalia. Almost certainly, such clear views of dissected structures can contribute to our understanding of human embryonic development.
Explore the source record for details and available documents.
Guinea pig endometrial cells were isolated and maintained in cell culture. Our investigation of these cultures for the presence of epidermal growth factor (EGF) binding activity demonstrated that these cells possess EGF receptors. Upon the addition of EGF to growing cell cultures, an increase in the rate of cell growth resulted along with a higher saturation density. The binding of EGF to these cells is saturable at 40-50 ng/ml., and the receptors demonstrate "down-regulation" in response to hormonal challenge at 37 degrees C.
Collagen-like proteins have been found in the egg shell membranes of the hen. Materials similar to types I and V collagens were detected in each of the two layers of this membrane, the thick outer membrane and the thin inner membrane. Collagen was extracted by acid-pepsin digestion and isolated by differential salt precipitation. Identification of type-specific collagen-like material was established by coelectrophoresis on SDS-polyacrylamide gels using known collagen standards. These bands were susceptible to digestion by bacterial collagenase. From differential staining of the gels it was estimated that the ratio of collagen types I:V was approximately 100:1. Further confirmation of these biochemical results was obtained with immunofluorescence microscopy using type-specific antisera against chicken types I and V collagen with the indirect sandwich technique. Both the inner and outer shell membranes contained the two types of collagen. Within each membrane, the large, coarse 2.5-micron fibers contained predominantly type I collagen-like material, while type V collagen was mainly associated with the delicate narrower fibers of approximately 0.6-micron diameter. These tended to be concentrated in the inner membrane. At the electron microscopic level, both types of fibers were coated with glycoproteins that stained positively with ruthenium red. The deposition of these collagen-like substances by the hen oviduct on to the surface of the developing egg is an additional example of interstitial-type collagen synthesis and secretion by epithelial rather than by mesenchymal cells.
In order to study the invasive characteristics of a patient-derived malignant melanoma cell line (C8146A), interactions of the tumor cells with a basement membrane model were studied in vitro with transmission and scanning electron microscopy. Tumor cells were cultured on amniotic basement membranes for 1, 2, and 3 day intervals and exposed to the anticancer agent Actinomycin D (Act D). Localization of the subcellular sites of drug-cell interaction was performed with protein A sepharose-purified rabbit anti-Act D IgG and immunofluorescence microscopy. The action of the rRNA inhibitor prevented a significant number of C8146A cells from invading the membrane as compared with the untreated cells. These data support the utilization of Act D as an anticancer agent, but pose several considerations regarding the mechanism of anti-inhibitory activity.
In an attempt to diagnose a suspected amelanotic melanoma tumour, we examined a variety of tissue and cell samples from one patient at the ultrastructural level, which consisted of single cell suspensions of tumour cells with and without DOPA treatment, tumour cells after culture in agar with and without DOPA treatment, and single tumour cells hetero-transplanted into a nude mouse. Premalanosomes were not observed in sections of the amelanotic tumour with routine electron microscopy. Osmophilic-dense bodies, suggestive of melanosomes, were noted in the single cells in suspension treated with DOPA and cells grown in agar without DOPA treatment. Definitive premelanosomes, with an identifiable striated matrix, were only observed in cells grown into colonies in agar and treated with DOPA. Positive L-DOPA reaction products were noted in the golgi complex, endoplasmic reticulum closely related to the golgi (GERL), and in vacuoles from cells grown in agar. As controls, Cloudman S91 53.1 melanoma cells were evaluated as single cells in suspension or as colonies after culture in agar, both with and without DOPA treatment. Premelanosomes were always observed in this established melanoma cell line while DOPA-treated cells contained positive L-DOPA reaction products. The overall findings identified the tumour as amelanotic melanoma and indicated that both DOPA treatment and culture in agar were needed for the demonstration of premelanosomes.
