Isolation and characterisation of a bipotential haematopoietic cell line.
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Biomedical subjects
Publications and source records attributed to T D Allen.
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Cultured hepatocellular carcinoma cells were studied during anchorage-independent growth in semi solid medium (Methocel). The regular occurrence of mitotic figures both at the surface and within the colonies precludes the possibility of such colonies being formed by re-aggregation. The estimated population doubling time in the three-dimensional (3-D) colonies is consistent with those two-dimensional of (2-D) colonies. Structures resembling bile canaliculi were observed between the closely opposed membranes from the well packed adjacent cells. Cell surface and ultrastructural features of the colonies and individual cells are presented and comparisons made with 2-D growth of normal and malignant liver cells in vitro. The formation of 3-D colonies may not only be an assay for transformed cells but also for predicting the type of tumors produced by re-innoculation of the in vitro transformed cells.
The growth potential of normal and transformed rat liver epithelial cells in culture was investigated under various experimental conditions. The control cells failed to form colonies over a base layer of living parent cells, whereas transformed cells formed distinct colonies. Contact with living normal liver cells is required for the growth inhibition of the untransformed cells. Malignant cells capable of producing hepatocellular carcinomas in vivo exhibited signs of invasion in vitro. Altered properties of hepatic cells during the early stages of transformation can be assessed by plating such cell population over a living base layer of normal liver cells.
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Maintenance of myelopoiesis and pluripotential stem cell production for prolonged periods in vitro hitherto has been limited to mouse bone marrow culture. In an effort to adapt the system for use in higher species, particularly in human and non-human primates, studies were undertaken using the prosimian species, Tupaia glis (tree shrew). In a number of experiments the duration of sustained normal hematopoiesis observed in cultures of this species, following a single inoculum of 5 X 10(6)--10(7) bone marrow cells, with or without addition of fresh allogeneic bone marrow exceeded 1 yr. Analysis of suspension cells obtained by weekly demidepopulation of such cultures revealed production of CFU-C, differentiating neutrophils, and basophils at high levels. Direct comparison with murine cultures indicated that in both species a complex series of cellular interactions takes place within an adherent environment of marrow-derived endothelial cells, macrophages, and fat-containing cells. Certain functional and ultrastructural features served to distinguish murine from Tupaia marrow cultures, and the prolonged duration of in vitro hematopoiesis in the latter species could be attributed to a regenerative capacity possessed by its adherent hematopoietic microenvironment. The availability of this primate marrow culture system should facilitate studies of hematopoiesis, viral leukemogenesis, and transplantation biology, which have more direct relevance to man than that provided by the existing murine system.
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Eight patients with congenital microphallus were investigated. Plasma luteinizing hormone, follicle-stimulating hormone, testosterone and androstenedione levels were obtained in all cases. In addition, the response to the administration of human chorionic gonadotropin, luteinizing horomone-releasing hormone and adrenocorticotropic hormone, the assessment of testicular histology by electron microscopy and the measurement of dihydrotestosterone formation by preputial skin were determined in some patients. The results of these studies were compared to similar studies in 6 normal prepubertal boys, 4 boys with bilateral cryptorchidism, 1 male infant with anorchia and 1 adult with hypogonadotropic hypogonadism. The clinical and endocrinological findings in the 8 patients with microphallus can be divided into 2 distinct categories. In 5 patients the disorder is familial, gonadotropin levels are low and there is a normal response to stimulation with chorionic gonadotropin. The data are compatible with the possibility that 3 (possibly 5) of the 8 patients with microphallus have hypogonadotropic hypogonadism. In the other group the cases are sporadic, serum luteinizing hormone and follicle-stimulating hormone levels are elevated and plasma testosterone failed to increase after short-term treatment with chorionic gonadotropin. In these patients a primary testicular disorder appears to be responsible. Experimental and clinical evidence suggests that microphallus results from defective testicular function during the second and third trimesters of pregnancy, either as the result of defective gonadotropin secretion or defective androgen synthesis.
Simultaneous measurements of the intravesical pressure, electromyographic activity of the anal sphincter and the urinary flow rate in 17 children with dysfunctional voiding problems have shown a variety of unusual patterns, each distinct for the particular child but all with the common denominator of failure to coordinate detrusor and sphincter activity. We postulate that these patterns represent persistence of the transitional phase in the development of micturitional control whereby the child learns to prevent involuntary wetting by forceful contraction of the external urethral sphincter.
The clinical and endocrinological features of 15 male subjects with small but normally formed external genitalia are reviewed. Nine of these patients had a simple microphallus alone, while the other 6 patients exhibited associated central nervous system defects. The endocrine data obtained from both groups were the same and were consistent with defective hypothalamic function. Parenteral testosterone seems to be the present treatment of choice but we believe it should be started early in life to take advantage of the greater hyperplastic response of the young.
The clinical and endocrinological features of 15 male subjects with small but normally formed external genitalia are reviewed. Nine of these patients had a simple microphallus alone, while the other 6 patients exhibited associated central nervous system defects. The endocrine data obtained from both groups were the same and were consistent with defective hypothalamic function. Parenteral testosterone seems to be the present treatment of choice but we believe it should be started early in life to take advantage of the greater hyperplastic response of the young.
A liquid culture system is described whereby proliferation of haemopoietic stem cells (CFU-S), production of granulocyte precursor cells (CFU-C), and extensive granulopoiesis can be maintained in vetro for several months. Such cultures consist of adherent and non-adherent populations of cells. The adherent population contains phagocytic mononuclear cells, "epithelial" cells, and "giant fat" cells. The latter appear to be particularly important for stem cell maintenance and furthermore there is a strong tendency for maturing granulocytes to selectively cluster in and around areas of "giant fat" cell aggregations. By "feeding" the cultures at weekly intervals, between 10 to 15 "population doublings" of functionally normal CFU-S regularly occurs. Increased "population doublings" may be obtained by feeding twice weekly. The cultures show initially extensive granulopoiesis followed, in a majority of cases, by an accumulation of blast cells. Eventually both blast cells and granulocytes decline and the cultures contain predominantly phagocytic mononuclear cells. Culturing at 33 degrees C leads to the development of a more profuse growth of adherent cells and these cultures show better maintenance of stem cells and increased cell density. When tested for colony stimulating activity (CSA) the cultures were uniformly negative. Addition of exogenous CSA caused a rapid decline in stem cells, reduced granulopoiesis and an accumulation of phagocytic mononuclear cells.
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The records of 21 children with neuropathic bladder disease are reviewed. The natural history in these cases has been consistent with that of an acquired disorder and the results of urodynamic testing have supported Hinman's contention that the disease is basically a functional one, caused by a discoordination between detrusor contraction and sphincter relaxation. Bladder retraining and specific medication have yielded far better results than were obtained previously by surgical measures alone.
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Simultaneous measurements of the intravesical pressure, electromyographic activity of the anal sphincter and the urinary flow rate in 17 children with dysfunctional voiding problems have shown a variety of unusual patterns, each distinct for the particular child but all with the common denominator of failure to coordinate detrusor and sphincter activity. We postulate that these patterns represent persistence of the transitional phase in the development of micturitional control whereby the child learns to prevent involuntary wetting by forceful contraction of the external urethral sphincter.
For strictures of the posterior urethra lying superior to the urogenital diaphragm, a transpubic approach with resection of the pubic symphysis has been employed. This has permitted primary excision of the scar with reanastomosis of the urethra, thus correcting the defect in a single operation without the introduction of hair-bearing skin and with preservation of the external urethral sphincter. The results in five cases have been most gratifying.