Fibronectin (FN) distribution was compared among samples of normal human dermis, hypertrophic scar, keloid, and granulation tissues from deep injuries. Localization was established by use of fibronectin antibodies and the indirect immunofluorescence method. Fresh-frozen tissue was sectioned on a cryostat and examined by epifluorescence. Hypertrophic scar and keloid demonstrated heavy deposition of FN, which conformed to the nodular characteristics of the lesions. Intense localization occurred in granulation tissue over fibroblasts which were stellate and vesiculated, and over small blood vessels. FN-staining was weak in areas over fibroblasts which were more rounded and nonvesiculated. Staining for FN was also minimal over the collagen in normal dermis and the deeper, larger collagen fascicles in the lesions. Fibroblasts cultured from normal dermis, hypertrophic scar, and keloid for 5-6 weeks were intensely stained for FN. Extracellular matrix was heavily positive in cultures from the lesions compared with those from normal dermis.
Explore the source record for details and available documents.
A tumorigenic human cell line was derived from a patient with metastatic melanoma. Cells were treated with adriamycin or actinomycin D in order to assess morphological alterations induced by these anticancer agents. Exposure to 0.01 micrograms/ml adriamycin for one hour caused no observable morphological abnormalities as determined by SEM, while 0.1, 1.0 and 10.0 micrograms/ml concentrations of adriamycin produced surface alterations in the form of blebs, filopodia, microvilli and cell rounding. These alterations may be drug-affected changes of the cell surface or may reflect phases of the cell cycle directed by adriamycin action on the nucleus. Cell morphology appeared normal by SEM for 0.01, 0.1, 1.0 and 10.0 micrograms/ml concentrations of adriamycin when the cells were allowed to recover in drug-free media for 24 hours after initial drug incubation. Concentrations of 0.1, 1.0, and 10.0 micrograms/ml actinomycin D for 24 hours created morphological alterations characterized by cell rounding and long, dendritic-like processes. TEM of colonies treated in soft agar for 24 hours with either 1.0 micrograms/ml adriamycin or 1.0 micrograms/ml actinomycin D showed major morphological effects identified by increased cytoplasmic vacuolization and nuclear disintegration.
Clonogenic assays have been widely adopted for the investigation of hematopoietic and human tumor stem cell biology. Inasmuch as specific, whole colonies need to be analyzed morphologically, we used various methods for fixing and embedding individual colonies in situ that allowed macroscopic, light microscopic (LM), immunofluorescence, and transmission electron microscopic (TEM) evaluation of the intact colony. Melanoma colonies stained with Masson's Trichrome, hematoxylin and eosin (H&E), periodic acid-Schiff, Best's carmine, Page-Green method for inclusion bodies, and Snook's reticulum revealed cellular and extracellular components by LM. Ultrastructural studies revealed specific cellular organelles and extracellular components. Immunofluorescence studies demonstrated cell-surface fibronectin, a high molecular weight, adhesive glycoprotein. Myeloma colonies contained a heterogeneous cell population and produced amyloid fibers that were observed by TEM. Fixation and embedding the colonies in agar for TEM has several advantages over centrifugation methods and other conventional techniques for collecting cells in that (a) an entire specific colony can be studied, (b) there is excellent preservation of the cell and its spatial orientation in the colony, and (c) the extracellular matrix (ECM) of the colony is preserved for immunohistochemical analysis.
Monoclonal and conventional antibodies against collagen types I and II were used for immunofluorescence and immuno-electron microscopic studies of developing chick corneas (5-day-old embryos to adult) and embryonic limb cartilages. Secondary antibodies were labeled with rhodamine or ferritin. We found that the 5-day primary corneal stroma stains uniformly at the light microscope level with both monoclonal and rabbit antibodies to collagen types I and II. At the electron microscope level, the striated fibrils are stained by these antibodies. After invasion by fibroblasts (7-day-old embryos), type I collagen becomes the predominant collagen within most of the stroma, whereas type II becomes progressively localized in subepithelial (Bowman's membrane) and subendothelial (Descemet's membrane) regions. In the adult the only remaining type II reactivity is in Descemet's membrane. In this structure, both the nodes and strands stain positively for type II. In embryonic cartilage, on other hand, type II collagen is organized as nonstriated fibrils. Thus, during avian corneal development, radical changes occur both in the the types of collagens present and in their distribution. In addition, it seems that the same genetic type of collagen can take several morphologic forms, depending on the environment fibrils as well as in the nodes and strands of Descemet's membrane.
Explore the source record for details and available documents.